United States Office of Water EPA 822-P-14-001
Environmental Protection 4304T May 2014
Agency
External Peer Review Draft
Aquatic Life Ambient Water Quality
Criterion for
Selenium - Freshwater
\2014 *S
U.S. Environmental Protection Agency
Office of Water
Office of Science and Technology
Washington, D.C.
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Table of Contents
Table of Contents ii
List of Tables iv
List of Figures v
Notices vii
Foreword viii
Acknowledgements ix
Acronyms xi
1 Executive Summary 1
2 Introduction and Background 6
2.1 History of the EPA Selenium AWQC for Aquatic Life 6
3 Problem Formulation 9
3.1 Overview of Selenium Sources and Occurrence 9
3.2 Environmental Fate and Transport of Selenium in the Aquatic Environment 13
3.2.1 Selenium Species in Aquatic Systems 13
3.2.2 Bioaccumulation of Selenium in Aquatic Systems 15
3.3 Mode of Action and Toxicity of Selenium 17
3.4 Narrow Margin between Sufficiency and Toxicity of Selenium 20
3.5 Interactions with Mercury 21
3.6 AssessmentEndpoints 21
3.7 Measures of Effect 24
3.7.1 Fish Tissue 25
3.7.2 Water 29
3.7.3 Summary of Assessment Endpoints and Measures of Effect 30
3.7.4 Conceptual Model of Selenium Effects on Aquatic Life 31
3.8 Analysis Plan 32
3.8.1 Analysis Plan for Derivation of the Chronic Fish Tissue-Based Criterion Element. 32
3.8.2 Analysis Plan for Derivation of Duration of Fish Tissue Criterion Elements 34
3.8.3 Analysis Plan for Derivation of Chronic Water-based Criterion Element 34
3.8.4 Analysis Plan for Intermittent-Exposure Water-based Criterion Element
Derivation 38
4 Effects Analysis for Freshwater Aquatic Organisms 39
4.1 Chronic Tissue-Based Selenium Criterion Element Concentration 39
4.1.1 Acceptable Studies of Reproductive Effects 40
4.1.2 Summary of Acceptable Studies of Fish Reproductive Effects 51
4.1.3 Invertebrate Chronic Effects 54
4.1.4 Summary of Relevant Invertebrate Tests 56
4.1.5 Derivation of Tissue Criterion Element Concentrations 57
4.2 Chronic Water Column-based Selenium Criterion Element 62
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4.2.1 Translation from Fish Tissue Concentration to Water-Column Concentration 62
4.2.2 Equation Parameters 71
4.2.3 Food-Web Models 80
4.2.4 Classifying Categories of Aquatic Systems 81
4.2.5 Deriving Protective Water Column Concentrations for Lentic and Lotic System
Categories 85
4.2.6 Derivation of Averaging Period for Chronic Water Criterion Element 91
4.3 Intermittent-Exposure Water Criterion Element: Derivation from the Chronic Water
Criterion Element 92
5 National Criterion for Selenium in Fresh Waters 96
6 Site-specific Criteria 100
7 Effects Characterization 102
7.1 Fish 102
7.1.1 Principles for Using Studies for which ECioS Cannot Be Calculated 102
7.1.2 Reproductive Effects in Catfish (Ictaluridae) 103
7.1.3 Reproductive Studies Not Used in the Numeric Criterion Derivation 105
7.1.4 Salmo GMCV: EPA Re-analysis of a Key Study Used in Criterion Derivation 107
7.1.5 Influence of Curve-fitting on Calculation ofLepomis GMCV Ill
7.1.6 Impact of Number of Tested Species on Criterion Derivation 112
7.1.7 Conversions between Concentrations in Different Tissues 112
7.1.8 Studies of Non-Reproductive Effects 113
7.1.9 Comparison of Fish Chronic Reproductive Effects and Chronic Non-Reproductive
Effects 130
7.2 Water 133
7.2.1 Validation of Translation Equation for Developing Water Column
Concentrations 133
7.2.2 Evaluating the Protectiveness of the Final Water-Column Criterion Values 135
7.2.3 Uncertainty in Bioaccumulation of Total Dissolved Selenium 138
7.3 Protection of Threatened or Endangered Species 139
7.4 Aquatic-Dependent Wildlife is Beyond the Scope of this Aquatic Criteria Derivation.
140
8 References 141
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List of Tables
Summary of the External Peer Review Draft Freshwater Selenium Ambient Chronic Water
Quality Criterion for Protection of Aquatic Life (See Section 5 for the complete
criterion statement.) 4
Table 1. Predominant chemical forms of selenium in discharges associated with different
activities and industries 13
Table 3. 1985 Guidelines Minimum Data Requirements Summary Table Reflecting the
Number of Species and Genus Level Mean Values Represented in the Chronic Toxicity
Dataset for Selenium in Freshwater 28
Table 4. Summary of Assessment Endpoints and Measures of Effect Used in Criteria
Derivation for Selenium 30
Table 5. Maternal Transfer Reproductive Toxicity Studies 52
Table 6a. Ranked Genus Mean Chronic Values for Fish Reproductive Effects 54
Table 6b. Ranked Invertebrate Whole-Body Chronic Values with Translation to Expected
Accompanying Fish Egg-Ovary Concentrations 57
Table 6c. Four lowest Genus Mean Chronic Values for Fish Reproductive Effects 58
Table 7a. Tested Reproductive-Effect Egg-Ovary (EO) Concentrations Converted to Whole-
Body (WB) Concentrations 59
Table 7b. The lowest four reproductive-effect whole-body GMCVs 60
Table 8a. Tested Reproductive-Effect Egg-Ovary (EO) Concentrations Converted to Muscle
(M) Concentrations 61
Table 8b. The lowest four reproductive-effect fish muscle GMCVs 61
Table 9. EPA-derived Trophic Transfer Function (TTF) Values for Freshwater Aquatic
Invertebrates 76
Table 10. EPA-Derived Trophic Transfer Function (TTF) Values for Freshwater Fish 77
Table 11. EPA-Derived Egg-Ovary to Whole-Body Conversion Factor (CF) Values 79
Table 12. Site-Specific Data for the 132 Species-Site Combinations and Translation of the
Egg-Ovary Criterion Concentraiton Element to a Water Column Concentration.51 85
Table 13. Summary of water column criterion element concentration values translated from
the egg-ovary criterion element 91
Table 14. Representative Values of the Intermittent Water Criterion Concentration Element 95
Table 15. 2014 External Peer Review Draft Freshwater Selenium Ambient Water Quality
Chronic Criterion for Aquatic Life 97
Table 16. Correlation matrix (values of r) for Ictaluridae and Centrarchidae abundance and
for selenium food chain contamination for the Hyco Reservoir data reported by
Crutchfield (2000) 104
Table 17. Freshwater Chronic Values from Acceptable Tests - Non-Reproductive Endpoints
(Parental Females Not Exposed.) 125
Table 18. Comparison of criterion attainment using the water column and egg-ovary
concentration values in lentic aquatic systems 137
Table 19. Comparison of selenium criterion attainment using the water column and
egg-ovary concentration values in lotic aquatic systems 137
Table 20. Binary classification statistics for lentic and lotic aquatic systems 138
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List of Figures
Figure 1. Map indicating deposits of selenium in mining regions 11
Figure 2. Areas of western U.S. susceptible to selenium contamination (gray) and where
agricultural land is irrigated (green) 12
Figure 3. Diagram of selenium partitioning, bioaccumulation, and effects in the aquatic
environment 31
Figure 4. Conceptual model for translating the selenium egg-ovary concentration to a water-
column concentration 38
Figure 5. Distribution of (a) reproductive-effect GMCVs for fish measured as egg-ovary
concentrations (diamond markers), (b) the reproductive-effect value for mosquito fish
(square marker), a live-bearer measured as adult whole-body but translated to an
equivalent egg-ovary concentration using the median conversion factor 1.71, and (c)
invertebrate effect concentrations (triangle markers) measured as whole-body but
translated to the equivalent fish egg-ovary concentrations expected in an accompanying
fish assemblage, through the median trophic transfer factor of 1.27 from Table 10 and
the median egg-ovary conversion factor of 1.71 from Table 11 58
Figure 6. Distribution of (a) reproductive-effect GMCVs for fish, measured as egg-ovary
concentrations but converted to whole-body concentrations as shown in Table 7, (b) the
reproductive-effect value for mosquito fish, a live-bearer already measured as adult
whole-body, and (c) invertebrate effect concentrations measured as whole-body and
translated to equivalent fish whole-body concentrations through the median trophic
transfer factor of 1.27 from Appendix B 60
Figure 7. Example aquatic system scenarios and the derivation of the equation parameter
TTFcomposite 68
Figure 8. Effect of relative sample collection time on correlation coefficients of selenium
measurements in particulate material, and invertebrate and fish tissue 73
Figure 9. Distribution of EF values for 69 aquatic sites derived from published studies and
grouped into 4 categories 83
Figure 10. Distribution of EF values for the same 69 aquatic systems as shown in Figure 9
grouped into 2 categories (lentic and lotic) 84
Figure 11. Probability distribution of the water-column concentrations translated from the
egg-ovary criterion at lentic and lotic aquatic sites 89
Figure 12. Illustration of intermittent spike exposure occurring for a certain percentage of
time (e.g., 10%) over a 30-day period, and background exposure occurring for the
remaining percentage of time (e.g., 90%) 93
Figure 13. Concentration-response relationships of brown trout deformities (a-b), survival
(c-d), and deformities+survival (e-f) in response to selenium concentrations in eggs 110
Figure 14. Fitting the Hermanutz et al. (1992, 1996) data to yield (a) the 12.68 mg Se/kg
ECio (dotted line) with TRAP measuring error vertically, and (b) a possible alternative
18.40 mg Se/kg ECio (dashed line), reducing horizontal error by running TRAP after
combining two points averaging 49.85% normal (absence of edema) into one point at
their geometric mean exposure of 24.56 mg Se/kg, and combining the two points
averaging 19.3% normal into one at their geometric mean exposure, 24.45 mg Se/kg Ill
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Figure 15. Distribution of (a) non-reproductive effect genus mean values for fish measured
as whole-body concentrations or muscle concentrations converted to whole body, and
(b) invertebrate effect concentrations converted to equivalent fish whole-body
concentrations using a trophic transfer factor of 1.27, both compared to the
reproductive effect whole-body FCV 131
Figure 16. Scatter plot of predicted versus measured concentrations of selenium in fish.
Dashed line shows unity y = x line 135
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Notices
This document has been reviewed by the Health and Ecological Criteria Division, Office
of Science and Technology, U.S. Environmental Protection Agency, and is approved for
publication.
When published in final form, this document will provide guidance to States and Tribes
authorized to establish water quality standards under the Clean Water Act (CWA), to protect
aquatic life from toxic effects of selenium. Under the CWA, States and Tribes are to establish
water quality criteria to protect designated uses. State and tribal decision makers retain the
discretion to adopt approaches on a case-by-case basis that differ from this guidance when
appropriate. While this document contains EPA's draft scientific recommendations regarding
ambient concentrations of selenium that protect aquatic life, it does not substitute for the CWA
or EPA's regulations; nor is it a regulation itself. Thus, it cannot impose legally binding
requirements on EPA, States, Tribes, or the regulated community, and might not apply to a
particular situation based upon the circumstances. EPA may change this draft document in the
future. This document has been approved for publication by the Office of Science and
Technology, Office of Water, U.S. Environmental Protection Agency.
Mention of trade names or commercial products does not constitute endorsement or
recommendation for use. This document can be downloaded from:
http://www.epa.gov/waterscience/criteria/aqlife.html
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Foreword
Section 304(a)(l) of the Clean Water Act of 1977 (P.L. 95-217) requires the
Administrator of the Environmental Protection Agency to publish water quality criteria that
accurately reflect the latest scientific knowledge on the kind and extent of all identifiable effects
on health and welfare that might be expected from the presence of pollutants in any body of
water, including ground water. This document is a new proposal of an ambient water quality
criterion (AWQC) for the protection of aquatic life based upon consideration of all available
information relating to effects of selenium on aquatic organisms and comments received from
U.S. EPA staff.
The term "water quality criteria" is used in two sections of the Clean Water Act, section
304(a)(l) and section 303(c)(2). The term has a different program impact in each section. In
section 304, the term represents a non-regulatory, scientific assessment of ecological effects. The
criterion presented in this document is such a scientific assessment. If water quality criteria
associated with specific stream uses are adopted by a state as water quality standards under
section 303, they become enforceable maximum acceptable pollutant concentrations in ambient
waters within that state. Water quality criteria adopted in state water quality standards could have
the same numerical values as criteria developed under section 304. However, in many situations
states might want to adjust water quality criteria developed under section 304 to reflect local
environmental conditions and human exposure patterns. Alternatively, states may use different
data and assumptions than EPA in deriving numeric criteria that are scientifically defensible and
protective of designated uses. It is not until their adoption as part of state water quality standards
that criteria become regulatory. Guidelines to assist the states and Indian tribes in modifying the
criteria presented in this document are contained in the Water Quality Standards Handbook (U.S.
EPA 1994). This handbook and additional guidance on the development of water quality
standards and other water-related programs of this agency have been developed by the Office of
Water.
This draft document is guidance only. It does not establish or affect legal rights or
obligations. It does not establish a binding norm and cannot be finally determinative of the issues
addressed. Agency decisions in any particular situation will be made by applying the Clean
Water Act and EPA regulations on the basis of specific facts presented and scientific information
then available.
7
Elizabeth Southerland
Director
Office of Science and Technology
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Acknowledgements
Technical Analysis Leads
Charles Delos, U.S. EPA, Office of Water, Office of Science and Technology, Health and
Ecological Criteria Division, Washington, DC
Gary Russo, U.S. EPA, Office of Water, Office of Science and Technology, Standards and
Health Protection Division, Washington, DC
russo.gary@epa.gov
Technical Contributor and Primary Contact Person
Joseph Beaman, U.S. EPA, Office of Water, Office of Science and Technology, Health and
Ecological Criteria Division, Washington, DC
beaman.joe@epa.gov
V ^f
Editors
Elizabeth Behl and Kathryn Gallagher, U.S. EPA, Office of Water, Office of Science and
Technology, Health and Ecological Criteria Division, Washington, DC
Reviewers (2010-2011)
Lisa Huff, U.S. EPA, Office of Water, Office of Science and Technology, Washington, DC
Joseph Beaman, U.S. EPA, Office of Water, Office of Science and Technology, Washington, DC
Intra-Agency Panel Peer Reviewers (2014)
Dale Hoff and Charles Stephan, U.S. EPA, Office of Research and Development, Mid-Continent
Ecology Division, Duluth, MN
Cindy Roberts, U.S. EPA, Office of Research and Development, Office of Science Policy,
Washington, DC
Jim Lazorchak, U.S. EPA, Office of Research and Development, National Exposure Research
Laboratory, Cincinnati, OH
Keith Sappington and Nancy Andrews, U.S. EPA, Office of Chemical Safety and Pollution
Prevention, Office of Pesticide Programs, Arlington, VA
Jeff Gallagher, U.S. EPA, Office of Chemical Safety and Pollution Prevention, Office of
Pollution Prevention and Toxics, Washington, DC
Laura Phillips and Scott Wilson, U.S. EPA, Office of Water, Office of Wastewater Management,
Washington, DC
Rosaura Conde, Ruth Chemerys, and Eric Monschein, U.S. EPA, Office of Water, Office of
Wetlands, Oceans, and Watersheds, Washington, DC
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Lars Wilcut and Jim Keating, U.S. EPA, Office of Water, Office of Science and Technology,
Washington, DC
Cheryl Atkinson and Frank Borsuk, U.S. EPA Region 3, Philadelphia, PA, and Wheeling, WV
Joel Hansel, U.S. EPA Region 4, Atlanta, GA
Angela Vincent, U.S. EPA Region 5, Chicago, IL
Lareina Guenzel, U.S. EPA Region 8, Denver, CO
Diane Fleck, Eugeina McNaughton, and Daniel Oros, U.S. EPA Region 10, San Francisco, CA
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Acronyms
AE Assimilation Efficiency
AWQC Ambient Water Quality Criteria
BAF Bioaccumulation Factor
CCC Criterion Continuous Concentration
CF Conversion Factor
CV Chronic Value (expressed in this document as an EC20 or MATC)
CWA Clean Water Act
ECx Effect Concentration at X Percent Effect Level
EF Enrichment Factor
EPA Environmental Protection Agency
EO Egg Ovary
FCV Final Chronic Value
GMCV Genus Mean Chronic Value
IR Ingestion Rate
ke Rate of selenium loss
ku Rate of selenium uptake
LOEC Lowest Observed Effect Concentration
M Muscle
MATC Maximum Acceptable Toxicant Concentration (expressed mathematically as the
geometric mean of the NOEC and LOEC)
MDR Minimum Data Recommendations or Requirements
NPDES National Pollutant Discharge Elimination System
NOEC No Observed Effect Concentration
SMCV Species Mean Chronic Value
TMDL Total Maximum Daily Load
TRAP EPA's Statistical Program: Toxicity Relationship Analysis Program (Version
1.21)
TTF Trophic Transfer Factor
WB Whole body
WQBLS Water Quality-based Effluent Limitations
WQC Water Quality Criteria
WQS Water Quality Standards
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1 Executive Summary
This document sets forth the basis for and derivation of the Clean Water Act, Section
304(a) water quality criterion for protecting aquatic life from harmful effects of selenium, a
naturally occurring chemical element that is nutritionally essential in small amounts, but toxic at
higher concentrations. This assessment provides a critical review of all data quantifying the
toxicity of selenium to aquatic organisms, and provides a basis for a criterion that will assure
protection of population assemblages offish, amphibians, aquatic invertebrates, and plants.
Although selenium may cause acute toxicity, the most deleterious effect on aquatic
organisms is due to its bioaccumulative properties. Organisms in aquatic environments exposed
to selenium accumulate it primarily through their diet, and not directly through water (Chapman
et al. 2010). It is also recognized that selenium toxicity occurs primarily through transfer to the
eggs and subsequent reproductive effects. Consequently, in harmony with the recommendations
of expert panels (U.S. EPA 1998, Chapman et al. 2010) and with peer review and public
comments on the U.S. EPA (2004) draft, the Agency has developed a chronic criterion reflective
of the effects of selenium concentrations in the reproductive tissues offish species. The 2014
freshwater criterion for selenium is composed of four parts, or elements. The recommended
elements are for (1) a fish egg/ovary element; (2) a fish whole-body and/or muscle element; (3) a
water-column chronic element for lentic or lotic waterbody types; and (4) a water-column
intermittent element for lentic or lotic waterbody types to account for potential chronic effects
from repeated, short-term exposures to this bioaccumulative pollutant. All criterion elements are
intended to protect aquatic life from the chronic effects of exposure to total selenium. The
criterion is not intended to address concerns about selenium toxicity to aquatic-dependent
wildlife such as aquatic bird species. Because the factors that control the bioaccumulation of
selenium vary from location to location, a site-specific criterion for the protection of aquatic life
can be developed as needed (Appendix I), when establishing allowable concentrations in water
or resident fish.
The toxicity studies relevant to the derivation of the two fish tissue selenium criterion
elements involve (a) extended-duration dietary exposure, and (b) measurement of total selenium
in the tissue of the target organism. Selenium either in fish whole-body or in muscle is usually
measured in non-reproductive studies - tests measuring the survival and growth of organisms,
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for example, juvenile fish, exposed to elevated concentrations of selenium in their diet. Selenium
in eggs or ovaries is measured in reproductive studies - tests measuring the health of the
offspring of adult female fish exposed to elevated dietary selenium either in the lab or in the
field. Selenium accumulation in the eggs of the exposed adult female prior to spawning has been
shown to yield a statistically significant occurrence of deformities and reduced survival of the
offspring.
The outcome of assessing both reproductive and non-reproductive studies under
laboratory and field conditions ultimately led EPA to the conclusion, consistent with expert
consensus (Chapman et al. 2009, 2010), that reproductive effects, linked to egg-ovary selenium
concentrations, are of greater ecological concern and provide a more reliable basis for the
criterion than non-reproductive (e.g., survivorship, growth) endpoints. Reproductive effects have
been linked to observed reductions in the populations of sensitive fish species in waterbodies
having elevated concentrations of selenium (Young et al. 2010). Applying the species sensitivity
distribution concepts from the U.S. EPA (1985) Guidelines to the available data, the draft egg-
ovary criterion element is 15.2 milligrams selenium per kilogram dry weight (mg Se/kg dw),
based on 19 reproductive studies with nine fish genera.
The egg-ovary criterion element is expected to protect aquatic invertebrates and plants (in
addition to fish) because field experience, corroborated by the available laboratory toxicity
studies, indicates that these taxa are less sensitive than fish, based on the available data.
Mechanism of action information suggests that amphibians would have sensitivity comparable to
fish; however, EPA is not aware of existing amphibian studies of sufficient quality that can be
used for selenium criteria derivation.
EPA is recommending one criterion with two fish tissue-based and two water-based
criterion elements to protect against adverse effects of selenium on aquatic life. All four of these
elements are based on the same assessment endpoint, reproductive effects in freshwater fish.
EPA derived the values for the water-based criterion elements from the egg-ovary element by
assessing food-chain bioaccumulation at representative field sites across the continental United
States. EPA also used field observations to assess selenium enrichment in algae, detritus, and
sediment relative to water, and used field observations and laboratory data to quantify trophic
transfer functions from algae, detritus, and sediment into invertebrates, and from such prey into
fish. EPA also used field observations to assess selenium partitioning between the whole-body
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and the eggs or ovaries offish species. EPA tested and validated this approach using field data
on existing conditions at 132 species-site combinations with a range of bioaccumulation potential
due to different hydrologic and biotic characteristics. Two different, but related elements were
developed for the water column portion of the selenium chronic criterion; a monthly average
element and an element for intermittent exposures. Both water column elements are further
refined into two values; one for lentic waters (e.g., lakes and impoundments) and one for lotic
waters (e.g., rivers and streams). The lentic and lotic water values reflect the apparent difference
in enrichment from water into algae, detritus, and sediment in these two types of aquatic systems.
These lentic and lotic water concentrations were calculated based on the fish egg-ovary criterion
element (15.2 mg Se/kg dw). This egg-ovary value was converted to estimated fish whole-body
concentrations and fish muscle concentrations (for each species). Corresponding selenium
concentrations were then predicted at each trophic level downward through the food chain, until
arriving at predicted allowable water concentrations in lentic and lotic systems for the 132
species-site combinations. The 20* percentile of the distribution of predicted allowable site
median selenium concentrations in water yields the national monthly water criterion element
concentrations of 1.3 |ig/L in lentic waters and 4.8 |ig/L in lotic waters.
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Summary of the External Peer Review Draft Freshwater Selenium Ambient
Chronic Water Quality Criterion for Protection of Aquatic Life (See Section 5 for the
complete criterion statement.)
Media
Type
Criterion
Element
Magnitude
Duration
Frequency
Fish Tissue
Egg/Ovary1
15.2mg/kg
Instantaneous
measurement5
Never to be
exceeded
Fish Whole
Body or
Muscle2
8.1 mg/kg
whole body
or
11. 8 mg/kg
muscle
(skinless,
boneless filet)
Instantaneous
measurement5
Never to be
exceeded
Water Column3
Monthly
Average
Exposure
1.3 |ig/Lin
lentic aquatic
systems
4.8 ng/Linlotic
aquatic systems
30 days
Not more than
once in three
years on
average
Intermittent Exposure4
WQCint =
WQC30_day — Cbkgrnd(\ — f int)
1 int
Number of days/month with
an elevated concentration
Not more than once in three
years on average
Overrides any whole-body, muscle, or water column elements when fish egg/ovary
concentrations are measured.
r\
Overrides any water column element when both fish tissue and water concentrations are
measured.
3 Water column values are based on dissolved total selenium in water.
4 Where WQCso-dayis the water column monthly element, for either a lentic or lotic system, as
appropriate. Cbkgmd is the average background selenium concentration, and f;nt is the fraction of
any 30-day period during which elevated selenium concentrations occur, with f;nt assigned a
value >0.033 (corresponding to 1 day).
5 Instantaneous measurement. Fish tissue data provide point measurements that reflect integrative
accumulation of selenium over time and space in the fish at a given site. Selenium concentrations
in fish tissue are expected to change only gradually over time in response to environmental
fluctuations.
EPA recommends that states and tribes adopt into their water quality standards a
selenium criterion that includes all four elements, expressing the four elements as a single
criterion composed of multiple parts, in a manner that explicitly affirms the primacy of the
whole-body or muscle element over the water-column elements, and the egg-ovary element over
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any other element. Adoption of the fish whole-body or muscle elements into water quality
standards ensures the protection of aquatic life when fish egg or ovary tissue measurements are
not available, and adoption of the water-column elements ensures protection when neither fish
egg-ovary nor fish whole-body or muscle tissue measurements are available. (See Section 5.)
EPA recommends that when states implement the criteria for selenium under the National
Pollutant Discharge Elimination System (NPDES) permits program, states should establish
additional procedures to facilitate translation offish tissue criteria concentrations into water
concentration permit limits.
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2 Introduction and Background
National Ambient Water Quality Criteria (AWQC) are established by the United States
Environmental Protection Agency (EPA) under the Clean Water Act (CWA). As provided for by
the Clean Water Act, EPA reviews and from time to time revises 304(a) AWQC to ensure the
criteria are consistent with the latest scientific information. Section 304(a) aquatic life criteria
serve as recommendations to states and tribes in defining ambient water concentrations that will
protect against adverse ecological effects to aquatic life resulting from exposure to a pollutant
found in water from direct contact or ingestion of contaminated water and/or food. Aquatic life
criteria address the CWA goals of providing for the protection and propagation offish and
shellfish. When adopted into state water quality standards (WQS), these criteria can become a
basis for establishing National Pollutant Discharge Elimination System (NPDES) program
permit limits and Total Maximum Daily Loads (TMDLs).
2.1 History of the EPA Selenium AWQC for Aquatic Life
In 1980 EPA first published numeric aquatic life criteria for selenium in freshwater.
These criteria were based on water-only exposure (no dietary exposure). In order to address the
lack of consideration of bioaccumulation in the 1980 selenium criteria, in 1987 EPA published
updated selenium criteria to address field-based toxicity observed in aquatic ecosystems at levels
below the existing criteria values. The 1987 criteria were field-based and account for both the
water-column and dietary uptake pathways manifested at Belew's Lake, North Carolina (USA), a
cooling water reservoir that had been affected by selenium loads from a coal-fired power plant.
At that time EPA also provided an acute criterion of 20 jig/L derived from a reverse application
of an acute-chronic ratio obtained from conventional water-only exposure toxicity tests applied
to the 5 |ig/L chronic value based on dietary and water column exposure in Belew's Lake.
In 1998-1999 EPA published a revised acute criterion, a formula that recognized that the
two oxidation states, selenate and selenite, appeared to have substantially different acute
toxicities. This acute criterion assumed water-only exposure, neglecting the dietary uptake
pathway. In addition, subsequent research has demonstrated that sulfate levels may affect
selenate toxicity in water-only exposures.
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In 1998 EPA held a peer consultation workshop to evaluate new science available for
selenium relevant to the selenium aquatic life criterion. EPA concluded, and the peer reviewers
agreed, that fish-tissue values better represent chronic adverse effects of selenium than the
conventional water concentration approach used by EPA to protect aquatic life, because chronic
selenium toxicity is primarily based on the food-chain bioaccumulation route, not a direct
waterborne route.
In 2004 EPA published a draft chronic whole-body fish-tissue criterion with a water-
based monitoring trigger in the summer and fall. The critical effect considered at that time was
the impact on survivorship based on overwintering stress to bluegill sunfish. An acute criterion
was estimated at that time that addressed concerns with the species of selenium present and
adjusted for sulfate levels; however, it did not address the dietary uptake pathway.
Further refinement of the fish tissue approach occurred in 2009 based on the findings of a
Pellston scientific workshop on the ecological risk assessment of selenium (Chapman et al. 2009,
2010). As presented by Chapman et al. (2009), some key findings resulting from that workshop
are:
• Diet is the primary pathway of selenium exposure for both invertebrates and vertebrates.
• Traditional methods for predicting toxicity on the basis of exposure to dissolved [water-
column] concentrations do not work for selenium because the behavior and toxicity of
selenium in aquatic systems are highly dependent upon site-specific factors, including
food web structure and hydrology.
• Selenium toxicity is primarily manifested as reproductive impairment due to maternal
transfer, resulting in embryotoxicity and teratogenicity in egg-laying vertebrates.
For this 2014 external peer review draft, EPA has conducted a new literature review and
reanalyzed data considered in the 2004 and 2009 draft criteria documents. The 2014 criterion
reflects the latest scientific consensus (e.g., Chapman et al. 2010) on the reproductive effects of
selenium on aquatic life and their measure in aquatic systems. The criterion presented here
supersedes all previous national aquatic life water quality criteria for selenium. Because of the
bioaccumulative nature of selenium, EPA is recommending a national chronic criterion that is
expressed as 4 elements. Two elements are fish tissue-based (concentrations in the egg-ovary and
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whole-body or muscle tissue). Two elements are water column based (a 30 day value and/or an
intermittent value derived from equation). The 30-day average water concentration is protective
against chronic effects of selenium derived from modeling selenium bioaccumulation via the
food web in lotic and lentic waterbody types. To address intermittent exposures that could
contribute to chronic effects from selenium bioaccumulation, EPA is also recommending an
intermittent exposure water concentration element intended to limit cumulative exposure, based
on the chronic 30-day water criterion. These water quality criterion elements apply to the total of
all oxidation states (selenite, selenate, organic selenium, and any other forms).
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3 Problem Formulation
Problem formulation provides a strategic framework for water quality criteria
development by focusing the effects assessment on the most relevant chemical properties and
endpoints. The structure of this effects assessment is consistent with EPA's Guidelines for
Ecological Risk Assessment (U.S. EPA 1998).
This ecological effects assessment defines a scientifically-defensible water quality
criterion for selenium under section 304(a)(l) of the Clean Water Act. The goal of the Clean
Water Act is to protect and restore the biological, chemical and physical integrity of waters of
the U.S. Clean Water Act Section 304(a)(l) requires EPA to develop criteria for water quality
that accurately reflect the latest scientific knowledge. These criteria are based solely on data and
best professional scientific judgments on toxicological effects. Criteria are developed following
overarching guidance outlined in the Agency's Guidelines for Deriving Numerical National
Water Quality Criteria for the Protection of Aquatic Organisms and Their Uses (Stephan et al.
1985).
States and authorized tribes may adopt EPA's recommended criteria into their water
quality standards to protect designated uses of water bodies, or they may modify EPA's criteria
to reflect site-specific conditions, or they may derive criteria using other scientifically-defensible
methods, all subject to EPA review and approval.
3.1 Overview of Selenium Sources and Occurrence
Selenium is a naturally occurring element present in sedimentary rocks and soils. It is
also present in the environment as methyl derivatives of selenium (Ranjard et.al., 2003). There
are around 40 known selenium-containing minerals, some of which can have as much as 30%
selenium, but all are rare and generally occur together with sulfides of metals such as copper,
zinc and lead (Emsley 2011). The distribution of organic-enriched sedimentary rocks, shales,
petroleum source rocks, ore deposits, phosphorites, and coals, in which selenium typically co-
occurs, is well characterized in the United States (Presser et al. 2004) (see Figures 1 and 2). Two
major anthropogenic activities cause selenium mobilization and introduction into aquatic
systems. The first is the mining of metals, minerals and refinement and use of fossil fuels; the
second is irrigation of selenium-rich soils.
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Mining activities bring selenium deposits to the surface, where they are exposed to
physical weathering processes (Figure 1). The release of selenium related to resource extraction
activities is most common in the phosphate-rich beds of southeast Idaho and adjacent areas of
Wyoming, Montana, and Utah, and in coal mining areas in portions of West Virginia, Kentucky,
Virginia, and Tennessee (Presser et al. 2004). When selenium-containing minerals, rocks, and
coal are mined, selenium can be mobilized when ore and waste materials are crushed, increasing
the surface area and exposure of material to weathering processes. Selenium contamination of
surface waters can also occur when sulfide deposits of iron, uranium, copper, lead, mercury,
silver, and zinc are released during the mining and smelting of these metal ores. When coal is
burned for power production, selenium can enter surface waters as drainage from fly-ash ponds
and fly-ash deposits on land (Gillespie and Baumann 1986). The refining of crude oil containing
high levels of selenium can also be a major source of loading in certain water bodies (Maher et
al. 2010).
Irrigation of selenium-rich soils for crop production in arid and semi-arid regions of the
country (Figure 2) can mobilize selenium and move off-site in surface water runoff or via
leaching into ground water. Deposits of Cretaceous marine shales have weathered to produce
high selenium soils in many areas of the western US (Lemly, 1993c). Selenium is abundant in
the alkaline soils of the Great Plains, and some ground waters in California, Colorado, Kansas,
Oklahoma, South Dakota and Wyoming contain elevated concentrations of selenium due to
weathering of and leaching from rocks and soils. In semi-arid areas of the West, irrigation water
applied to soils containing soluble selenium can leach selenium from the soil. The excess water
(in tile drains or irrigation return flow) containing selenium can run off into nearby basins,
ponds, or streams. For example, elevated selenium levels at the Kesterson Reservoir in California
were reported to come from agricultural irrigation return flow collected in tile drains (Ohlendorf
etal. 1986).
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Figure 1. Map indicating deposits of selenium in mining regions.
Light green shading indicates lower selenium concentrations (< 7.2 mg/L), whereas darker green
shading indicates higher selenium concentrations (> 7.2 mg/L) in underlying geology. Source of
Map: SAIC, 2008.
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\C-~ ^
I .
0 250 500 KILOMETERS
EXPLANATION
\n'j MJMC|iltlilf In . naiJHriiuiiim Vvh. r-' ••', if-ivfH-ri
IndPX Is gfpaler than 2 5 -mcl ntKf* gpokftlr iniis .itv- rrutnh
1 ;| I-T ' r> i i •.'••II >•• l.-n:.ir, num.. <..i';ii..'U.ii / •(. )•• i it-
M.:'1 •!•'! JJjlll Ulflll .ll I. lltll
Figure 2. Areas of western U.S. susceptible to selenium contamination (gray) and where
agricultural land is irrigated (green).
Overlap of gray and green show areas susceptible to selenium discharge from irrigation. Note:
Eastern U.S. is not as susceptible since selenium does not occur at surface where agricultural
practices can mobilize selenium. Source of Map: Seilor et al. 1999 page 31.
Atmospheric emissions of selenium can originate from several sources including power
plants and other facilities that burn coal or oil, selenium refineries that provide selenium to
industrial users, base metal smelters and refineries, resource extraction industries, milling
operations, and end-product manufacturers (e.g., semiconductor manufacturers)(ATSDR, 2003).
Airborne selenium particles can settle either on surface waters or on soils from which selenium is
transported and deposited into water bodies through conveyances or runoff.
The chemical form of selenium that dominates a location is usually dependent on its
sources, effluent treatments, and biogeochemical processes in the receiving waters. Table 1
shows the predominant form of selenium that is associated with different activities and
industries.
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Table 1. Predominant chemical forms of selenium in discharges associated with
different activities and industries.
Selenium Form
Selenate
Selenite
Organoselenium
Sources
Agricultural irrigation drainage
Treated oil refinery effluent
Mountaintop coal mining/ valley fill leachate
Copper mining discharge
Oil refinery effluent
Fly ash disposal effluent
Phosphate mining overburden leachate
Treated agricultural drainage (in ponds or lagoons)
Source: Presser and Ohlendorf 1987; Zhang and Moore 1996; Cutter and Diego-McGlone 1990.
3.2 Environmental Fate and Transport of Selenium in the Aquatic Environment
As a member of Group 16 of the Periodic Table, selenium is a non-metallic chemical
element with chemical activity and physical properties similar to sulfur and tellurium. Selenium
speciation has important influences on the fate of the element, and therby occurrence. Because
reproductive effects are based on selenium concentrations in fish egg/ovary tissue, the effects are
integrated across forms of selenium; thus water column values are based on total selenium
exposure.
3.2.1 Selenium Species in Aquatic Systems
^^ r\
The primary selenium species present in water are the anions selenate (SeC>4 or Se[VI]),
r\
selenite (SeOs , or Se[IV]) and organo-selenide (e.g., selenomethionine or org-Se[II]). Selenate
usually predominates in well-aerated surface waters such as rivers and streams, especially under
alkaline conditions, and is associated with calcareous soils. In water selenite tends to dominate in
slow moving waters such as lakes and reservoirs. In soils, selenite is more typically found in
acidic conditions (McLean and Bledsoe 1992). Organoselenium containing carbon-selenium
chemical bonds is also found in water. The proportion of these different forms of selenium found
in aquatic systems can vary. Factors that enhance selenium mobility in soils are; alkaline pH,
high selenium concentration, oxidizing conditions, and high concentrations of other anions that
strongly adsorb to soils, in particular phosphate (Balistrieri and Chao 1987).
The distribution of selenium among dissolved species cannot be predicted from
thermodynamic equilibrium alone. Biological (kinetically driven) processes are just as important
as geochemical processes in determining the forms of selenium that are present (Cutter and
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Bruland 1984). Biological processes are difficult to predict from environmental characteristics,
so conventional speciation modeling is problematic for selenium. On the other hand, selenium is
one of the few elements for which the different species can be directly measured at
environmental concentrations (Cutter and Bruland 1984; Cutter and Cutter 2004). These data
show that geologic and anthropogenic sources often release mostly selenate (U.S. EPA 1992),
which is not reactive with particle surfaces, and is highly mobile in soils. Some types of bacteria
r\
convert selenate to elemental selenium in sediments (Oremland 1990). Selenate (SeC>4 ) in the
r\
water column is taken up only slowly by bacteria, especially if competition with sulfate (SC>4 )
is involved. Selenite is more reactive because of its more polar character, and tends to adsorb to
soils and soil constituents (McLean and Bledsoe 1992). Evidence indicates that selenite is
reduced rapidly, even before uptake in some cases, making it difficult to distinguish between
uptake and metabolic processes (Milne 1998). Freshwater phytoplankton process selenate and
selenite by different mechanisms, leading to different concentrations within the cell (Riedel et al.
1991).
In nature, adverse effects from selenium are determined by a sequence of processes. The
transformation of selenium to organic forms by organisms via methylation acts to increase the
solubility and therefore bioavailability (Simmons and Wallschlager 2005). Generally, selenides
(Se2~) can be the precursors to organic selenides (e.g. methylated selenides, soluble seleno-amino
acids), and also relatively insoluble inorganic selenides. When any form of selenium is taken up
at the base of the food web by plants and microbes, it is converted to organo-selenide (Wrench,
1978). Organo-selenide is released back to the water column as these cells die or are consumed
(Lee and Fisher, 1994), where some selenite is formed. In water, macrophytes and other plants
(algae, phytoplankton) can readily take up selenite and selenate and incorporate selenium in the
tissue as selenomethionine. In general, selenium concentrations in algae, microbes, sediments, or
suspended particulates are 100-500 times higher than dissolved concentrations in selenate
dominated environments such as streams and rivers. But when selenite or organo-selenide is
proportionately more abundant, the ratio can be 1000-10,000, such as in wetlands, some
estuaries, the oceans, and pure phytoplankton cultures. This variability of particulate
concentrations relative to dissolved concentrations is a major cause of the variability in the
relationship between selenium in water and selenium in organisms (Luoma and Presser 2009).
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Methylated selenides produced by biological reduction of selenite, usually occur at very
low concentrations in water relative to the inorganic selenium species, and differ between
flowing (lotic) and standing (lentic) fresh waters (Simmons and Wallschlager 2005). The
accumulation patterns for selenium from water to sediment identify higher rates of methylation
in lentic compared with lotic environments, and correspondingly higher accumulation rates of
selenium in biota in lentic environments (USEPA, 2004). The result is a build-up of
proportionately more organo-selenides and selenite as selenium is recycled through the base of
food webs, and proportionately less selenate. This unidirectional build-up of potentially reactive
forms, especially in environments where water residence times are extended (e.g., wetlands,
estuaries, and lakes) is a key factor in the ecological risks posed by selenium. Anaerobic
microbial reduction of selenate and selenite to insoluble elemental selenium can represent an
important mechanism for removing selenium from water and transferring it to sediments (Lemly.
2004).
3.2.2 Bioaccumulation of Selenium in Aquatic Systems
Dissolved selenium uptake by animals is slow, whatever the form, such that under
environmentally relevant conditions, dissolved selenium in the water column makes little or no
direct contribution to bioaccumulation in animals (Lemly 1985a; Ogle and Knight 1996), but
does influence the concentration of selenium in particulate matter. Selenium bioaccumulation in
aquatic organisms occurs primarily through the ingestion of food (Fan et al. 2002; Ohlendorf et
al. 1986; Saiki and Lowe 1987; Presser and Ohlendorf 1987; Luoma et al. 1992; Presser et al.
1994; Chapman et al. 2010). However, unlike other bioaccumulative contaminants such as
mercury, the single largest step in tissue selenium accumulation in aquatic environments occurs
at the base of the food web where algae and other microorganisms accumulate selenium from
water by factors of up to one million-fold (Orr et al. 2012; Stewart et al. 2010). Bioaccumulation
and transfer through aquatic food webs constitute the major biogeochemical pathways of
selenium in aquatic ecosystems. Dissolved selenium oxyanions are primarily absorbed by aquatic
producers (trophic level 1 organisms), including phytoplankton and bacteria, and biotransformed
into elemental selenium and organoselenium. These organisms, together with other particle-
bound selenium sources, constitute the particulate selenium fraction in the water column.
Selenium can then be transferred from these trophic level 1 organisms to aquatic primary
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consumers such as zooplankton, insect larvae, larval fish, and bivalves (trophic level 2), and then
to predators such as fish and birds (trophic level 3 and above).
In addition to the water concentration of selenium, selenium bioaccumulation depends on
several factors specific to each aquatic system. These factors include:
Water residence time. Residence time is a measure of the average time a water molecule
will spend in a specified region of space. Residence time influences both the proportion of
selenium found in particulate and dissolved forms and the predominant form of selenium.
Organisms in waters with long residence times such as lakes, ponds, reservoirs, wetlands or
estuaries will tend to bioaccumulate more selenium than those living in waters with shorter
residence times such as rivers and streams (ATSDR 2003; Luoma and Rainbow 2005; Orr et al.
2006; Simmons and Wallschlagel 2005; EPRI2006). Several interrelated factors underlie
selenium's greater bioaccumulation potential in slow moving systems such as food web
complexity, and the organic content and reduction/oxidation potential of sediments. In addition,
the biogeochemical processes that result in more bioavailable forms of selenium (selenite and
organoselenium) occur to a greater extent in waterbodies with long residence times because
sediments where these chemical reactions take place tend to settle in depositional areas rather
than being transported downstream. As a result, selenium toxicity in flowing waters with short
residence times may only be apparent far downstream of their selenium sources, whereas waters
with long residence times are more likely to exhibit selenium toxicity near their sources (Presser
& Luoma 2006).
^^
Distribution of selenium between particulate and dissolved forms. Selenium is found in
both particulate and dissolved forms in water. The proportion of selenium found in particulate
matter (algae, detritus, and sediment) is important because it is the primary avenue for selenium
entering into the aquatic food web (Ohlendorf et al. 1986; Saiki and Lowe 1987; Presser and
Ohlendorf 1987; Luoma et al. 1992; Presser et al. 1994; Presser & Luoma 2006; Luoma and
Rainbow 2005). Direct transfer of selenium from water to animals is a small proportion of total
exposure.
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Bioaccumulation in prey. Trophic level 1 organisms such as algae and bacteria, detritus,
and other forms of particulate material containing selenium are ingested by trophic level 2
organisms such as mollusks, planktonic crustaceans, and many insects, increasing the
concentration of selenium in the tissues of these organisms. Differences in the physiological
characteristics of these organisms result in different levels of bioaccumulation. Also, selenium
effects on invertebrates typically occur at concentrations higher than those that elicit effects on
vertebrates (e.g., fish and birds) that prey upon them. Additionally, mollusks such as mussels and
clams accumulate selenium to a much greater extent than planktonic crustaceans and insects
(although not to toxic levels) due to higher ingestions rates of both particulate-bound (algae) and
dissolved selenium from the water column through filter feeding, as well as the lower rate at
which they eliminate selenium (Luoma and Rainbow 2005). Because egg-laying vertebrates are
the most sensitive groups to selenium (Janz et al. 2010), oviparous vertebrate consumers such as
fish and birds are consequently the most vulnerable groups to selenium poisoning and the focal
point of most environmental assessments (Ogle and Knight 1996, Stewart et al., 2010).
Trophic transfer to predators. Bioaccumulation of selenium by higher trophic level
organisms, such as trophic level 3 and 4 fish, is highly influenced by the food web of the aquatic
environment. For example, fish that primarily consume mollusks will exhibit greater selenium
bioaccumulation than fish that consume primarily insects or crustaceans from waters with the
same concentration of dissolved selenium because mollusks tend to accumulate selenium at
higher concentrations than other trophic level 2 organisms, as noted above (Luoma and Presser,
2009).
3.3 Mode of Action and Toxicity of Selenium
Selenium is a naturally occurring chemical element that is also an essential micronutrient.
Trace amounts of selenium are required for normal cellular function in almost all animals.
However, excessive amounts of selenium can also have toxic effects, with selenium being one of
the most toxic of the biologically essential elements (Chapman et al. 2010). Egg-laying
vertebrates have a lower tolerance than do mammals, and the transition from levels of selenium
that are biologically essential to those that are toxic occurs across a relatively narrow range of
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exposure concentrations (Luckey and Venugopal 1977; USEPA 1987, 1998; Haygarth 1994;
Chapman et al. 2009, 2010).
Selenium is a member of the sulfur group of nonmetallic elements and consequently the
two chemicals share similar characteristics. Selenium can replace sulfur in two amino acids, the
seleno-forms being selenomethionine and selenocysteine. It has been a long-standing hypothesis
that the cause of malformations in egg-laying vertebrates is due to the substitution of selenium
for sulfur in these amino acids and their subsequent incorporation into proteins causing
disruption of the structure and function of the protein. When present in excessive amounts,
selenium is erroneously substituted for sulfur, resulting in the formation of a triselenium linkage
(Se-Se-Se) or a selenotrisulfide linkage (S-Se-S), either of which was thought to prevent the
formation of the normal disulfide chemical bonds (S-S). The end result was thought to be
distorted, dysfunctional enzymes and protein molecules that impaired normal cellular
biochemistry (Diplock and Hoekstra 1976; Reddy and Massaro 1983; Sunde 1984).
Recent research, however, suggests that selenium's role in oxidative stress plays a role in
embryo toxicity, whereas selenium substitution for sulfur does not. The substitution of
selenomethionine for methionine does not appear to affect either the structure or function of
proteins (Yuan et al. 1998; Mechaly et al. 2000; Egerer-Sieber et al. 2006). The reason is
apparently due to selenium not being distally located in selenomethionine and therefore its effect
on the tertiary structure of the protein is insulated. Although the incorporation of
selenomethionine into proteins is concentration-dependent (Schrauzer 2000), selenocysteine's
incorporation into proteins is not (Stadtman 1996). This suggests that neither selenomethionine
nor selenocysteine affect protein structure or function. In fact, Se as an essential micronutrient is
incorporated into functional and structural proteins as selenocysteine.
The role of selenium-induced oxidative stress in embryo toxicity and teratogenesis
appears to be related to glutathione homeostasis. A review of bird studies by Hoffman (2002)
showed exposure to selenium altered concentrations and ratios of reduced to oxidized glutathione
thereby increasing measurements of oxidative cell damage. Palace et al. (2004) suggested
oxidative stress due to elevated selenium levels results in pericardial and yolk sac edema in
rainbow trout embryos. Evidence for the role of oxidative stress in selenium toxicity is growing
but mechanistic studies are needed to better understand its effects on egg-laying vertebrates. For
a more in depth discussion on the mechanism of toxicity at the cellular level including the
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evidence against sulfur substitution as a cause and the role of oxidative stress see Janz et al.
(2010).
The most well-documented, overt and severe toxic symptoms in fish are reproductive
teratogenesis and larval mortality. Egg-laying vertebrates appear to be the most sensitive taxa,
with toxicity resulting from maternal transfer to eggs. Selenium consumed in the diet of adult
female fish is deposited in the eggs, when selenium replaces sulfur in vitellogenin, which is
transported to the ovary and incorporated into the developing ovarian follicle (Janz et al. 2010),
the primary yolk precursor. In studies involving young organisms exposed through transfer of
selenium from adult female fish into their eggs, the most sensitive diagnostic indicators of
selenium toxicity in vertebrates occur when developing embryos metabolize organic selenium
that is present in egg albumen or yolk. It is then further metabolized by larval fish after hatching.
A variety of lethal and sublethal deformities can occur in the developing fish exposed to
selenium, affecting both hard and soft tissues (Lemly 1993b). Developmental malformations are
among the most conspicuous and diagnostic symptoms of chronic selenium poisoning in fish.
Terata are permanent biomarkers of toxicity, and have been used to identify impacts of selenium
on fish populations (Maier and Knightl994; Lemly 1997b). Deformities in fish that affect
feeding or respiration can be lethal shortly after hatching. Terata that are not directly lethal, but
distort the spine and fins, can reduce swimming ability and overall fitness. Because the rate of
survival of deformed young would be less than that for normal young, the percentage of
deformed adults observed during biosurveys will likely understate the underlying percentage of
deformed young, although quantitation of the difference is ordinarily not possible.
In summary, the most sensitive indicators of selenium toxicity in fish larvae are effects
modulated through the reproductive process and exhibited in fish larvae as teratogenic
deformities such as skeletal, craniofacial, and fin deformities, and various forms of edema that
result in mortality (Lemly 2002). The toxic effect generally evaluated is the reduction in the
number of normal healthy offspring compared against the starting number of eggs. In studies of
young organisms exposed to selenium solely through their own diet (rather than via maternal
transfer), reductions in survival and/or growth are the effects that are generally evaluated.
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3.4 Narrow Margin between Sufficiency and Toxicity of Selenium
Selenium has a narrow range encompassing what is beneficial for biota and what is
detrimental. Selenium is an essential nutrient that is incorporated into functional and structural
proteins as selenocysteine. Several of these proteins are enzymes that provide cellular
antioxidant protection. Selenium is an essential element required as a mineral cofactor in the
biosynthesis of glutathione peroxidases. All of the classic glutathione peroxidases contain
selenium and are found to be involved in the catalytic reaction of these many enzymes (Allan
1999). The major function of the glutathione peroxidases involves the reduction of hydrogen
peroxide to water at the expense of the oxidation of glutathione, the enzyme's cofactor, an
important antioxidant process at normal dietary levels.
Aquatic and terrestrial organisms require low levels of selenium in their diet to sustain
metabolic processes, whereas concentrations of selenium that are only an order of magnitude
greater than the required level have been shown to be toxic to fish due to generation of reactive
oxidized species, resulting in oxidative stress. Dietary requirements in fish have been reported to
range from 0.05 to 1.0 mg Se/kg dw (Watanabe et al. 1997). Selenium requirements for optimum
growth and liver glutathione peroxidase activity in channel catfish were reported as 0.25 mg
Se/kg dw (Gatlin and Wilson 1984). Estimated selenium dietary requirements in hybrids of
striped bass, based on selenium retention, were reported as 0.1 mg Se/kg dw (Jaramillo 2006).
Selenium deficiency has been found to affect humans (U.S. EPA 1987), sheep and cattle (U.S.
EPA 1987), deer (Oliver et al. 1990), fish (Thorarinsson et al. 1994; Wang and Lovell 1997;
Wilson et al. 1997; U.S. EPA 1987), aquatic invertebrates (Audas et al. 1995; Caffrey 1989;
Cooney et al. 1992; Cowgill 1987; Cowgill and Milazzo 1989; Elendt 1990; Elendt and Bais
1990; Harrison et al. 1988; Hyne et al. 1993; Keating and Caffrey 1989; Larsen and Bjerregaard
1995; Lim and Akiyama 1995; Lindstrom 1991; U.S. EPA 1987; Winner 1989; Winner and
Whitford 1987), and algae (Doucette et al. 1987; Keller et al. 1987; Price 1987; Price et al. 1987;
Thompson and Hosja 1996; U.S. EPA 1987; Wehr and Brown 1985). The predominance of
research on selenium deficiency in invertebrates and algae are related to optimizing the health of
test organisms cultured in the laboratory.
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3.5 Interactions with Mercury
The most well known interactions with selenium occur with both inorganic and organic
mercury, and are generally antagonistic (Micallef and Tyler 1987; Cuvin and Furness 1988;
Paulsson and Lundbergh 1991; Siegel et al. 1991; Southworth et al. 1994; Ralston et al. 2006),
with the most likely mechanism being the formation of metabolically inert mercury selenides
(Ralston et al. 2006; Peterson et al. 2009). However, other studies have found interactions
between mercury and selenium to be additive (Heinz and Hoffman 1998) or synergistic
(Huckabee and Griffith 1974; Birge et al. 1979). The underlying mechanism for these additive
and synergistic interactions between mercury and selenium are unknown.
3.6 Assessment Endpoints
Assessment endpoints are defined as "explicit expressions of the actual environmental
value that is to be protected" and are defined by an ecological entity (species, community, or
other entity) and its attribute or characteristics (U.S. EPA 1998). Assessment endpoints may be
identified at any level of organization (e.g., individual, population, community). In the context of
the Clean Water Act, aquatic life criteria for toxic pollutants are typically determined based on
the results of toxicity tests with aquatic organisms in which unacceptable effects on growth,
reproduction, or survival occurred. The goal of criteria is to protect the diversity, productivity,
and stability of aquatic communities. To achieve this goal, the endpoint of criteria assessment is
the survival, growth, and reproduction of a high percentage of species of a diverse assemblage of
freshwater aquatic animals (fish, amphibians, and invertebrates) and plants. Toxicity data is
aggregated into a sensitivity distribution that indicates the impact of the toxicant under study to a
variety of genera representing the broader the aquatic community. Criteria are designed to be
protective of the vast majority of aquatic animal species in an aquatic community (i.e.,
approximately 95* percentile of tested aquatic animals representing the aquatic community). As
a result, health of the aquatic ecosystem may be considered as an assessment endpoint indicated
by survival, growth, and reproduction.
To assess potential effects on the aquatic ecosystem by a particular stressor, and develop
304(a) aquatic life criteria under the CWA, EPA typically requires the following as outlined in
the Agency's 1985 Guidelines (Stephan et al. 1985):
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Acute toxicity test data (mortality, immobility, loss of equilibrium) for aquatic animals
from a minimum of eight diverse taxonomic groups is required. The diversity of tested species is
intended to ensure protection of various components of an aquatic ecosystem. In the case of
bioaccumulative compounds like selenium, these acute toxicity studies do not address risks that
result from exposure to chemicals via the diet (through the food web). They also do not account
for the slow accumulation kinetics of many bioaccumulative compounds such as selenium and
may underestimate effects from long-term accumulation in different types of aquatic systems
(SAB 2005).
Because the most sensitive adverse effects of selenium are reproductive effects on the
offspring of exposed fish, chronic effects are the focus of this selenium assessment. Shorter-term
intermittent or pulsed exposure of selenium may result in bioaccumulation through the aquatic
food web and consequently may adversely affect fish reproduction; such measures of effect are
estimated from chronic assessment endpoints in the 2014 selenium criterion document. Available
acute toxicity data based on acute water column-only exposure (i.e., LCso's) are not used in this
assessment to derive a traditional acute toxicity criterion because acute effects are not the effects
of concern for the bioaccumulative chemical selenium.
Chronic toxicity test data (longer-term survival, growth, or reproduction) for aquatic
animals are needed from a minimum of eight diverse taxonomic groups. The diversity of tested
species is intended to ensure protection of various components of an aquatic ecosystem. Specific
minimum data recommendations or requirements (MDRs) identified for development of criteria
in the 1985 EPA Ambient Water Quality Criteria Guidelines require aquatic animal toxicity data
from:
1. the family Salmonidae in the class Osteichthyes ,
2. a second family in the class Osteichthyes, preferably a commercially
or recreationally important warmwater species (e.g., bluegill, channel
catfish, etc.),
3. a third family in the phylum Chordata (may be in the class
Osteichthyes or may be an amphibian, etc.),
4. a planktonic crustacean (e.g., cladoceran, copepod, etc.),
5. a benthic crustacean (e.g., ostracod, isopod, amphipod, crayfish, etc.),
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6. an insect (e.g., mayfly, dragonfly, damselfly, stonefly, caddisfly,
mosquito, midge, etc.),
7. a family in a phylum other than Arthropoda or Chordata (e.g.,
Rotifera, Annelida, Mollusca, etc.), and
8. a family in any order of insect or any phylum not already represented.
Acceptable quantitative chronic values for selenium are available for six of the above
eight MDRs of Stephan et al. (1985) (requirements 1, 2, 3, 6, 7, and 8). Acceptable information
indicating relative insensitivity in accord with the approach of U.S. EPA (2008b) is available for
the remaining two items (4 and 5). Consequently, the chronic selenium criterion was derived
using the genus-level sensitivity distribution approach per the 1985 Guidelines (Stephan et al.
1985).
The Guidelines also require at least one acceptable test with a freshwater alga or vascular
plant. If plants are among the aquatic organisms most sensitive to the material, results of a plant
in another phylum should also be available. No tests were conducted that evaluated a
biologically relevant endpoint of an important aquatic plant species in which the concentrations
of selenite or selenate were measured. Therefore, plant endpoints were not used in this criteria
derivation, consistent with the relative sensitivity perspective of Chapman et al. 2010. A
summary of studies investigating the toxicity of selenium on aquatic plants is provided in
Appendix E.
The available scientific evidence indicates that for selenium, critical assessment
endpoints are offspring mortality and severe development abnormalities that affect the ability of
fish to swim, feed and successfully avoid predation, resulting in impaired recruitment of
individuals into fish populations. Selenium enrichment of reservoir environments (e.g., Belews
Lake, NC (Lemly 1985), Hyco Reservoir (DeForest 1999), and Kesterson Reservoir, CA
(Ohlendorf 1986) provide examples of adverse effects that occur through bioaccumulative
processes at different levels of biological organization, and comprise integrated whole-ecosystem
examples of trophic transfer resulting in population-level reductions of resident species.
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3.7 Measures of Effect
Each assessment endpoint requires one or more "measures of ecological effect," which
are defined as changes in the attributes of an assessment endpoint itself or changes in a surrogate
entity or attribute in response to chemical exposure. Ecological effects data are used as measures
of direct and indirect effects to growth, reproduction, and survival of aquatic organisms.
The amount of toxicity testing data available for any given pollutant varies significantly,
depending primarily on whether any major environmental issues are raised. An in-depth
evaluation of available data on selenium has been performed by EPA to determine data
acceptability (see Stephan et al. 1985 for additional detail).
In traditional chronic tests used in many EPA aquatic life criteria documents, organisms
are exposed to contaminated water but fed a diet grown in uncontaminated media not spiked with
the toxicant prior to introduction into the exposure chambers. Such tests are not suitable for
deriving a criterion for a bioaccumulative pollutant unless (1) effects are linked to concentrations
measured in appropriate tissues, and (2) the route of exposure does not affect the potency of
residues in tissue. For selenium, the first condition might be met, but the second condition is not,
because the route of selenium exposure appears to influence the potency of a given tissue residue
(Cleveland et al. 1993; Gissel-Nielsen and Gissel-Nielsen 1978). Consquently, toxicity tests with
water-only exposures (and any tests not relying on dietary exposure) are not included in this
assessment.
Selenium toxicity is primarily manifested as reproductive impairment due to maternal
transfer, resulting in embryo mortality and teratogenicity. Measurements offish tissue are most
closely linked to the chronic adverse effects of selenium (Chapman et al. 2010), since chronic
selenium toxicity is based on the food-chain bioaccumulation route, not a direct waterborne
route. In this selenium criterion document, water-column criterion element concentrations for
selenium were derived from fish tissue concentrations by modeling selenium transfer through the
food web. The next sections describe approaches used to establish selenium effects
concentrations in fish tissue and to relate the concentrations in fish tissue to concentrations in
water.
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3.7.1 Fish Tissue
Chronic measures of effect concentrations are the ECio,EC2o, No Observed Effect
Concentration (NOEC), Lowest Observed Effect Concentration (LOEC), and Maximum
Acceptable Toxicant Concentration (MATC). The ECio is the concentration of a chemical that is
estimated to result in a 10 percent effect in a measured chronic endpoint (e.g., growth,
reproduction, and survival); the EC20 corresponds to 20 percent effect. The NOEC is the highest
test concentration at which none of the observed effects are statistically different from the
control, as determined by hypothesis testing. The LOEC is the lowest test concentration at which
observed effects are found to be statistically different from the control. The MATC is the
geometric mean of the NOEC and LOEC.
Wherever possible, estimates of selenium concentrations associated with a low level of
effect (i.e., ECio) were calculated for each study using the computer program TRAP, Toxicity
Relationship Analysis Program (U.S. EPA 2008, 2011). The program is based on a regression
approach that models the level of adverse effects as a function of increasing concentrations of the
toxic substance. With the fitted model it is possible to estimate the contaminant concentration
associated with a small effect. Adverse effects were modeled as a sigmoid function of the
logarithmic concentrations of the toxic substance. In most cases the following logistic equation
was fit to concentration-response data using the TRAP software (U.S. EPA 2008a):
y =
\ + e(
where yo is the background response level at a selenium concentration of zero and S is the slope
at XSQ, the selenium concentration associated with a 50% reduction in y, the response level,
relative to yo. In all analyses, selenium concentrations were log-transformed. In a few cases
another mathematical function with a subtly different shape was used.
Only studies with a reference site (field surveys) or control treatment (experimental
studies) were included in the analysis, because response levels at these low (background)
selenium concentrations were the most influential points for calculating the estimated response
level at a selenium concentration of zero (yo).
When considering the use of the ECio versus the EC20, an ECio was determined to be a
more appropriate endpoint for tissue-based criteria given the nature of exposure and effects for
this bioaccumulative chemical. EC2os have historically been used in the derivation of EPA
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criteria applicable to the water medium. While water concentrations may vary rapidly over time,
tissue concentrations of bioaccumulative chemicals are expected to vary gradually. Thus, where
concentrations of selenium in fish tissue approach an effect threshold, there is potential for
sustained impacts on aquatic systems, relative to chemicals that are not as bioaccumulative. This
calls for use of a lower level of effect to attain sufficient protection. Further, the ECio was also
preferred over the NOEC or LOEC as these measures of effect are highly influenced by study
design, specifically the particular concentrations tested, the number of concentrations tested, the
number of replicates for each concentration, and the number of organisms in each replicate. As
noted by Campbell (2011), ECios and NOECs are generally of similar magnitude, but ECios have
the advantage of being more reproducible than NOECs (Van der Hoeven et al. 1997; Warne and van
Dam 2008). NOECs and MATCs are generally presented if calculated by the original
investigators, but were not used where an ECio could be calculated. The four lowest egg-ovary
Genus Mean Chronic Values (GMCVs), whose exact values influence the calculation of the egg-
ovary criterion, are all based solely on ECios. NOECs contribute to some of the GMCVs for less
sensitive species.
Comparing Effect Concentrations in Different Fish Tissues
In this document, chronic values are presented as tissue concentrations of selenium in
units of mg/kg dry weight (dw). Studies of chronic toxicity of selenium to aquatic organisms
measure concentrations in distinct tissues (e.g., whole body, ovaries, eggs, muscle, and liver) and
report these values as either wet weight (ww) or dw. Studies reporting tissue concentrations only
based on wet weight were converted to dry weight using tissue-specific and species-specific
conversion factors. When wet to dry weight conversion factors were not available for a given
species, conversion factors for a closely related taxon were used. In deriving the document's
primary criterion, that for egg or ovary tissue, chronic values are for those tissues directly
measured in the study. Tissue-to-tissue conversions (e.g., to estimate concentrations in an
unmeasured tissue from a study's measured tissue) involve some uncertainty because of
variability in tissue concentration ratios (Osmudson et al. 2007; deBruyn et al. 2008). Such
conversions were not needed for obtaining the egg-ovary chronic values. Tissue-to-tissue
conversions were needed for calculating the reproductive toxicity-based whole-body and muscle
chronic criterion element and water criteria concentration elements. Researchers often report
concentrations of selenium in fish eggs or ovaries (Holm et al. 2005; Kennedy et al. 2000;
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Hermanutz et al. 1996). The selenium concentrations in eggs and ovaries are usually assumed to
be approximately equal. Osmundson et al. (2007) found reduced levels of selenium in ovaries
after spawning, presumably due to the loss of selenium through spawning and release of eggs
with relatively high concentrations of selenium. In this document, concentrations of selenium in
ovaries are considered equivalent to concentrations of selenium in eggs because most studies
measured selenium in the ovaries prior to spawning.
The overall assessment was structured to include both reproductive and non-reproductive
studies. Selenium in eggs or ovaries is used in reproductive (maternal transfer) studies, and
conversions to whole body or muscle tissue resulting in reproductive effects were estimated.
Direct measurements of selenium in whole-body or muscle are used for non-reproductive studies
to examine non-reproductive, chronic effects.
Selenium Fish Tissue Toxicity Data Fulfilling Minimum Data Needs
The toxicity data currently available for genera and species fulfilling the 1985 Guidelines
recommendations for calculation of freshwater chronic criterion are described in Section 4.1.1
4.1.2 and Appendix C and summarized in Table 3.
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Table 3. 1985 Guidelines Minimum Data Requirements Summary Table Reflecting
the Number of Species and Genus Level Mean Values Represented in the Chronic Toxicity
Dataset for Selenium in Freshwater.
Freshwater Minimum Data Requirement
1 . Family Salmonidae in the class Osteichthyes
2. Second family in the class Osteichthyes,
preferably a commercially or recreationally
important warmwater species
3. Third family in the phylum Chordata (may be
in the class Osteichthyes or may be an
amphibian, etc.)
4. Planktonic Crustacean
5. Benthic Crustacean
6. Insect
7. Family in a phylum other than Arthropoda or
Chordata (e.g., Rotifera, Annelida, or
Mollusca)
8. Family in any order of insect or any phylum
not already represented
Total
Genus Mean Chronic
Value (GMCV)
3
3
>
See text ^
See text
1
1
1
14
Species Mean Chronic
Value (SMCV)
5
3
4
See text
See text
1
1
1
17
The first three of these MDRs in Appendix C are easily fulfilled by the fish species
represented in Section 4.1.1 and Appendix C. Because the field observations of contaminated
sites have found effects on fish and birds in the absence of changes in invertebrate assemblages,
scientific studies on invertebrates on chronic toxicity of dietary selenium have been very limited.
The few dietary chronic toxicity studies that are available for invertebrate species indicate that
they are among the more tolerant aquatic taxa, with available data indicating invertebrate mean
chronic values ranging from approximately 3 to 12 times higher than the fish whole body
criterion value recommended in this document (data provided in Section 4.1.3) The above
invertebrate data address MDRs 6-8, leaving only MDRs 4 and 5, for the planktonic and benthic
crustaceans, to be addressed. Because the 5* percentile calculation methods for the Final Chronic
Value (FCV) per the Guidelines use actual numeric values for the GMCVs of only the four most
sensitive (fish) genera in the selenium dataset, it is only necessary to know that the more tolerant
genera have GMCVs that are greater than those of the low four. A recommendation in the draft
white paper on Aquatic Life Criteria for Contaminants of Emerging Concern Part I (U.S. EPA
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2008b), which was supported by the Science Advisory Board, states "because only the four most
sensitive genus mean chronic values (GMCVs) are used in the criteria calculations, chronic
testing requirements for a taxon needed to meet an MDR should be waived if there is sufficient
information to conclude that this taxon is more tolerant than the four most sensitive genera." If
this concept is applied to the selenium criterion 5* percentile calculations, actual GMCVs for
MDRs 4 and 5 (the two crustacean MDRs) should be waived and counted in the total number of
GMCVs in the dataset, based on (a) the difference in the measured effect values discussed above,
and (b) the lack of observed invertebrate field effects linked to selenium (for example, as
concluded by Lemly 2002, pages 21-23, and Janz et al. 2010). It is thus concluded that there is
adequate data to fulfill the data needs for developing a chronic selenium criterion.
The total number of GMCVs available to derive the chronic criterion is 14. These include
nine fish genera from Section 4.1.1 (Salmo, Lepomis, Micropterus, Oncorhynchus, Pimephales,
Gambusia, Esox, Cyprinodon, and Salvelinus) [Added to these are the tested invertebrate genera
Centroptilum, Brachionus, and Lumbriculus from Section 4.1.3], and lastly the two waived
genera for MDRs 4 and 5 (crustaceans).
3.7.2 Water
Because fish tissue measurements of selenium are not available for many waters, the EPA
is estimating chronic measures of effect in the water-column using the chronic effect level
measured by fish tissue. The chronic criterion element for the water column is the 30-day
average concentration that corresponds to the concentration of selenium in fish tissue estimated
to result in a 10 percent effect in fish in the water body type under consideration (lotic or lentic
water bodies as described below in Section 4.2.4). The chronic criterion element for the water-
column is derived by modeling trophic transfer of selenium through the food web resulting in the
fish tissue concentration that yields the chronic reproductive effects of concern.
The EPA collaborated with the United States Geological Survey to develop and peer-
review (ERG 2008) a model relating the concentration of selenium in fish tissue to the water-
column. The approach is based on bioaccumulation and trophic transfer through aquatic system
food-webs. Model parameters are calculated using both field and laboratory measurements of
selenium in water, particulate material (algae, detritus and sediment), invertebrates, fish whole-
body, and fish egg-ovary. This model (which is a set of equations) is described in more detail in
the Analysis Section 4.2.
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3.7.3 Summary of Assessment Endpoints and Measures of Effect
The typical assessment endpoints for aquatic life criteria are based on effects on growth,
reproduction, or survival of the assessed taxa. These measures of effect on toxicological
endpoints of consequence to populations are provided by results from toxicity tests with aquatic
plants and animals. The toxicity values (i.e., measures of effect expressed as genus means) are
used in the genus sensitivity distribution of the aquatic community to derive the aquatic life
criteria. Endpoints used in this assessment are listed in Table 4.
Table 4. Summary of Assessment Endpoints and Measures of Effect Used in
Criteria Derivation for Selenium.
Assessment Endpoints for the Aquatic
Community
Measures of Effect
Survival, growth, and reproduction of
freshwater fish, other freshwater vertebrates,
and invertebrates
For effects from chronic exposure:
1. EC 10 concentrations in egg and ovary, for
offspring mortality and deformity.
2. Estimated reproductive ECio in whole
body and muscle.
3. Estimated concentrations (|ig/L) in water
linked to egg-ovary ECioS by food web-
modeling.
4. Intermittent water concentrations yielding
exposure equivalent to the above.
For acutely lethal effects:
Acute toxicity effects based on standard
water column-only toxicity testing are not
provided here for selenium, due to the
dominant significance of chronic effects.
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3.7.4 Conceptual Model of Selenium Effects on Aquatic Life
Selenium Sources
Naturally elevated selenium in soils - agricultural irrigation practices (Western US only)
Mining activities - coal, metals and sulfide minerals, phosphate
Selenium in Water Column
Dissolved - selenate, selenite, selenides
Particle-bound - selenate, selenite, selenides
Algal/Plant Transformation and Enrichment:
Inorganic forms <-»• Organoselenium forms
• As function of sorption to particulates (sediment, algae, detritus)
As function of system hydrodynamics, lotic & lentic systems, residence time
Initial Trophic Transfer of Organoselenium
from phytoplankton, periphyton, macrophytes, detritus, & sediment
I
Secondary Trophic Transfer of
Organoselenium from
macroinvertebrates/
icthyoplankton/ other zooplankton
V^ J
Tertiary Trophic Transfer of
Organoselenium from lower
trophic level fish
To higher trophic level fish
V J
4
(Reproductive Impairment.
Larval skeletal deformities.
Larval mortality.
Population decline
Loss of species and community
change
Figure 3. Diagram of selenium partitioning, bioaccumulation, and effects in the aquatic
environment.
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The above conceptual model links sources, transformation and uptake through media
phases, and consumer transfer and dynamics reflective of the movement of selenium through
ecosystems (Figures 3). Diet is the dominant pathway of selenium exposure for both
invertebrates and vertebrates. Selenium moves from water to particulates, a collection of biotic
and abiotic compartments that includes primary producers, detritus, and sediments, which form
the base of aquatic food webs. Transfer from particulates to primary consumers (e.g.,
macroinvertebrates) to fish is species specific. Knowledge of the food web is one of the keys to
determining which biological species or other ecological characteristics will be affected. Other
important parameters include rates of input of selenium into the system, hydraulic residence
time, and selenium speciation in water and particulates.
3.8 Analysis Plan
During the development of CWA section 304(a) criteria, EPA assembles all available test
data and considers all the relevant data that meet acceptable data quality and test acceptability
standards. This criterion update document is specific to selenium in fresh water. Chronic
criterion elements for selenium are protective concentrations measured in fish tissue and related
protective water concentrations generated using food-web modeling. Further modeling is used to
estimate short-term concentrations in water from intermittent or pulsed exposures that are
protective against the chronic effect. Available data indicate freshwater plants are not more
sensitive to selenium than freshwater animals, thus, a plant criterion element was not developed.
3.8.1 Analysis Plan for Derivation of the Chronic Fish Tissue-Based Criterion Element
Data for possible inclusion in the selenium dataset were obtained primarily by search of
published literature using EPA's public ECOTOX database and includes studies provided to
EPA in public comments. These studies were screened for data quality as described in the
"Guidelines for Deriving Numerical National Water Quality Criteria for the Protection of
Aquatic Organisms and Their Uses" (Stephan et al. 1985), but adjusted for factors related to
dietary lab or field exposure, which were not considered at the time the Guidelines were written.
Chronic toxicity studies were further screened to ensure they contained the relevant
chronic exposure conditions of selenium to aquatic organisms (i.e., dietary, or dietary and
waterborne selenium exposure), measurement of chronic effects, and measurement of selenium
in tissue(s). It has been well established that diet is the primary route of exposure that controls
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selenium toxicity to fish, the taxonomic group considered to be the most sensitive to chronic
selenium exposure (Coyle et al. 1993; Hamilton et al. 1990; Hermanutz et al. 1996, Chapman et
al. 2010). Furthermore, the toxic potency offish tissue residues acquired by routes other than
dietary exposure do not appear to be equivalent to those acquired by through diet (Cleveland et
al. 1993; Gissel-Nielsen and Gissel-Nielsen 1978). For example, Cleveland et al. (1993) used
water-only and dietary exposure in separate tests with bluegill. Although water-only exposure
required hundreds of ug/L to significantly elevate the tissue concentrations, those tissue
concentrations began producing effects at 3-4 mg Se/kg dw WB. In contrast, through dietary
exposure, no effects were observed at 14 mg Se/kg dw WB, the highest tissue concentration
tested. This is hypothesized to be a result of the fact that the form or speciation of selenium
differs among exposure routes (diet, water), and that the different forms have differing toxicities.
Because water-only chronic exposure would rarely if ever occur in the environment at that
magnitude, the criterion derivation uses only those studies in which test organisms were exposed
to selenium in their diet, either via laboratory or field exposure, and either alone or in
conjunction with elevated water exposure, because such studies most closely replicate real-world
exposures (diet and/or diet plus water). This approach accords with findings and
recommendations of the 2009 SET AC Pellston Workshop (Chapman et al. 2009, 2010).
EPA grouped studies based on whether the effects were chronic reproductive (e.g.,
effects on offspring survival or morphology) or chronic non-reproductive (e.g., juvenile growth
and survival). At the 2009 Pellston workshop (Chapman et al. 2009, 2010), a group of 46 experts
in the area of ecological assessment of selenium in the aquatic environment agreed that the most
important toxicological effects of selenium in fish arise following maternal transfer of selenium
to eggs during vitellogenesis, resulting in selenium exposure when hatched larvae undergo yolk
absorption. Such effects include larval mortality or permanent developmental malformations,
such as skeletal and craniofacial deformities. Therefore, the chronic fish-tissue-based criteria are
based on reproductive effects only.
The egg-ovary Species Mean Chronic Values (SMCVs) were calculated from the chronic
values (ECios and occasionally NOECs) obtained from the relevant toxicity tests. Genus Mean
Chronic Values (GMCVs) were calculated from the SMCVs and then rank-ordered from least to
most sensitive. The egg-ovary Final Chronic Value (FCV) was calculated from regression
analysis of the four most sensitive GMCVs, in this case extrapolating to the 5* percentile of the
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distribution represented by the tested genera. The FCV directly serves as the fish tissue egg-
ovary criterion concentration element without further adjustment because the underlying ECios
represent a low level of effect (per the 1985 Guidelines).
For the whole-body and muscle criteria concentrations, the egg-ovary Genus Mean
Chronic Values were converted to estimated equivalent whole-body or muscle GMCVs. The
criterion concentration element expressed as whole-body or as muscle concentration was
calculated in manner similar to the egg-ovary criterion element using conversion factors
described below, from their respective genus-level sensitivity distributions.
3.8.2 Analysis Plan for Derivation of Duration of Fish Tissue Criterion Elements
A numerical value for the fish tissue criterion elements averaging period, or duration, is
specified as instantaneous, because fish tissue data provide point, or instantaneous,
measurements that reflect integrative accumulation of selenium over time and space in the fish at
a given site. Selenium concentrations in fish tissue are expected to change only gradually over
time (Section 4.2.6 and Appendix G) in response to environmental fluctuations, thus, there
would be relatively little practical difference between outcomes with different possible
specifications of the duration of an averaging period. When fish tissue concentrations are
excessive, their duration can be assumed to occur for a sufficient length of time to elicit the
effect associated with that concentration.
3.8.3 Analysis Plan for Derivation of Chronic Water-based Criterion Element
The relationship between the ambient concentration of selenium in water and the
concentration of selenium in the eggs or ovaries offish is primarily through trophic transfer of
selenium, which is greatly affected by site-specific conditions. The EPA is using a method based
on a peer-reviewed model to derive water concentrations from the egg-ovary criterion that
explicitly recognizes partitioning of selenium in water and particulate material (algae, detritus,
and sediment), and trophic transfer from particulate material to aquatic invertebrates, from
invertebrates to fish, and the partitioning in fish whole-body and fish eggs and ovaries. The
method is composed of four main steps:
1. Formulate a mathematical equation relating the concentration of selenium in the eggs and
ovaries offish to the ambient concentration of selenium in the water column.
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2. Develop parameters needed to use the mathematical equation formulated in step 1 from
available empirical or laboratory data related to selenium bioaccumulation in aquatic
organisms.
3. Account for bioaccumulation variability across aquatic sites by evaluating the
bioaccumulation potential at the base of the aquatic food web, and classifying categories of
aquatic systems where a single water column concentration would be adequately protective.
4. Apply a statistical threshold to the distribution of translated water column concentrations for
each aquatic system category to derive a protective water column criterion for each aquatic
system category.
The EPA worked with the United States Geological Survey to derive a translation
equation to estimate the site-specific concentration of selenium in the water column
corresponding to the egg-ovary criterion concentration. This equation utilizes a mechanistic
model of bioaccumulation previously published in peer-reviewed scientific literature (Luoma et.
al., 1992; Wang et. al., 1996; Luoma and Fisher, 1997; Wang, 2001; Schlekat et al. 2002b;
Luoma and Rainbow 2005; Presser and Luoma 2006; Presser and Luoma 2010; Presser 2013).
The equation uses site-specific food web models, species-specific bioaccumulation parameters
(CF and TTF), and a site-specific enrichment factor (EF) to calculate a site-specific water
column concentration element from the egg-ovary criterion element.
Empirical or laboratory data related to selenium bioaccumulation in aquatic organisms
are needed to calculate species-specific TTF and CF parameters and site-specific EF parameter.
The EPA obtained these data by searching published literature using EPA's public ECOTOX
database and other publication databases. Studies were screened using the same data quality
guidelines as described above. Relevant studies contained selenium measurements from field
studies (water, particulate material, and aquatic organisms) or contained laboratory data on
physiological parameters of selenium bioaccumulation in aquatic organisms. Literature searches
for information on selenium associated with particulate matter included searches for data on all
forms of algae, detritus, inorganic suspended material, and sediment.
The EPA compiled a database of selenium concentration measurements from acceptable
field studies. Measurements were accepted if the study indicated the samples were collected in
the field, and the study identified the unit of measure, the media from which the measurement
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was made, the location from where the sample was taken, and the date the sample was collected.
The EPA only used data from studies with adequately described field collection protocols and
where concentrations were within the bounds of concentrations found using modern, rigorous
protocols in similar systems (Safiudo-Wilhelmy et al. 2004). The spatial precision of field data
sample collection locations were generally at the level of site, although aggregate measurements
was also included if exposure conditions were considered similar (e.g., averages of single or
composite measurements from several locations in the same aquatic system). The temporal
precision of sample collection times were usually at the level of the day they were collected,
although some studies only provided enough information to determine the week, month, or year.
If the day a series of samples were collected was not reported but the study provided information
that indicated the samples were taken concurrently, the sample precision was noted, but a single
effective collection date was assigned to all the samples.
The EPA also compiled a database of physiological coefficients for food ingestion rate
(IR), selenium assimilation efficiency (AE), and rate of selenium loss (ke). Coefficients were
accepted if the studies provided either the actual measurements or sufficient information to
derive them, and were reported in standard units (ke. /d; AE: %; IR: g/g-d) or could be converted
to standard units. Even though IR can be highly variable (Whitledge and Hayward 2000) IR
values of surrogate species were occasionally used.
The EPA accounts for bioaccumulation variability across aquatic sites by evaluating the
parameter EF (representing the partitioning of selenium between the dissolved and paniculate
state) from representative aquatic systems. The parameter EF is a measure of bioaccumulation
potential because it quantifies the transfer of selenium from the water column to particulate
material, which is the single most influential step in selenium bioaccumulation (Chapman et al.
2010). The EPA calculated EF values for a set of aquatic systems and applied statistical methods
to distinguish categories with similar bioaccumulation characteristics. On this basis a single
water column concentration is deemed adequately protective when it is derived using data from
aquatic sites in the same category. The EPA translated the egg-ovary criterion element into a set
of water concentration values and derived a water column criterion element for each aquatic
system category using a percentile of the water column concentrations for each category. To
ensure adequate protection, the EPA selected the 20* percentile of the distribution of median
water column values from 54 studies across the nation as the statistical cut-off. Using binary
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classification statistics, the EPA examined the the performance of the 20* percentile water
column values by evaluating how likely it would be that meeting the the water column criterion
element would achieve attainment of the fish tissue criteria element. In this analysis EPA used an
independent data set composed of measured concentrations of selenium to complete a
verification or "ground-truthing" of the selected 20* percentile water column values to evaluate
their protectveness, and found that the use of the 20* percentile water column would prevent
potential exceedances of the fish tissue criterion value over 90% of the time, i.e., false negative
conclusions regarding fish tissue exceedances would be minimized using the selected 20*
percentile water column value for the water column criteria derivation. Figure 4 diagrams the
conceptual framework the EPA used to derive water column criterion element from the egg-
ovary criterion element.
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Egg-ovary criterion
Representative aquatic systems
Egg/ovary - whole body Conversion Factors
Concentrations in whole-body offish
Trophic Transfer Functions (TTF)
Concentrations in prey
Trophic Transfer Functions (TTF)
Concentrations in paniculate material
Site-specific Enrichment Factors (EF)
Distribution of water concentrations
*
Statistical threshold
Water criterion
Figure 4. Conceptual model for translating the selenium egg-ovary concentration to a
water-column concentration.
3.8.4 Analysis Plan for Intermittent-Exposure Water-based Criterion Element Derivation
Like the chronic water criterion element, the intermittent-exposure criterion element is
intended to protect against cumulative exposure that would cause exceedance of the fish tissue
criterion element. Consequently, it is derived directly from the chronic water criterion element,
algebraically rearranging it to establish a limit on an intermittent elevated concentration that
occurs a specified percentage of time.
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4 Effects Analysis for Freshwater Aquatic Organisms
The following sections present the derivation of the freshwater aquatic life chronic tissue-
based criterion elements (fish egg-ovary, whole-body and/or muscle), chronic water column-
based criterion element, and intermittent water column-based criterion element for selenium.
These criterion element concentrations are developed to protect against the same effect,
reproductive impairment in fish due to maternal transfer to offspring, resulting in mortality and
teratogenicity. The fish whole-body and muscle criterion element concentrations are derived by
conversion of egg/ovary effect concentrations to whole-body and muscle effect concentrations.
The water column-based criterion element concentrations are translated from the egg-ovary
concentrations using food web modeling. The intermittent water column-based criterion element
concentrations are derived by algebraic rearrangement of the 30-day chronic water criterion
element concentrations to single day-steps.
4.1 Chronic Tissue-Based Selenium Criterion Element Concentration
Data were obtained primarily by search of published literature using EPA's public
ECOTOX database. In addition, EPA considered studies submitted with comments during the
review of the 2004 draft selenium criteria, and studies provided in response to an October 2008
Federal Register Notice of Data Availability. All available, relevant, and reliable chronic toxicity
values were incorporated into the appropriate selenium AWQC tables and used to recalculate the
Criterion Chronic Concentration (CCC) as outlined in detail in the 1985 Guidelines. The most
recent literature search extended to July 2013.
The chronic values determined from acceptable chronic toxicity studies were separated
into reproductive endpoint and non-reproductive endpoint categories. Although both sets of
endpoints assess effects due to selenium on embryo/larval or juvenile development, survival and
growth, the fundamental difference in these two categories of endpoints is exposure (inherent in
test design). That is, the fundamental difference is whether the aquatic organisms (e.g., fish)
were directly exposed to selenium in the diet and water column or via maternal transfer of
selenium to the eggs/ovaries prior to reproduction. In studies with reproductive endpoints,
parental females are exposed to selenium and the contaminant is transferred from the female to
her eggs. In the selenium-exposed females, selenium replaces sulfur in vitellogenin, the primary
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yolk precursor, which is transported to the ovary and incorporated into the developing ovarian
follicle (Janz et al. 2010). In most but not all of these studies, progeny from these females were
not additionally exposed to aqueous selenium. The chronic values derived for the reproductive
effects (survival, deformities, and edema) are based on the concentration of selenium in the eggs
or ovary, the tissues most directly associated with the observed effects. In contrast, in studies
grouped under non-reproductive effects (usually larval and/or juvenile survival or growth), the
tested fish had no maternal pre-exposure to selenium. Chronic values for non-reproductive
effects are based on the concentration of selenium in tissues measured in the study: muscle, liver
and/or whole body.
The reproductive endpoint studies applied to the derivation of the chronic criterion
elements are described below. Less definitive reproductive studies, not directly applied to the
criterion derivation are described in Section 7.1.3 and in Appendix D. Nonreproductive studies
are described in Section 7.1.8.
4.1.1 Acceptable Studies of Reproductive Effects
Below is a brief synopsis of the experimental design, test duration, relevant test
endpoints, and other critical information regarding the calculation of each specific chronic value.
The studies in this section involve effects on the offspring of exposed female fish. Data are
summarized in Table 5. Details of these studies are contained in Appendix C.
Cyprinidae
Pimephales promelas (fathead minnow)
Schultz and Hermanutz (1990) examined the effects of selenium transferred from
parental fish (females) on fathead minnow larvae. The parental fathead minnows were originally
exposed to selenite that was added to artificial streams in a mesocosm study. The selenite entered
the food web and contributed to exposure from the diet. Spawning platforms were submerged
into treated and control streams. The embryo samples that were collected from the streams were
brought into the laboratory and reared in incubation cups which received stream water dosed
with sodium selenite via a proportional diluter. Edema and lordosis were observed in
approximately 25 percent of the larvae spawned and reared in natural water spiked with 10 ug
Se/L. Selenium residues in ovaries of females from the treated stream averaged 5.89 mg/kg ww
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or 23.85 mg/kg dw (applying 75.3 average percent moisture for fathead minnow eggs/ovaries
from GEI Associates (2008) and Rickwood et al. (2008)).
The reproductive SMCV for fathead minnows is <23.85 mg Se/kg dw in ovary/eggs
based on the Schultz and Hermanutz (1990) study. Because it is a "less than" value, the
sensitivity of fathead minnows relative to the species having lower SMCVs is not certain.
However, the Young et al. (2010) observation that fathead minnow populations remained after
selenium contamination of Belews Lake had eliminated most other fish species, including
bluegill and largemouth bass, indicates that fathead minnows at that site were not as sensitive as
those species.
Esocidae
Esox lucius (northern pike)
Muscatello et al. (2006) collected spawning northern pike from four sites near a uranium
milling operation in north-central Saskatoon, Canada, with egg concentrations ranging from 2.7
to 48 mg Se/kg dw. Milt and ova were stripped from gravid fish, eggs were fertilized in the field
and then incubated in the laboratory for observations and measurements. The test was terminated
when the majority of the fry exhibited swim-up and had absorbed the yolk.
Mean egg diameter, fertilization success and cumulative embryo mortality were not
significantly different among the sites. Significant increases in percent total deformities
including edema, skeletal deformities, craniofacial deformities and fin deformities were observed
in fry originating from pike collected at the medium exposure site. The concentrations of
selenium in the northern pike eggs collected at the reference and low exposure site were very
similar, as were the percent total deformities in embryos/fry. The geometric mean of selenium in
the eggs of the adult females at the reference and low exposure sites was 3.462 mg Se/kg dw and
the corresponding arithmetic mean of the percent total deformities was 13.20%. There were only
4 adult females from exposed sites, and all had relatively similar concentrations in their eggs, all
close to the geometric mean concentration of 34.00 mg Se/kg dw. Likewise, all four exposed
females had relatively similar percent total deformities, not far from their arithmetic mean of
33.40%. This is not a sufficient level of effect for applying TRAP to determine an ECio.
Furthermore, the relatively large spread between the two clusters of exposure concentrations
(3.462 and 34.00 mg Se/kg dw) would render a NOEC and LOEC unreliable and unsuitable for
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defining a threshold. That is, the NOEC and LOEC would be a "greater than" and "less than"
values, >3.462 and <34.00 mg Se/kg dw respectively, providing little information on the
sensitivity of northern pike compared to other species.
Instead, making use of the clustering of data at low exposure and effects and at elevated
exposure and effects, the effect level for the elevated exposure eggs was normalized to the low
exposure condition and rescaled to a 0-100% range. The rescaled (i.e., Abbott-adjusted) percent
of total deformities for the elevated exposure eggs was 24% (relative to the low exposure eggs).
Thus the concentration of selenium in the elevated exposure eggs (34 mg Se/kg dw) was
equivalent to an EC 24, and is the only effects concentration that can be calculated for this test,
given the limitations in the range of concentrations tested and effects observed. Although the
EC24 is not directly translatable to an ECio for use in determining the criterion, it is useful for
comparison with the EC24 in other species in order to determine species sensitivity rank. The
EC24 for skeletal deformities from the Holm et al. (2005) study of rainbow trout, calculated to be
30.9 mg Se/kg dw in eggs, is slightly lower than the northern pike value, indicating these two
species may be similar in tolerance, with the northern pike being slightly more tolerant (see
Appendix C for more details.)
Salmonidae
Seven publications provide quantitative data on effects of selenium on salmonid
embryo/larval survival and deformity used in calculating criteria values. All involve wild-caught
adults from selenium contaminated streams, spawned for effects determination; exposure was
through the parents. These data are for rainbow trout (Oncorhynchus mykiss), cutthroat trout
(Oncorhynchus clarki), Dolly Varden (Salvelinus malma) and brown trout (Salmo truttd) and are
discussed below.
Oncorhynchus mykiss (rainbow trout)
Holm (2002) and Holm et al. (2005) obtained eggs and milt from ripe rainbow trout
collected from reference streams and streams containing elevated selenium from an active coal
mine in Alberta, Canada. In 2000, 2001 and 2002 eggs were fertilized and monitored until swim-
up stage in the laboratory, for percent fertilization, deformities (craniofacial, fmfold, and spinal
malformations), edema, and mortality. The temperature at which embryos were incubated was
8°C in 2000, with the exception of rainbow trout, which were incubated at 5°C in 2001 (Holm
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2002; Holm et al. 2005). No significant differences among sites were observed for percent
fertilization and mortality. Percentages of embryonic deformities and edema were significantly
different among streams, but rates of incidence of deformities at Wampus Creek, one of the
reference streams, were often similar to or higher than deformities at streams with elevated
concentrations of selenium (see Holm summary in Appendix C). The measurement of selenium
in the otolith layers of rainbow trout collected in this watershed showed low selenium exposure
in the fish's early life and a higher exposure to selenium during the fish's adult years (Palace et
al. 2007), suggesting that individuals that reach adulthood do not tend to start their lives in
elevated exposure streams even though they may reside there later.
Estimates of effect concentrations for the combined data (i.e., 2000 through 2002) were
derived with a fitted logistic equation (TRAP). Proportion of skeletal deformities in rainbow
trout embryos was the most sensitive endpoint with an EC 10 of 21.1 mg Se/kg dw and an EC20 of
28.4 mg Se/kg dw. Holm (2002) and Holm et al. (2005) reported egg selenium concentrations in
wet weight. Egg wet weight was converted to dry weight assuming 61.2% moisture in rainbow
trout eggs (Seilor and Skorupa 2001).
Oncorhynchus clarki lewisi (westslope cutthroat trout)
In a field study similar to those conducted by Holm et al. (2005) and Kennedy et al.
(2000), Rudolph et al. (2008) collected eggs from Westslope cutthroat trout from Clode Pond
(exposed site) and O'Rourke Lake (reference site). Clode Pond is on the property of Fording
River Coal Operations in Southeast British Columbia with reported selenium concentrations of
93 |ig/L. O'Rourke Lake is an isolated water body into which Westslope cutthroat trout were
stocked in 1985, 1989 and 1992 and has selenium levels reported as <1 |ig/L. Eggs with the four
highest Se concentrations (86.3 to 140 mg/kg dw) collected from Clode Pond fish died before
reaching the laboratory. Of those eggs from both ponds that survived, there was no correlation
between egg selenium concentration and frequency of deformity or edema in the fry. The percent
of alevins (post hatch to swim-up stage) that died was related to the selenium concentration in
the eggs; the TRAP estimates of the ECio and EC20 for survival in the eggs are 24.11 mg Se/kg
dw and 28.73 mg Se/kg dw, respectively.
As a follow-up to the study by Rudolph et al. (2008), Nautilus Environmental (2011)
conducted a more extensive study with Westslope cutthroat trout at the same site. Adult
Westslope cutthroat trout were collected from lentic and lotic environments from locations near
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the mining operations. The lentic fish were primarily captured in Clode Pond, a settling area used
to improve water quality of the mining discharge. Lotic fish were collected from the Fording
River and its tributaries near the mining operation. Reference females were obtained from
Connor Lake which is located within the watershed but not exposed to mining discharges. The
researchers reared fertilized eggs from the caught females in the laboratory until they reached
swim-up fry stage. A subset of fry surviving at swim-up were reared for an additional 28 days.
The most sensitive endpoint was larval survival at swim-up with an ECio determined by TRAP
of 24.02 mg/kg egg dw. This result is very similar to the ECio of 24.11 mg/kg egg dw
determined for the data generated by Rudolph et al. (2008). See Appendix C for more details on
the Nautilus Environmental (2011) study.
Based on these two studies, for the ECio level of effects, the SMCV for cutthroat trout,
Oncorhynchus clarki, is 24.06 mg Se/kg dw in eggs derived from Rudolph et al. (2008) and
Nautilus Environmental (2011) (24.11 and 24.02 mg Se/kg dw, respectively).
Salvelinus fontinalis (brook trout)
This data was not used directly in the criterion calculations See Section 7.1 for discussion
of the available data.
Salvelinus malma (Dolly Varden)
Golder (2009) collected adult Dolly Varden from a reference site and two sites
downstream from the Kemess Mine in northern British Columbia, one with a high and one with a
moderate selenium exposure in the fall of 2008. Fertilized eggs were taken to the laboratory
where they were monitored for survival and deformities until 90% of the larvae reached swim-
up, approximately 5 months after fertilization. Alevin mortality was <1% in the treatments
collected from the exposed sites and not considered an effect. The prevalence of deformities
increased sharply after the selenium egg concentration exceeded 50 mg/kg dw (Appendix C).
The proportion of Dolly Varden larvae without any type of deformity (skeletal, craniofacial, and
finfold as well as edema) as a function of the log of the selenium concentration in the eggs using
TRAP produced an ECio value of 56.22 mg Se/kg dw and an EC20 value of 60.12 mg Se/kg dw.
Salmo trutta (brown trout)
Formation Environmental (2011) collected adult female and male brown trout from sites
with low and high selenium exposure in the proximity of a phosphate mine located in
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Southeastern Idaho in November 2007. Eggs were collected from 26 gravid females across three
sampling locations, fertilized with milt collected from several males from the same site and taken
to the laboratory for hatching and observation of larval malformations and survival. In addition
to the field collected fish, fertilized eggs of twelve females from two separate hatcheries were
used in the study. The study had two phases, hatch-to-swim up, and swim-up to 15 days post
swim-up. There are two experimental complications that affect the interpretation of these data:
(a) elevated deformity rates among the offspring that were to serve as hatchery-originated
method controls (very low selenium exposure) and among some of the low exposure field-
collected organisms, and (b) the accidental loss of a number of individuals from several
treatments during the 15-day post swim up portion of the test. This document's analysis of the
revised counts from AECOM (2012) builds upon but supersedes EPA's 2012 analysis (Taulbee
et al. 2012), peer reviewed by ERG (2012). For the full test, hatch through 15-days post swim up,
combining wild and hatchery fish, there are six ECios corresponding to survival, free of
deformity, and combined survival and free of deformity, all for both the lab accident worst case
assumption (fry lost in the lab accident during the 15-day portion of the study were assumed to
have been dead or deformed) and the optimistic assumption (fry lost had the same rates of
mortality and deformity as those not lost). These six ECios, shown in Section 7.1.4, fall in the
range 15.91 - 21.16 mg/kg egg dw. Appendix C presents details of the study. The chronic value
selected for the study was the low ECio of 15.91 mg/kg egg dw, for total deformities, assuming
the worst case scenario for fry lost in the lab accident.
Salmonidae SMCV and GMCV Summary
For the ECio level of effects, the SMCV for cutthroat trout, Oncorhynchus clarki, is 24.06
mg Se/kg dw in eggs derived from Rudolph et al. (2008) and Nautilus Environmental (2011)
(24.11 and 24.02 mg Se/kg dw respectively). The GMCV for the genus Oncorhynchus is 22.53
mg Se/kg dw in eggs, derived from the 21.1 mg Se/kg dw ECio from the combined Holm (2002)
and Holm et al. (2005) rainbow trout data, and the above mean of the Rudolph et al. (2008) and
Nautilus Environmental (2011) Westslope cutthroat trout studies (24.06 mg Se/kg dw). The
GMCV for the genus Salvelinus is the ECio value of 56.22 mg Se/kg dw for Dolly Varden (S.
malmd) from the Golder (2009) study, and the GMCV for the genus Salmo is the ECio value of
15.91 mg Se/kg dw for brown trout (S. trutta) from the Formation Environmental (2011) study.
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Poeciliidae
Data are available for two species in this family. These studies are not represented in
Table 5 because these species are live-bearing rather than egg-laying species, but the relative
tolerance of these species is accounted for in derivation of the criterion.
Gambusia holbrooki (eastern mosquitofish)
Staub et al. (2004) collected male and gravid female eastern mosquitofish from a
contaminated ash basin and a reference pond in July 1999. Male fish were used for measuring
standard metabolic rate and the reproductive endpoints, brood size and percent viability of live
offspring at parturition were measured using the live-bearing females. Standard metabolic rates
of males, brood size of females, and offspring viability were not significantly different between
sites. Average concentrations of selenium in females were 11.85 and 0.61 mg/kg dw in the
contaminated ash basin and reference sites, respectively. The chronic value in whole body tissue
is >11.85 mg Se/kg dw whole-body (Appendix C). In a community of equally exposed fish taxa
(fish taxa having whole body tissue concentrations >11.85 mg Se/kg dw), the median egg-ovary
concentration among egg-laying fish would be expected to be 1.71 higher, or >20.26 mg Se/kg
dw.
Gambusia a/finis (western mosquitofish)
Western mosquitofish were collected in June and July 2001 from two sites in the
grassland water district, Merced County, California that were contaminated with selenium and
two reference sites in the same area with relatively low selenium exposure (Saiki et al. 2004).
Seventeen to 20 gravid females (mosquitofish are live-bearers) from each site were held in the
laboratory for two weeks to quantify live and dead births and to make other measurements. Live
and dead fry were visually examined under low magnification with a binocular microscope for
evidence of external abnormalities (teratogenic symptoms such as spinal curvature, missing or
deformed fins, eyes and mouths and edema). The percentage of live births was high at both
selenium-contaminated sites (96.6 to 99.9%) and reference sites (98.8 to 99.2%). There were no
obvious anomalies (e.g., deformities, edema) observed during the study. The concentration of
selenium in 4 postpartum females from the site with the highest selenium concentration ranged
from 13.0 to 17.5 mg Se/kg dw (geometric mean of the high and low is 15.1 mg Se/kg dw). The
chronic value in whole body tissue is >15.1 mg Se/kg dw (Appendix C). Similar to Staub et al.
(2004), this value can be converted to egg-ovary concentrations that would be expected in
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accompanying egg-laying fish, by multiplying by the median fish egg-ovary to whole-body
concentration ratio, 1.71. This yields >25.82 mg Se/kg dw equivalent egg-ovary.
Gambusia, which have been reported to be tolerant to selenium contamination, are often
one of the few remaining species at sites with high levels of selenium contamination (Cherry et
al. 1976; Lemly 1985a; Saiki et al. 2004; Young et al. 2010, Janz et al. 2010). The two studies
discussed above support this observation with a GMCV of >13.4 mg Se/kg dw in whole body
tissue, combining these "greater than" values as described in Section 7.1.1. It may be concluded
that this genus is not among the most sensitive to selenium.
Cyprinodontidae
Cyprinodon macularius (desert pupfish)
Besser et al. (2012), using a diet of oligochaete Lumbriculus that had fed on selenized
yeast, exposed desert pupfish to six levels of dietary and waterborne selenium. Five-week old
juveniles (Fo) were exposed for 85 days, during which time survival and growth were measured.
Upon reaching maturity at the end of this time, the 60-day reproductive study was begun, during
which FI eggs were collected, counted, and further tested for percent hatch, survival, growth, and
deformities. The authors observed no significant differences in pupfish survival, growth, total
egg production, hatch, or deformities among treatments. Although the authors noted a potential
interaction between the timing of egg production and treatment, a comprehensive re-analysis of
this data, described in Appendix C, indicated that the phenomenon was neither statistically nor
biologically significant. It is concluded that the egg concentration, 27 mg Se/kg (dw), for the
test's highest treatment was not sufficiently high to define a concentration-response curve.
Although desert pupfish is thus not among the most sensitive species, the slightly reduced
survival observed at 27 mg Se/kg suggests that the ECio may be close to that concentration, as
also noted by the authors.
Centrarchidae
Lepomis macrochirus (bluegill sunfish)
In a laboratory study, Doroshov et al. (1992) exposed adult bluegill for 140 days to three
dietary concentrations of seleno-L-methionine added to trout chow. Near the end of the
exposure, ripe females were induced to ovulate and ova were fertilized in vitro with milt stripped
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from males. Fertilized eggs were sampled for fertilization success and selenium content. They
were also used in two tests, (a) a larval development study during the first 5 days after hatching,
and (b) a 30-day embryo-larval test. In the 5-day larval test, the average proportion of larvae with
edema was 0% at an egg concentration of 8.33 mg Se/kg (the first treatment), 5% at an egg
concentration of 19.46 mg Se/kg dw (the second treatment), and 95% at an egg concentration of
38.39 mg Se/kg dw (the highest treatment). The latter two were statistically different from the
control (0% edema). All edematous larvae died in the high treatment. In the 30-day larval
survival test, statistical difference from the control was only found in the highest test treatment
for survival and growth (length and weight) measurements. The ECio calculated with TRAP for
the incidence of edema in the 5-day larval bioassay is 20.05 mg Se/kg dw in eggs.
A similar study with similar results was done by Coyle et al. (1993) in which two year
old pond-reared bluegill sunfish were exposed in the laboratory and fed (twice daily ad libitum)
Oregon moist™ pellets containing increasing concentrations of seleno-L-methionine. Water
concentrations were nominal 10 jag Se/L. The fish were grown under these test conditions for
140 days. Spawning frequency, fecundity, and percentage hatch were monitored after 60 days
when spawning began to occur. There was no effect of the combination of the highest dietary
selenium concentration (33.3 mg Se/kg dw) on adult growth, condition factor, gonadal somatic
index, or the various reproductive endpoints (Appendix C). The survival of newly hatched
larvae, however, was markedly reduced; only about 7 percent survived to 5 days post-hatch in
the high dietary treatment. The TRAP-calculated ECio value for larval survival is 24.55 mg
Se/kg dw in eggs.
Hermanutz et al. (1992), and Hermanutz et al. (1996) exposed bluegill sunfish to sodium
selenite spiked into artificial streams (nominal test concentrations: 0, 2.5, 10, and 30 jag Se/L)
which entered the food web, thus providing a simulated field-type exposure (waterborne and
dietary selenium exposure). In an effort originally intended to improve the rigor of the statistical
analysis of the Hermanutz et al. (1996) data, Tao et al. (1999) re-examined the raw data records
and made corrections to the counts. This criteria document considers the Hermanutz et al. (1992)
data and the Tao et al. (1999) re-examination of Hermanutz et al. (1996).
These data come from a series of three studies lasting from 8 to 11 months, conducted
over a 3-year period. All three studies began with exposure to adult bluegill sunfish in the fall,
and respective studies ended in the summer of the following year. Winter temperatures averaged
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4.6, 4.1 and 4.5°C and spawning months (June-July) averaged 26.4, 23.9 and 22.4°C,
respectively for Studies I, II and III. Spawning activity was monitored in the stream, and embryo
and larval observations were made in situ and from fertilized eggs taken from the streams and
incubated within egg cups in the laboratory. None of the adult bluegill exposed to the highest
concentration of selenium in the water (Study I, mean measured concentration equal to 29.4
Hg/L) survived the entire exposure period (although a few did survive to spawn). Embryo-larval
effects were observed in the selenium-dosed streams in both Study I and Study II. The incidence
of edema, lordosis, hemorrhage and larval survival in the one stream concentration common to
both Study I and II, 10 |ig/L, ranged from 80 to 100 percent, 5 to 18 percent, 27 to 56 percent,
and 29 to 58 percent, respectively (combined egg cup and nest observations). Edema, lordosis,
and hemorrhage in the lowest stream concentration in Study II, 2.5 |ig/L, ranged from 0 to 4
percent, 0 to 25 percent, and 3.6 to 75 percent, respectively (combined egg cup and nest
observations); larval survival was 71.6 percent (72 and 75 percent in the control streams). See
Hermanutz 1996 and 1992 in Appendix C for more detail. The above effects were not observed
in larvae from fish that were not exposed to elevated concentrations of selenium (control
treatment). The mean concentrations of selenium in bluegill ovaries ranged from 0.8 to 2.5
mg/kg dw in the control 7.6 to 10.9 mg/kg dw in the 2.5 |ig/L treatment, and 17.7 to 30.0 mg/kg
dw in the 10 jag Se/L treatment (values represent both Study I and II for the control and 10 |ig/L
treatments).
The embryo-larval data for continually exposed streams of Studies I and II of this
experiment were combined and analyzed in response to measured selenium concentration in the
maternal ovaries (mg/kg dw) using TRAP. In the recovering streams of Study II and Study III no
effects were observed at concentrations that had previously caused effects in the continually
exposed streams. Recovering streams were therefore excluded from this criteria document's
analysis because they do not reflect the type of system that water quality criteria are most
commonly applied to, those receiving existing waterborne pollutant discharges. That is, Study III
consisted of the addition of new adult bluegill to the same streams that had received the 2.5, 10,
and 30 ug/L sodium selenite during previous studies, but with all continued external dosing of
selenite halted. The adult bluegills exposed only to dietary selenium present in the Study III food
web accumulated selenium to levels very near to those accumulated during Study II in which
aqueous selenium was also present demonstrating the importance of diet on selenium
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accumulation. There were no effects (no effect on larval survival, 0 percent deformities, 0
percent hemorrhaging), on the bluegill progeny in Study III even from fish that accumulated
26.04 and 10.17 mg/kg dw in the recovering 10 ug/L streams, and 15.92 mg/kg dw in the
recovering 30 ug/L stream. The absence of effects at high tissue levels in the recovering streams
of Study III provides experimental corroboration for the field observations of biological recovery
in Belews Lake and Hyco Reservoir after selenium loads were reduced but while tissue
concentrations remained relatively high (Lemly 1997a; Crutchfield 2000; Finley and Garrett
2007). Because Study III involved new (naive) fish added to previously contaminated streams,
neither acclimation nor adaptation would seem to explain this phenomenon. Overall, the
implication is that for some period of time, recovering systems might possibly exceed tissue
criteria concentrations even though the effects of selenium have been mitigated.
Of the three endpoints that attained model convergence when analyzed by TRAP (%
edema, % lordosis, and % hemorrhage), % edema relative to ovary selenium concentration was
the most sensitive yielding an ECio of 12.68 mg Se/kg and an EC20 of 13.66 mg Se/kg dw ovary.
The data for % lordosis indicate that it is a less sensitive endpoint. The data for % hemorrhage
showed no conclusive concentration-response relationship: responses at concentrations 7.6 and
50.5 mg Se/kg dw varied widely without any relationship to selenium concentration, precluding
its use for estimating an ECio. The ECio value of 12.68 mg/kg Se dw (larval edema in response
to Se concentration in the parental ovaries) is considered an environmentally conservative
chronic value for this bluegill study. (See Appendix C for more discussion of this study, and
Section 7.1.5 for more discussion of this chronic value).
The SMCV for bluegill reproductive endpoints based on ECio values is 18.41 mg Se/kg
dw in egg/ovary, based on the ECio values of Doroshov et al. (1992), Coyle et al. (1993), and
Hermanutz et al. (1992, and 1996 as corrected by Tao et al. 1999)
Micropterus salmoides (largemouth bass)
A laboratory study was conducted by Carolina Power & Light (1997) in which adult
largemouth bass obtained from a commercial supplier were fed an artificial diet spiked with a
gradient of selenomethionine for several months. Similar exposure studies were conducted in
1995 and 1996 resulting in successful spawning of adults. Approximately 100 eggs from each
spawn were monitored for mortality and deformities up to the larval swim-up stage. The authors
combined survival and deformities into a single metric (i.e., survival as normal offspring). The
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average concentration of selenium in the ovaries ranged from 3.1 mg/kg dw in the control to 77.6
mg/kg dw in the high dietary treatment (53.1 mg/kg dw). The percent survival of larval
largemouth bass as a function of the selenium concentration in the parental ovary using TRAP
produced an ECio of 20.35 mg/kg dw and an £€20 of 23.60 mg/kg dw (Appendix C).
4.1.2 Summary of Acceptable Studies of Fish Reproductive Effects
Table 5 summarizes the effect concentrations obtained from reproductive studies with
fish.
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Table 5. Maternal Transfer Reproductive Toxicity Studies.
Species
Pimephales promelas
fathead minnow
Esox lucius
northern pike
Oncorhynchus mykiss
rainbow trout
Oncorhynchus clarki
lewisi
Westslope cutthroat
trout
Oncorhynchus clarki
lewisi
Westslope cutthroat
trout
Salvelinus malma
Dolly Varden
Reference
Schultz and
Hermanutz 1990
Muscatello et al.
2006
Holm 2002;
Holm et al.
2003; Holm et
al. 2005
Rudolph et al.
2008
Nautilus
Environmental
2011
Golder 2009
Exposure route
dietary and
waterborne
(mesocosm:
Monticello)
dietary and
waterborne (field:
Saskatoon, Sask.)
dietary and
waterborne
(field: Luscar
River, Alberta)
dietary and
waterborne (field:
Clode Pond, BC)
dietary and
waterborne (field:
Clode Pond &
Fording River,
BC)
dietary and
waterborne
(field: Kemess
Mine NW British
Columbia)
Toxicological
endpoint
LOEC for larval
edema and lordosis
EC24 larval
deformities
EC 10 for skeletal
deformities
ECio for alevin
mortality
EC 10 for survival
at swim-up
EC 10 for total
deformities
Chronic value,
mg/kg dwa
<23.85 Ob
34.00 E
21.1Eb
24.11 E
24.02 E
56.22 E
SMCV
mg/kg dw
<23.85 O
34.00 E
21. IE
24.06 E
56.22 E
GMCV
mg/kg dw
<23.85 O
34.00 E
22.53 E
56.22 E
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52
Draft Document
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Species
Salmo trutta
brown trout
Cyprinodon
macularius desert
pupfish
Lepomis macrochirus
bluegill
Lepomis macrochirus
bluegill
Lepomis macrochirus
bluegill
Micropterus salmoides
Largemouth bass
Reference
Formation
Environmental
2011
Besser et al.
2012
Doroshov et al.
1992
Coyle et al.
1993
Hermanutz et al.
1992
Hermanutz et al.
1996
Carolina Power
& Light 1997
Exposure route
dietary and
waterborne (field:
Lower Sage Creek
& Crow Creek,
ID)
dietary and
waterborne (lab)
dietary
(lab)
dietary and
waterborne (lab)
dietary and
waterborne
(mesocosm:
Monticello)
dietary (lab)
Toxicological
endpoint
EC 10 for larval
survival
Estimated ECio for
offspring survival
ECio larval edema
EC 10 for larval
survival
ECio for larval
edema
ECio for larval
mortality &
deformity
Chronic value,
mg/kg dwa
15.91E
27 E
20.05 E
24.55 E
12.68 Ob
20.35 O
SMCV
mg/kg dw
15.91 E
27 E
18.41E
20.35 O
GMCV
mg/kg dw
15.91 E
27 E
18.41E
20.35 O
a All chronic values reported in this table are based on the measured concentration of selenium in egg/ovary tissues.
E - concentration reported in egg; O - concentration reported in ovary
b Tissue value converted from ww to dw. See Appendix C for conversion factors.
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Table 6a presents the Genus Mean Chronic Values from acceptable studies of
reproductive effects in fish.
Table 6a. Ranked Genus Mean Chronic Values for Fish Reproductive Effects.
Rank
9
8
7
6
5
4
O
2
1
GMCV
(mg Se/kg dw EO)
56.22
<34
27
>25.82estim. EO*
(> 15.1 meas. WB)
<23.85
22.53
20.35
18.41
15.91
Species
Dolly Varden,
Salvelinus malma
Northern pike,
Esox lucius
Desert pupfish,
Cyprinodon macularius
Eastern mosquitofish,
Gambusia holbrooki
Western mosquitofish,
Gambusia affmis
Fathead minnow,
Pimephales promelas
Cutthroat trout,
Oncorhynchus clarki
Rainbow trout,
Oncorhynchus mykiss
Largemouth bass,
Micropterus salmoides
Bluegill sunfish,
Lepomis macrochirus
Brown trout,
Salmo trutta
SMCV
(mg Se/kg dw EO)
56.22
<34
27
> 20.26 estim. EO*
(>1 1.85 meas. WB)
> 25. 82 estim. EO*
(> 15.1 meas. WB)
<23.85
24.06
21.1
20.35
18.41
15.91
*For mosquitofish, a live bearer, the egg-ovary concentrations is that estimated to be
typical for other fish species in an assemblage sharing the same exposure. Combining
of its two "greater than" SMCVs into its GMCV follows the principles of Section
7.1.1.
4.1.3 Invertebrate Chronic Effects
Below is a brief synopsis of the experimental design of the available invertebrate chronic
toxicity tests, and the resulting chronic values.
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Brachionus calyciflorus (rotifer)
Dobbs et al. (1996) exposed Brachionus calyciflorus to selenate in natural creek water for
25 days in a three-trophic level food chain test system. This is one of two laboratory-based
experiments (also see Bennett et al. 1986) that involved exposing algae to selenium (in this case
as sodium selenate) in water, and subsequently feeding the algae to rotifers which were in turn
fed to fish (fathead minnows). In the Dobbs et al. (1996) study, the rotifers and fish were
exposed to the same concentrations of sodium selenate in the water as the algae, but consumed
selenium bioaccumulated in next lower trophic level. Rotifers did not grow well at
concentrations exceeding 108.1 jig Se/L in water, and the population survived only 6 days at
selenium concentrations equal to or greater than 202.4 jig Se/L in the water (40 jig Se/g dw in
the algae). Regression analysis of untransformed growth data (dry weight) determined 4 day
post-test initiation resulted in a calculated ECio of 37.84 jig Se/g dw tissue.
Lumbriculus variegatus (oligochaete, blackworm)
Although not intended to be a definitive toxicity study for blackworms, Besser et al.
(2006) evaluated the bioaccumulation and toxicity of selenized yeast to the oligochate,
Lumbriculus variegatus, which was intended to be used for dietary exposure in subsequent
studies with the endangered desert pupfish, Cyprinidon macularius. Oligochaetes fed selenized-
yeast yeast diets diluted with nutritional yeast (54 to 210 mg Se/kg) had stable or increasing
biomass and accumulated Se concentrations as high as 140 mg/kg dw. The oligochaetes fed the
undiluted selenized-yeast (826 ug/g Se dry wt.) showed reduced biomass. The effect level is
considered >140 mg Se/kg dw.
^^
Centroptilum triangulifer (mayfly)
Mayfly larvae (Centroptilum triangulifer) were exposed to dietary selenium contained in
natural periphyton biofilms to eclosion (emergence) (Conley et al. 2009). The periphyton fed to
the mayfly larvae were exposed to dissolved selenite (radiolabeled 75Se) in November 2008 (12.6
and 13.9 |ig/L) and in January 2009 ( 2.4, 2.4, 4.9, 10.3, and 10.7 |ig/L). Periphyton
bioconcentrated Se an average of 1113-fold over the different aqueous selenium concentrations
(see table below). Twenty 4 to 6-day old mayfly larvae were exposed for 4.5 to 6 weeks to each
of the periphyton diets until the larvae eclosed to subimagos (final pre-adult winged stage). The
subimagos were allowed to emerge to the adult imago stage which deposited their egg masses in
Petri dishes. Selenium was measured in postpartum adults along with their dry weights and
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clutch size.
Selenium increased in concentration from periphyton to the adult mayflies (trophic
transfer factor) an average of 2.2-fold. The authors observed a reduction in fecundity with diets
containing more than 11 mg Se/kg dw, which is considered the dietary threshold for this study.
Using the trophic transfer factor of 2.2, the periphyton selenium concentration of 11 mg/kg dw
translates to an adult mayfly selenium concentration of 24.2 mg/kg dw.
4.1.4 Summary of Relevant Invertebrate Tests
The available measured invertebrate whole-body effect concentrations and their
translation to fish egg-ovary concentrations are shown in Table 6b. Because the intent of this
assessment is to derive a concentration expressed in terms offish tissue, effect concentrations
expressed as in terms of invertebrate tissue need to be translated across media in order to
compare invertebrate effect concentrations to egg-ovary concentrations that would occur in a fish
assemblage that would accompany the invertebrates. That is, using the bioaccumulation
modeling approach of Section 4.2, invertebrate whole-body effect concentrations have been
translated to fish egg-ovary concentrations using (a) the median trophic transfer factor of 1.27
from Table 10 and (b) the median whole-body to egg-ovary conversion factor of 1.71 from Table
11. This yields a combined conversion factor of 1.27 x 1.71 = 2.17. Mean whole body chronic
values ranging from 24.2 mg/kg for the mayfly (Centroptilum traingulifer) to greater than 100
mg/kg for the oligochaete (Lumbrilicus variegatus),which is approximately 3 to 12 times higher
than the fish whole body criterion.
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Table 6b. Ranked Invertebrate Whole-Body Chronic Values with Translation to
Expected Accompanying Fish Egg-Ovary Concentrations.
SMCV & GMCV
as measured
(mg Se/kg dw WB)
>100
37.84
24.2
SMCV & GMCV as estimated EO
concentration in an accompanying
fish assemblage
(mg Se/kg dw EO)
>217
82.11
52.6
Species
Oligochaete,black
Lumbriculus variegatus
Rotifer,
Brachionus calyciflorus
Mayfly,
Centroptilum triangulifer
4.1.5 Derivation of Tissue Criterion Element Concentrations
Data used to derive the final chronic value were differentiated based on the effect
(reproductive and non-reproductive effects). Acceptable chronic toxicity data on fish
reproductive effects are available for 9 fish genera. Acceptable chronic toxicity data on non-
reproductive effects are available for 7 fish genera and 3 invertebrate genera. The fish non-
reproductive effects data were not used to calculate criteria because they were more variable and
less reproducible than the data on reproductive effects. The SSD is predominantly populated with
data on fish species because field evidence demonstrated that fish communities were affected in
situations having no observable change in the accompanying diverse array of invertebrate
communities. As a result, decades of aquatic toxicity research have focused primarily on fish.
The studies that have been done with invertebrates (Table 6c, Section 4.1.2) have shown them to
be more tolerant than most of the tested fish species. While potentially sensitive due to
physiologic similarities to fish, amphibian effects clearly attributable to selenium are largely
unknown (Unrine et al. 2007; Hopkins et al. 2000; Janz et al. 2010). Hopkins et al. (2000)
reported that amphibian larvae at sites receiving coal combustion wastes appear to efficiently
accumulate selenium in their tissues and possibly due to selenium have exhibited axial
malformations
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Fish Egg-Ovary Concentration
The lowest four GMCVs from Table 6a are shown below in Table 6c.
Table 6c. Four lowest Genus Mean Chronic Values for Fish Reproductive Effects.
Relative Sensitivity
Rank
4
O
2
1
Genus
Oncorhynchus
Micropterus
Lepomis
Salmo
GMCV
(mg Se/kg dw egg-ovary)
22.53
20.35
18.41
15.91
With N=14 GMCVs, the 5* percentile projection yields an egg/ovary criterion of 15.2 mg
Se/kg dw egg/ovary, lower than the most sensitive fish species tested, brown trout (Salmo trutta).
The egg/ovary criterion element concentration is compared to the distribution of egg/ovary
chronic values in Figure 5.
M 7
~O
KM TCC
_™ ZDO
01
l/l
bO 1 9Q
u
C CA
3
(0 09
i
^ 16
.c
iZ
» Fish
• Mosquitofish
A Inverteb.
___ EOFCV
A *
A
0 20 40 60 80 100
Percentile
Figure 5. Distribution of (a) reproductive-effect GMCVs for fish measured as egg-ovary
concentrations (diamond markers), (b) the reproductive-effect value for mosquito fish
(square marker), a live-bearer measured as adult whole-body but translated to an
equivalent egg-ovary concentration using the median conversion factor 1.71, and (c)
invertebrate effect concentrations (triangle markers) measured as whole-body but
translated to the equivalent fish egg-ovary concentrations expected in an accompanying
fish assemblage, through the median trophic transfer factor of 1.27 from Table 10 and the
median egg-ovary conversion factor of 1.71 from Table 11
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Fish Whole-Body Criterion Element Concentration
Using the egg-ovary to whole-body conversion factors of the bioaccumulation modeling
approach presented subsequently in Section 4.2, Table 7a shows the conversion of reproductive-
effect egg-ovary concentrations to whole-body concentrations. It can be seen that for some
species, the conversion was done in a single step, using a measured egg-ovary (EO) to whole-
body (WB) ratio specific to the taxon. But for other species, it was done in two steps, first
converting to muscle (M) and then applying a generic M/WB ratio.
Table 7a. Tested Reproductive-Effect Egg-Ovary (EO) Concentrations Converted to
Whole-Body (WB) Concentrations.
Taxon
Salvelinus
Esox
Cyprinodon
Pimephales
O. mykiss
O. clarkii
Onchyrhynchus
Micropterus
Lepomis
Salmo
EO
Chronic
Value
56.22
34.00
27.00
23.85
21.10
24.06
22.53
20.35
18.41
15.91
EO/WB
Ratio
1.61
2.39
1.21
2.00
2.44
2.30
2.37
1.45
2.13
1.45
Calculated
WB Repro
Chronic
Value
34.90
14.23
22.31
11.94
8.64
10.46
9.51
14.03
8.63
10.97
Basis for EO/WB Ratio
(from Appendix B)
Median Dolly Varden EO/M (1.264)
x median fish M/WB (1.274)
Median northern pike EO/M (1.875)
x median fish M/WB (1.274)
Median desert pupfish EO/WB
Median Cyprinidae EO/WB
Median rainbow trout EO/M (1.916)
x median fish M/WB (1.274)
Median cutthroat trout EO/M (1 .805)
x median fish M/WB (1.274)
Using geomean of species ratios
yields geomean of SMCVs
Median Centrarchidae EO/WB
Median bluegill EO/WB
Median brown trout EO/WB
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Table 7b. The lowest four reproductive-effect whole-body GMCVs.
Relative Sensitivity
Rank
4
3
2
1
Genus
Pimephales
Salmo
Oncorhynchus
Lepomis
GMCV
(mg Se/kg dw whole-body)
11.94
10.97
9.51
8.63
Because the factors used to convert egg-ovary to whole-body concentrations vary across
species, the whole-body rankings differ from the egg-ovary rankings. With N=14 GMCVs, the
5* percentile projection yields a whole body criterion of 8.13 mg Se/kg dw whole-body, slightly
lower than the most sensitive fish species tested, bluegill (Leopmis machrochirus) The fish
whole body criterion is compared to the distribution offish whole body chronic values in Figure
6.
TCC nn
mo -i -) o nn
*oT
en
mo CA nn
u
c 39 nn
0 32.00
n ifi nn
(§ ib-uu
«
2 snn
!§ Ann
(
# Fish
A Inverteb.
___ WBFCV
X Mosquitofish
A
•
.*****
) 20 40 60 80 100
Percentile
Figure 6. Distribution of (a) reproductive-effect GMCVs for fish, measured as egg-ovary
concentrations but converted to whole-body concentrations as shown in Table 7, (b) the
reproductive-effect value for mosquito fish, a live-bearer already measured as adult whole-
body, and (c) invertebrate effect concentrations measured as whole-body and translated to
equivalent fish whole-body concentrations through the median trophic transfer factor of
1.27 from Appendix B.
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Fish Muscle Criterion Element Concentration
Using the egg-ovary to muscle conversion factors of the bioaccumulation modeling
approach presented subsequently in Section 4.2, Table 8a shows the conversion of reproductive-
effect egg-ovary concentrations to whole-body concentrations. For all but Cyprinodon (desert
pupfish), the conversion could be done in a single step, applying an EO/M ratio specific to the
taxon.
Table 8a. Tested Reproductive-Effect Egg-Ovary (EO) Concentrations Converted to
Muscle (M) Concentrations.
Taxon
Salvelinus
Esox
Cyprinodon
Pimephales
O. mykiss
O. clarkii
Onchyrhynchus
Micropterus
Lepomis
Salmo
EO
Chronic
Value
56.22
34
27
23.85
21.1
24.1
22.56
20.35
18.41
15.91
EO/M
Ratio
1.264
1.875
0.950
1.590
1.916
1.805
1.860
1.187
1.375
1.135
Calculated
Muscle Repro
Chronic
Value
44.48
18.13
28.42
15.00
11.01
13.35
12.124
17.15
13.39
14.02
Basis for EO/M Ratio
(from Appendix B)
Median Dolly Varden EO/M (1.264)
Median northern pike EO/M (1.875)
Median desert pupfish EO/WB divided
by median fish M/WB (1.274)
Median Cyprinidae EO/M (1.590)
Median rainbow trout EO/M (1.916)
Median cutthroat trout EO/M (1.805)
Using geomean of species ratios yields
geomean of SMCVs
Median Micropterus EO/M (1 . 1 87)
Median bluegill EO/M (1.375)
Median brown trout EO/M (1.135)
Table 8b. The lowest four reproductive-effect fish muscle GMCVs.
Relative Sensitivity
Rank
4
3
2
1
Genus
Pimephales
Salmo
Lepomis
Oncorhynchus
GMCV
(mg Se/kg dw muscle)
15.00
14.02
13.39
12.12
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Because the factors used to convert egg-ovary to muscle concentrations vary across
species, the whole-body rankings differ from both from the egg-ovary rankings and the muscle
rankings. With N=14 GMCVs, the 5th percentile projection yields a muscle criterion of 11.8 mg
Se/kg dw muscle, lower than muscle value for the most sensitive fish genus tested, Oncorhyncus.
4.2 Chronic Water Column-based Selenium Criterion Element
4.2.1 Translation from Fish Tissue Concentration to Water-Column Concentration
The chronic water column selenium criterion element is derived by translating the egg-
ovary concentration to an equivalent water concentration. The EPA worked with the United
States Geological Survey to derive a translation equation that utilizes a mechanistic model of
bioaccumulation previously published in peer-reviewed scientific literature (Luoma et. al., 1992;
Wang et. al., 1996; Luoma and Fisher, 1997; Wang, 2001; Schlekat et al. 2002b; Luoma and
Rainbow 2005; Presser and Luoma 2006; Presser and Luoma 2010; Presser 2013). This model
quantifies bioaccumulation in animal tissues by assuming that net bioaccumulation is a balance
between assimilation efficiency from diet, ingestion rate, rate of direct uptake in dissolved forms,
loss rate, and growth rate. The basic model is given as:
Where:
[(ku*Cwater)+(AExIRxCfood)]
(ke+g)
tissue
ku
AE
IR
Cfood
(Equation 1)
= Concentration of metal in water (|ig/L)
= Average concentration of metal in all tissues at steady-state (|ig/g)
= Efflux rate (/d)
= Growth rate (/d)
= Uptake rate (L/g-d)
= Assimilation efficiency (%)
= Ingestion rate (g/g-d)
= Concentration in food (|ig/g)
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Simplifying the Bioaccumulation Model
Specific application to selenium bioaccumulation permits the simplification of Equation 1
in two ways. One simplification is removing the parameter representing growth rate (g), and the
other simplification is removing the parameter representing direct aqueous uptake (&„).
Growth Rate
The growth rate constant g is included in Equation 1 because the addition of body tissue
has the potential to dilute the concentration of bioaccumulative chemicals when expressed as
chemical mass per tissue mass. For very hydrophobic chemicals with low excretion rates such as
polychlorinated biphenyls, growth can be an important factor in bioaccumulation estimates
(Connolly and Pedersen 1988). However, Luoma and Rainbow (2005) suggests that for
selenium, growth rate is a relatively inconsequential parameter under most circumstances. Food
consumption is typically high during periods of high growth rate. Because food consumption is
the primary route of selenium uptake in aquatic organisms (Ohlendorf et al. 1986a, b; Saiki and
Lowe 1987; Presser and Ohlendorf 1987; Lemly 1985a; Luoma et al. 1992; Presser et al. 1994,
Chapman et al. 2010), high consumption rates of selenium-contaminated food may counteract
the selenium dilution that occurs with the addition of body tissue during periods of fast growth.
The EPA evaluated the effect of removing the parameter g in the Equation 1 by
performing a sensitivity analysis. The EPA analyzed a series of hypothetical tissue concentration
estimates using Equation 1 with g ranging between 0 (no growth) and 0.2/day (a relatively high
rate of growth). In one analysis, tissue concentrations of selenium were estimated using static
values of IR. In a second analysis, tissue concentrations of selenium were estimated using values
of IR that were adjusted for growth rate using a method similar to the approach used in a model
of organic chemical accumulation in aquatic food webs (Thomann et al. 1992). As expected,
estimates of selenium tissue concentrations were significantly reduced at progressively higher
growth rates when IR remained constant. However, selenium concentrations remained fairly
steady or slightly increased with progressively higher growth rates when IR was adjusted for the
bioenergetics of growth. This analysis supports the hypothesis that a higher//? (and consequently
greater rate of selenium ingestion) associated with the higher bioenergetic requirements of
rapidly growing young fish tends to oppose the dilution of selenium in their tissues due to
growth, whereas a lower IR (and consequently lower rate of selenium ingestion) associated with
the lower bioenergetic requirements of slower growing older fish tends to oppose the
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bioconcentration of selenium in their tissues. The EPA concludes from this analysis that omitting
the growth rate parameter g is an appropriate simplification of Equation 1. A more detailed
description of this sensitivity analysis is provided in Appendix G.
Uptake Rate
The uptake rate constant ku is included in Equation 1 to account for direct absorption of
bioaccumulative chemicals in the dissolved phase. However, dietary intake of selenium is the
dominant source of exposure, suggesting that ku may also be relatively inconsequential for
selenium accumulation (Luoma and Rainbow 2005). Because aqueous uptake of selenium makes
up a small percentage of bioaccumulated selenium (Fowler and Benayoun 1976; Luoma et. al.,
1992; Roditi and Fisher 1999; Wang and Fisher 1999; Wang 2002; Schlekat et. al., 2004; Lee et.
al., 2006), Presser and Luoma (2010a, 2010b, 2013) deemed removal of ku from Equation 1 as an
acceptable simplification.
The EPA evaluated the effect of removing the parameter ku in the Equation 1 by
performing a sensitivity analysis. The EPA analyzed a series of tissue concentration estimates
using Equation 1 and a realistic range of ku values for trophic level 2 and trophic level 3
organisms. The analysis suggests that approximately 75% of selenium exposure in trophic level 2
organisms (invertebrates) and over 90% of selenium exposure in trophic level 3 organisms
occurs through consumption of selenium-contaminated food. The EPA concluded that omitting
the aqueous uptake rate constant ku is an appropriate simplification of Equation 1. A more
detailed description of this sensitivity analysis is provided in Appendix G.
Derivation of the Translation Equation
Disregarding growth (g) and uptake of selenium dissolved in water (ku x Cwater), Equation
1 becomes:
= AExIRxCfood
tissu e i
or:
AExIR
( — \/ (
tissue i food
(Equation 2)
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Because application of the bioaccumulation model applies to a single species, the
AExIR
combination of species-specific physiological parameters expressed as — remains
AExIR
constant for the species. Thus the EPA defines the expression as a single species-
specific Trophic Transfer Function (TTF) given as:
TTF=AExIR j£
ke (Equation 3)
AExIR
Substituting TTF for in Equation 2 yields:
Ctlssue = TTF x Cfood (Equation 4)
The trophic level of the organisms considered can be denoted by superscripts given as:
tlssue
(Equations)
Ctjssue as defined here represents the steady-state proportional concentration of selenium in the
tissue of trophic level 2 organisms relative to the concentration of selenium in their food source.
Using the same rationale, the average concentration of selenium in the tissues of trophic
level 3 organisms can be expressed as the concentration of selenium in its food multiplied by a
TTF which is given as:
tissue
(Equations)
For trophic level 3 organisms that consume trophic level 2 organisms, Cfood = Ctissue
Thus:
ctissue = TTF -xCtissue (Equation 7)
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Substituting c^ue in Equation 7 with TTF x Cfood in Equation 5 yields
/^ (Equations)
Defining the term cri3 as the concentration of selenium in fish tissue, defining the term
° ^tissue ' °
C/oorf as the concentration of selenium in living and nonliving particulate material ingested by
invertebrates, and expressing the product of all TTF values as a single term results in the
equation:
CT
-------
possible to differentiate bioaccumulative potential for different predator species and food webs
by modeling different exposure scenarios. For example, where the fish species of interest is a
trophic level 4 predator that primarily consumes trophic level 3 fish, the term jjp0™?0^ can be
represented as the product of all TTF parameters that includes the additional trophic level given
as:
TTFcomposlte = TTF™ x TTFTL3 x TTFTL2 (Equation 10)
where:
TT 9
TTF = the trophic transfer function of trophic level 2 species
TT ?
TTF = the trophic transfer function of the trophic level 3 species
ic transfer function of the trophic level 4 species
TTFcompoS«e = the product of all trophic transfer functions
Similarly, the consumption of more than one species of organism at the same trophic
level can also be modeled by expressing the TTF at a particular trophic level as the weighted
average of the TTFs of all species consumed given as:
TTFTLx = £ (jTF^ x wt ) (Equation 1 1)
/
where:
TTF> = the trophic transfer function of the ith species at a particular trophic level
w; = the proportion of the i* species consumed
These concepts can be used to formulate an expression of j'j'p^0^0^6 to model selenium
bioaccumulation in ecosystems with different consumer species and food webs. Figure 7
describes four example food web scenarios and the formulation of jj^0^0^^ to model selenium
bioaccumulation in each of them.
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A) Three trophic levels (simple);
B) Four trophic levels (simple);
C) Three trophic levels (mix within trophic levels);
TTFcomposi" = TTFT" x [(TTF™2 x Wl}+(TTF™2 x w2}]
K
' *
TTF?
D) Three trophic levels (mix across trophic levels);
comp°"'e=(TTFT" x Wl)+ (TTF™ x TTFTI2 x w2)
w,
E) Four trohic levels (mix across trohic levels);
Figure 7. Example aquatic system scenarios and the derivation of the equation parameter
TTFcomposite.
This parameter quantitatively represents all dietary pathways of selenium exposure for a
particular fish species within an aquatic system. The parameter is derived from species-specific
TTF values representing the food web characteristics of the aquatic system. wt, proportion of
species consumed. See text for further explanation.
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Because the EPA's objective is to derive an equation that translates a fish tissue
concentration of selenium to a water column concentration, the term Cwater is reintroduced into
Equation 9 by defining the enrichment function EF representing the steady state proportional
bioconcentration of dissolved selenium at the base of the aquatic food web given as:
C,
EF =•
'particular
r1
water (Equation 12)
where:
Cparticuiate = Selenium concentration in particulate material (|ig/g)
CWater = Concentration of selenium dissolved in water (|ig/L)
EF = Enrichment function (L/g)
IT^
Rearranging the terms of Equation 12:
c =EF*.C (Equation 13)
*— particulat -1-'-1 ^ ^ water \ ~l J
Substituting EF x C t for c in Equation 9 results in:
O Water ^* particulat ^
.-, -whole-body
water
'composite N
(Equation 14)
Solving for the concentration of selenium in water in Equation 14 results in:
1J//fiytt7- u vu,y
(Equation 15)
Because Equation 15 relates a concentration of selenium in water to the concentration of
selenium throughout all tissues of the body, and the intention here is to relate the concentration
of selenium in water to the concentration of selenium in the eggs or ovaries, Cwh0ie-body must be
converted to an equivalent concentration in eggs or ovaries. The EPA achieved this conversion
by incorporating a species-specific conversion factor (CF) into Equation 15. CF represents the
species-specific proportion of selenium in egg or ovary tissue relative to the concentration of
selenium in all body tissues and is given as:
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CF = ess-°vaiy
Cwhoie-body (Equation 16)
Where:
CF = Whole-body to egg-ovary conversion factor (dimensionless ratio).
= Selenium concentration in the eggs or ovaries offish (|ig/g)
y = Selenium concentration in the whole body of fish (|ig/g).
,-, _ J" egg-ovary
whole-body
Rearranging the terms of Equation 16 yields:
C
f>'^£~u v M y
CF (Equation 17)
Ce
Substituting Cwhok_body in Equation 15 with ess °™y yields the translation equation:
CF
x-r _ ^egg-ovary
T.4ifit/7f mm-r~tr'nmrinvit£> i—TI—r ^—» 1—r
(Equation 18)
TTT?composite
where TTFcomposlte equals the product of all trophic transfer functions from trophic level 2
through the target fish species.
Equation 18 establishes an ecosystem-dependent relationship between the concentration
of selenium in the eggs and ovaries of fish with the concentration of selenium in the water-
column. This approach explicitly recognizes the sequential transfer of selenium between
environmental compartments (water, paniculate material, invertebrate tissue, fish tissue, and
eggs and/or ovary tissue) by incorporating quantitative expressions of selenium transfer from one
compartment to the other. Because this approach uses food web modeling along with species-
specific TTF and CF parameters to quantify most of the transfer between compartments,
however, the only field measurements needed to relate selenium in egg-ovary and water are
measurements from the water-column and particulate material sufficient to calculate EF.
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4.2.2 Equation Parameters
Empirical or laboratory data related to selenium bioaccumulation in aquatic organisms
are needed to derive the equation parameters EF, TTF, and CF. The EPA obtained data from
published literature as described above, using the EPA database ECOTOX. The search resulted
in the retrieval of 54 acceptable studies containing a total of 6,838 selenium measurements at 610
aquatic sites (2,170 from water, 275 from algae, 30 from detritus, 780 from sediment, 1056 from
various species of invertebrates, and 2,170 from various species offish) and 34 acceptable
studies yielding 139 physiological constants (48 values of ke, 81 values of AE, and 10 values of
IR). The EPA used this database of selenium measurements to calculate site-specific EF values
and develop species-specific TTF and CF values in an unbiased and systematic manner. A more
detailed description of how the EPA calculates EF is described below. A more detailed
description of how the EPA calculates TTF and CF is described in Appendix B.
Derivation of Trophic Transfer Function (TTF) Values
The EPA derived TTF values for taxonomic groups of invertebrates and fish by either
using physiological coefficients found in the literature, or by evaluating the empirical
relationship between matched pairs of selenium measurements in organisms and the food they
consumed. When physiological coefficients were available, the EPA calculated a TTF value
using the equation:
k
e
Where:
k
e = Elimination rate constant (/d)
AE = Assimilation efficiency (%)
IR = Ingestion rate (g/g-d)
The EPA also derived TTF values using empirical measurements of selenium from field
studies. The EPA searched its database of available selenium measurements and identified
measurements taken from aquatic organisms. For each measurement from an aquatic organism,
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the EPA searched its selenium database again for additional measurements from other aquatic
organisms or particulate material that was collected from the same aquatic site and of a type
deemed likely to be ingested as a food source or in conjunction with feeding activity (i.e., a
lower trophic level). If multiple lower trophic level measurements were matched to an aquatic
organism measurement, the median of the lower trophic level measurements was calculated.
Each pair of measurements, one taken from an aquatic organism and the other taken from lower
trophic level organisms or particulate material, was designated as a matched pair.
Because selenium is transferred to aquatic animals primarily through aquatic food webs,
the observable concentration of selenium in different environmental compartments may vary
over time. To establish an appropriate time period with which to define matched pairs of
selenium measurements, the effect of sample collection time on the relationship between
selenium concentrations in different media was analyzed. The EPA defined matched pairs of
selenium measurements as described above using different relative collection time ranges and
estimated the strength of the relationship between the two measurements by calculating the
Pearson product-moment correlation coefficient (r).
Figure 8 shows the correlation coefficients for selenium measurements taken from the
same aquatic sites when the measurement collection times were systematically shifted relative to
one another. Each correlation coefficient was calculated from a set of data within a specified
range of relative collection times with respect to the higher trophic level. For example, the
correlation coefficient calculated from particulate and invertebrate measurements with a relative
sample collection time of 30 to 60 days were from invertebrate and particulate samples collected
at the same site, with the invertebrate samples collected 30 to 60 days after the particulate
samples. Similarly, the correlation coefficient calculated from particulate and invertebrate
measurements with a relative collection time of-60 to -30 days were from invertebrate and
particulate samples that were collected at the same site, with the invertebrate samples collected
30 to 60 days before the particulate samples.
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Particulate versus invertebrate
'1
IMi V'i*****"******,
<
rji
i
<
rJi?l
<
1
(
i
•
i n
1
<
i
r o
-1 -
Invertebrate versus fish
'l-
-i
i
00000000
OOo~rtrtrtOOOOOOOOOOOOOOOOOOOO
?^OCTsS>^°°°°°°°°°°°°°°°°°°°°
f.) l-~-> O O tO i—' i—' tO tO tO OJ OJ OJ OJ -t^ -t^ -t^ L/i L/i L/i ^ ^ ^ ^ ^
r—) O'-'iOOi—'-P^^lOOJ^^OtOL/iOOi—'-P^^lOOJ^^OtO
" 00000000000000000000
Number of days between col lection time of paired samples
Figure 8. Effect of relative sample collection time on correlation coefficients of selenium
measurements in particulate material, and invertebrate and fish tissue.
Error bars indicate the 95% confidence interval of r calculated using Fisher's r to z
transformation. Horizontal dashed line indicates r = 0; vertical dashed line indicates relative
collection time expected to have the highest correlation. The absence of a correlation coefficient
indicates an insufficient quantity of data at the specified relative collection time range.
The results of this analysis suggest that the relationship between selenium concentrations
in particulate material and invertebrate tissue and between invertebrate tissue and fish tissue is
somewhat insensitive to relative collection time within a one year time period. These results also
suggest that selenium becomes relatively persistent in the aquatic ecosystem once dissolved
selenium transforms to particulate selenium and becomes bioavailable. On the basis of this
analysis, the EPA concludes that selenium measurements from samples collected at the same
aquatic site within one year of each other are reasonable acceptability criteria for matched pairs
of measurements from the aquatic sites in the EPA database. Note that the EPA chose a relative
collection time period of one year on the basis of data taken from many different aquatic sites.
Individual aquatic sites may have selenium loads and/or bioaccumulation characteristics that
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require different relative collection time criteria to accurately characterize selenium
relationships.
After matched pairs of selenium measurements from samples collected in the field were
identified, the EPA evaluated two different analytical approaches to derive species-specific TTF
values from them. TTF was previously defined above as the steady state proportion relating the
concentration of selenium in the tissue of aquatic organisms to the concentration of selenium in
the food they ingest such that:
CtlsSue = TTF X Cfood (Equation 4)
Rearranging the terms of Equation 4 yields:
C
^^ tissue
r
^ food
Because TTF can be defined as the ratio of the concentration of selenium observed in the
tissue of an aquatic organism to the concentration of selenium observed in the tissue or material
the organism ingests, one approach for deriving TTF values from field data is to simply use the
ratio of the two values. The EPA evaluated this approach by calculating the ratios for all matched
pairs of selenium measurements, and for each species or taxonomic group, used a statistic of
central tendency of the distribution of ratios as the TTF value. An advantage of quantifying the
relationship between selenium in two environmental compartments using ratios is that it is a
simple and straightforward method that is conceptually similar to a bioaccumulation factor
(BAF). A disadvantage of this approach is that it presumes that the quality and quantity of data
used to derive the ratios adequately represents the relationship being characterized. Furthermore,
many aquatic organisms tend to bioaccumulate more metals at low environmental concentrations
(McGeer et al. 2003; Borgman et al. 2004; DeForest et al. 2007; USEPA 2007). Thus a
distribution of ratios could be biased toward larger values if the data are obtained from aquatic
systems with low selenium concentrations.
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Another analytical approach for deriving TTF values from matched pairs of selenium
measurements is to model the species-specific relationships using linear regression. One
possibility is to regress the concentration of selenium in the food of a particular species or
taxonomic group with the concentration of selenium in the organism's tissue, and use the
regression coefficient as the TTF. The EPA evaluated this approach by applying ordinary least
squares (OLS) linear regression on the matched pairs of data. The regression coefficient (slope of
the fitted line) was then taken as the TTF value for that species or taxonomic group. An
advantage of this regression approach is that it estimates the quantitative relationship of selenium
across a range of environmental concentrations in a manner that allows statistical assessment.
Disadvantages of this regression approach includes the assumption that the underlying data are
normally distributed, one or a few very high values can have a disproportionate influence on the
slope of the fitted line, and the fact that the bioaccumulation model does not account for a non-
zero y-intercept. Constraining the y-intercept to zero (also known as regression through the
origin or RTO) eliminates the added complexity of a non-zero y-intercept. However, RTO
further increases the disproportionate influence of one or a few high values on the slope of the
fitted line. Furthermore, RTO does not provide a straightforward way of evaluating goodness of
fit (Gordon 1981).
After evaluating both approaches, the EPA decided to use a hybrid approach by
designating the median of the ratio of matched pairs of selenium measurements as the TTF value,
but only if OLS linear regression of those data resulted in a significant (P < 0.05) fit and positive
regression coefficient. Requiring a significant positive OLS linear regression coefficient
confirms the relationship between selenium in organisms and the food they ingest is adequately
represented by the available data, and using the median of the individual ratios provides an
estimate of central tendency for that relationship that is less sensitive to potential bias due to
measurements taken from aquatic systems with very high or very low selenium concentrations.
The EPA calculated TTF values for 13 invertebrate species and 30 fish species that live in
freshwater aquatic environments in North America. The data used to derive these TTF values are
provided in Appendix B. The final T7F values are listed in Tables 9 and 10. The presence of
physiological coefficients for a tax on in Tables 9 and 10 indicates that the TTF values were
calculated using those parameters. The absence of physiological coefficients for a tax on indicates
the EPA derived the TTF value using field data. If a TTF value could be calculated from both
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physiological coefficients and field data, the EPA used the TTF value calculated from the
substantially larger number of field measurements so as to minimize statistical uncertainty.
Table 9. EPA-derived Trophic Transfer Function (TTF) Values for Freshwater
Aquatic Invertebrates.
Common name
Scientific name
AE
IR
ke
TTF
Crustaceans
amphipod
copepod
crayfish
water flea
Hyalella azteca
copepods
Astacidae
Daphnia magna
-
0.520
-
0.406
- ^^fl|
0.420
-
0.210
-
0.155
-
0.116
1.22
1.41
1.46
0.74
Insects ^ ^
dragonfly
damselfly
mayfly
midge
water boatman
Anisoptera
Coenagrionidae
Centroptilum triangulifer
Chironimidae
Corixidae
-
-
0.390
-
-
w
-
0.720
^^^S
-
-
-
0.220
-
-
1.97
2.88
1.28
1.90
1.48
Mollusks
asian clama
zebra mussel
Corbiculafluminea
Dreissena polymorpha
0.550
0.260
0.050
0.400
0.006
0.026
4.58
4.00
Annelids
blackworm
Lumbriculus variegatus
0.165
0.067
0.009
1.29
Other
zooplankton
zooplankton
-
-
-
1.89
a Not to be confused with Corbula amurensis
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Table 10. EPA-Derived Trophic Transfer Function (TTF) Values for Freshwater
Fish.
Common name
Scientific name
AE
IR
ke
TTF
Cypriniformes
bluehead sucker
common carp
creek chub
fathead minnow
flannelmouth sucker
longnose sucker
sand shiner
white sucker
Catostomus discobolus
Cyprinus carpio
Semotilus atromaculatus
Pimephales promelas
Catostomus latipinnis
Catostomus catostomus
Notropis stramineus A
Catostomus commersonii ^J
-
-
-
-
-
£•
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
1.04
1.29
1.12
1.57
1.06
0.90
1.83
1.18
Cyprinodontiformes
mosquitofish
northern plains killifish
western mosquitofish
Gambusia sp. ^F
Fundulus kansae
Gambusia a/finis
-
-
-
-
-
-
-
-
-
0.97
1.27
1.25
Esociformes
northern pike
Esox lucius
-
-
-
1.79
Gasterosteiformes
brook stickleback
Culaea inconstans
-
-
-
1.69
Perciformes
black crappie
bluegill
green sunfish
largemouth bass
striped bass
walleye
yellow perch
Pomoxis nigromaculatus
Lepomis macrochirus
Lepomis cyanellus
Micropterus salmoides
Morone saxatilis ^^
Sander vitreus
Percaflavescens
-
-
-
-
0.375
-
-
-
-
-
-
0.335
-
-
-
-
-
-
0.085
-
-
2.67
1.48
1.27
1.27
1.48
1.82
1.42
Salmoniformes
brook trout
brown trout
cutthroat trout
mountain whitefish
rainbow trout
westslope cutthroat trout
Salvelinus fontinalis
Salmo trutta
Oncorhynchus clarkii
Prosopium williamsoni
Oncorhynchus mykiss
Oncorhynchus clarkii lewisi
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
0.88
1.44
1.07
1.38
1.19
1.20
Scorpaeniformes
mottled sculpin
sculpin
Cottus bairdi
Cottus sp.
-
-
-
-
-
-
1.38
1.29
Siluriformes
black bullhead
channel catfish
Ameiurus melas
Ictalurus punctatus
-
-
-
-
-
-
0.91
0.73
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Derivation of Whole-Body to Egg-Ovary Conversion Factor (CF) Values
The parameter CF (conversion factor) in Equation 18 represents the species-specific
partitioning of selenium as measured in the whole-body and in egg-ovary tissue. The EPA
derived species-specific CF values by applying the same method used to derive species-specific
TTF values using empirical measurements of selenium concentrations in different tissues of the
same fish. To derive whole-body to egg-ovary CF values, the EPA defined matched pairs of
selenium measurements from the whole-body and from the eggs or ovaries measured from the
same individual fish or from matched composite samples. Egg-ovary concentration was defined
as a measurement from either the eggs or the ovaries. If multiple measurements from both eggs
and ovaries of the same individual or matched composite sample were available, the average
value was used. Similar to the procedure used to derive TTF values, the EPA first confirmed a
statistical relationship between egg-ovary and whole body selenium for each species using OLS
linear regression of the matched pairs of measurements. If the regression resulted in a significant
fit (P < 0.05) with a positive regression coefficient, the EPA calculated the ratio of the egg-ovary
to whole body selenium concentration of each matched pair and used the median ratio as the CF
value for the species.
Some selenium measurements in eggs and/or ovaries were only available with
corresponding measurements from muscle tissue. In those cases, the EPA estimated an
equivalent whole body concentration from the muscle concentration by applying a muscle to
whole-body conversion factor. The EPA derived the muscle to whole-body conversion factor
using the same method that was used to derive CF values. Matched pairs of muscle concentration
measurements were regressed on whole-body concentration, and if the regression resulted in a
significant fit (P < 0.05) with a positive regression coefficient, the median of the whole-body to
muscle concentration ratios was calculated for each species. Because data from only a few fish
species were available, the EPA used the median of the resulting species-specific muscle to
whole-body ratios as a single muscle to whole-body conversion factor of 1.27 for all fish species.
There were a sufficient number of egg-ovary and whole-body selenium measurements to
directly derive CF values for 9 species offish found in freshwater aquatic environments of North
America. The EPA derived CF values for 7 additional species using muscle and egg-ovary
selenium measurements and applying a muscle to whole-body conversion factor of 1.27. The
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data and methods used to derive the CF values are described in detail in Appendix B. The final
CF values are listed below in Table 11.
Table 11. EPA-Deriyed Egg-Ovary to Whole-Body Conversion Factor (CF) Values
Common name
Scientific name
CF
Cypriniformes
bluehead sucker
common carp
flannelmouth sucker
razorback sucker
roundtail chub
white sucker
Catostomus discobolus
Cyprinus carpio
Catostomus latipinnis
Xyrauchen texanus
Gila robusta
Catostomus commersonii
1.82
1.92
1.41
1.43
2.07
1.41
Esociformes
northen pike
Esox lucius
2.39
Perciformes
bluegill
green sunfish
smallmouth bass
Lepomis macrochirus
Lepomis cyanellus
Micropterus dolomieu
2.13
1.45
1.42
Salmoniformes
brook trout
brown trout
cutthroat trout
Dolly Varden
mountain whitefish
rainbow trout
Salvelinus fontinalis
Salmo trutta ^"^^
Oncorhynchus clarkii
Salvelinus malma
Prosopium williamsoni
Oncorhynchus mykiss
1.38
1.45
2.30
1.61
7.39
2.44
Calculation of Site-Specific Enrichment Factor (EF) Values
The single most influential step in selenium bioaccumulation occurs at the base of aquatic
food webs (Chapman et al. 2010). The parameter EF characterizes this step by quantifying the
partitioning of selenium between the dissolved and particulate state. EF can vary by at least two
orders of magnitude across aquatic systems (Presser and Luoma 2010). Uncertainty in translating
a fish tissue concentration of selenium to a water column concentration using Equation 18 is
minimized when site-specific empirical observations of dissolved and particulate selenium of
sufficient quality and quantity are used to accurately characterize EF. Thus the EPA only used
aquatic sites with sufficient data to calculate a reliable EF value.
The EPA searched its database of selenium measurements to identify measurements from
aquatic sites with sufficient particulate and water column data to calculate a reliable site-specific
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EF value. The EPA identified all the selenium measurements from algae, detritus, or sediment,
and then searched for corresponding water column measurements from samples collected at the
same aquatic site within one year of the particulate sample. If more than one water measurement
was available for any given parti culate measurement, the median was used. For each of these
matched pairs of parti culate and water measurements, the EPA calculated the ratio of parti culate
concentration to water concentration. If more than one ratio for any given category of parti culate
material (algae, detritus, or sediment) was calculated at an aquatic site, the EPA used the median
ratio. The geometric mean of the algae, detritus, and sediment ratios was used as the site EF.
Because there were at most only 3 possible values (one for algae, one for detritus, and one for
sediment), the EPA used the geometric mean in order to reduce the potential for one of the
values to have excessive influence on the final site EF value.
The availability of selenium measurements from paniculate material was limited. In
addition, the majority of parti culate measurements were from sediment samples with a
significantly lower correlation to selenium in water (r = 0.42) compared to algae (r = 0.65; Fisher
r-to-z transformation, P < 0.001) and detritus (r = 0.94; Fisher r-to-z transformation, P < 0.001).
Therefore, to reduce uncertainty in estimating site-specific EF values, the analysis was limited to
those aquatic sites with at least two paniculate selenium measurements with corresponding water
column measurements, and only used sediment measurements if there was at least one other
measurement from either algae or detritus. On the basis of these requirements, EF values were
calculated for 69 individual aquatic sites.
4.2.3 Food-Web Models
For the aquatic sites with a calculated EF value, the EPA modeled the food webs as
shown in Figure 9 for the fish species the studies indicated were present. Some of those studies
provided information about the species and proportions of organisms ingested by fish, either
through direct analysis of stomach contents, or examination of the presence and prevalence of
invertebrate species. For those studies, the EPA used that site-specific information in the food
web models. Most studies, however, did not provide site-specific food web information. In those
cases, the food web offish species present were modeled using information about their typical
diet and/or eating habits obtained from the NatureServe database (http://www.natureserve.org).
After the food web models were developed, the EPA identified the appropriate species-
specific TTF values for each model and calculated jj'pcomPosl-te^ Although TTF values were
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derived for several different taxa of invertebrates and fish (Tables 9 and 10), some of the food
web models included one or more taxa for which no TTF value was available. The EPA assigned
a TTF value to these taxa by sequentially considering higher taxonomic classifications until one
or more taxa for which a TTF value was available matched the tax on being considered. If the
lowest matching tax on was common to more than one species with a TTF value available, the
EPA used the median TTF from the matching species.
For example, one study reported selenium concentrations in Rhinichthys atratulus
(blacknose dace). Although the TTF value of Rhinichthys atratulus was not available, this
species is in the family Cyprinidae, which also includes Cyprinus carpio (common carp) and
Gila robusta (roundtail chub). Because Cyprinidae is the lowest taxonomic classification where
the fish species being considered matches a species with an available TTF value, the EPA used
the median of the common carp and roundtail chub TTF values for the TTF value of blacknose
dace. In another example, a study reported selenium concentrations in Sander vitreus, walleye.
Although the TTF value for Sander vitreus was not available, it is in the order Perciformes,
which is common to Pomoxis nigromaculatus (black crappie), Lepomis macrochirus (bluegill),
Lepomis cyanellus (green sunfish), and Micropterus dolomieu (smallmouth bass). Thus the EPA
used the median TTF values of those four fish species for walleye. Substitution methods using
other approaches such as feeding behavior yielded generally similar results.
For each food web model, a CF value was also identified for the targeted fish species
using the list of available values in Table 11. Although the EPA had a relatively large amount of
data to calculate a diverse set of species-specific CF values, some food web models included
taxa for which no CF value was listed in Table 11. A CF value was assigned to these taxa using
the same procedure that was used to assign TTF values by sequentially considering higher
taxonomic classifications until one or more taxa for which a CF value was available matched the
tax on of the targeted fish species.
4.2.4 Classifying Categories of Aquatic Systems.
Transformation reactions that convert dissolved selenium to particulate forms are the
primary route of entry into aquatic system food webs and a critical step in selenium
bioaccumulation and toxicity (Chapman et al. 2010). However, site-specific characteristics can
result in substantial bioaccumulation variability and consequently different risks of selenium
toxicity for a given concentration of dissolved selenium. One such site-specific characteristic is
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water residence time. Aquatic organisms living in waters with long residence times such as lakes,
ponds, reservoirs, wetlands or estuaries tend to bioaccumulate more selenium than those living in
waters with shorter residence times such as rivers and streams (ATSDR 2003; Luoma and
Rainbow 2005; Orr et al. 2006; Simmons and Wallschlagel 2005; EPRI2006). Because water
residence time is an almost universally described feature of aquatic systems, the EPA used this
characteristic to evaluate potential categories of aquatic systems where a single water-column
concentration value would be adequately protective.
The EPA evaluated potential categories of aquatic systems by analyzing the relationship
between residence time and EF. From the studies contributing data to the EPA's database of
available selenium measurements, the EPA identified the term used by the study authors to
describe the waterbody with respect to residence time. Of the 69 aquatic systems with a
calculated EF value, the study authors described them as either lakes (n = 7), reservoirs (n = 3),
ponds (n = 16), rivers (n = 6), creeks (n = 27), drains (n = 3), marshes (n = 3), washes (n = 2), or
streams (n = 2). Categories that did not have a sufficient number of values to perform a
meaningful analysis were combined into single categories on the basis of residence time. Thus
the EPA grouped aquatic sites into four categories: 1) lakes and reservoirs; 2) ponds and
marshes; 3) rivers; and 4) streams, drains, washes and creeks. Figure 9 summarizes the
distribution of EF values for these 69 aquatic sites when grouped into these 4 categories.
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EF
4 -
Lakes and Ponds and Rivers Streams, creeks,
reservoirs marshes (n = 6) drains, and washes
(n=10) (n=19) (n=34)
Figure 9. Distribution of EF values for 69 aquatic sites derived from published studies and
grouped into 4 categories.
Boxes indicate first quartile, third quartile, and median; whiskers indicate minimum and
maximum values (maximum value of the lakes and reservoirs category was allowed to exceed
the y-axis to better compare medians, means, and quartiles). Dashed line shows the grand median
of all data.
A Kolmogorov-Smirnov goodness of fit test indicated that the distribution of EF values
within each category was not normally distributed (lakes and reservoirs: P < 0.00001; ponds and
marshes: P < 0.00001; rivers: P < 0.002; streams, creeks, drains, and washes: P < 10"8). Thus the
EPA used nonparametric statistics to evaluate the central tendency of each group and test
differences between them. A Kruskal-Wallis test indicated a significant difference among the
medians of the 4 aquatic system categories (P < 0.0005). However, a multiple comparison of
mean ranks using Scheffe's S procedure indicated a significant difference only between the mean
ranks of the lakes and reservoirs category and the streams, creeks, drains, and washes category.
There were no significant differences in mean ranks among the other categories. The EPA
concludes from these data and analyses that currently available information does not adequately
differentiate among these 4 categories of aquatic systems with respect to selenium
bioaccumulation, and thus does not support the establishment of individual water concentration
values for all four categories of water body.
The grand median of all EF values from all categories was calculated and compared to
the median EF value of each category. The dashed line in Figure 11 shows the grand median EF
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value (0.62 L/g) for all 69 aquatic sites. All categories with a median greater than the grand
median are lentic aquatic systems, and all categories with a median less than the grand median
are lotic aquatic systems. The EPA evaluated the potential for differentiating aquatic system
categories on the basis of whether they are lentic or lotic by grouping EF values from lakes,
reservoirs, ponds, and marshes into the category lentic aquatic systems; and rivers, streams,
creeks, drains, and washes into the category lotic aquatic systems. Figure 10 summarizes theEF
values when grouped using these two categories.
EF
4 -
Lentic
(n=29)
Lotic
(n=40)
Figure 10. Distribution of EF values for the same 69 aquatic systems as shown in Figure 9
grouped into 2 categories (lentic and lotic).
Boxes depict first quartile, third quartile, and median; whiskers depict minimum and maximum
values (maximum value of lentic aquatic systems was allowed to exceed the y-axis to better
compare medians, means, and quartiles).
A Kolmogorov-Smirnov goodness of fit test indicates that the distribution of EF values in
O 1 f\ ^^ _^
the lentic (P < 10" ) and lotic (P < 10" ) categories are not normally distributed. Thus the EPA
again used nonparametric statistics to characterize the central tendency of each group and to test
differences between them. A Mann-Whitney U test indicates EF values the from lentic and lotic
categories are significantly different from each other (P < 0.001). The EPA concludes from these
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data and analyses that selenium bioaccumulation tends to be greater in lentic systems compared
to lotic systems. This result is also consistent with other studies (Hamilton and Palace 2001; Brix
et al. 2005; Orr et al. 2006). The EPA further concludes from this analysis that currently
available data and information support the establishment of separate water quality concentration
values for lentic and lotic systems.
4.2.5 Deriving Protective Water Column Concentrations for Lentic and Lotic System
Categories
To derive ambient water quality concentration values appropriate for lentic and lotic
aquatic systems, the EPA translated the egg-ovary criterion element to a set of water
concentration values on the basis of the EF values calculated for 69 aquatic sites and food web
models of the fish present in the aquatic systems. Because more than one fish species were often
present, many sites provided more than one translated water column concentration (one for each
species at each site). Thus the egg-ovary criterion element was translated into a total of 132
water column concentration values (50 values from 29 lentic aquatic systems and 82 values from
40 lotic aquatic systems).
The EPA used the distribution of water concentration values translated from the egg-
ovary criterion element to derive chronic water column criterion element values for lentic and
lotic aquatic systems. Table 12 shows the model parameters used to translate the egg-ovary
criterion element to site-specific water concentrations and Figure 11 shows distribution of the
translated values.
Table 12. Site-Specific Data for the 132 Species-Site Combinations and Translation
of the Egg-Ovary Criterion Concentraiton Element to a Water Column Concentration.3
Identification
Ref3
Bi
Bi
Bi
Bi
Bi
Bi
Bi
Bi
Site3
22
27
23
20
7
22
23
30
Species3
FM
FM
FM
ID
ID
ID
ID
NPK
Type
lentic
lentic
lentic
lentic
lentic
lentic
lentic
lentic
Model Parameters
EFb
2.37
0.87
1.21
2.31
0.88
2.37
1.21
1.70
CFC
2.00
2.00
2.00
1.45
1.45
1.45
1.45
1.71
T TplComPosite-d
2.35
2.35
2.35
2.56
2.56
2.56
2.56
1.98
Translated chronic water
criterion concentration
element
r e
^water
1.37
3.71
2.68
1.77
4.67
1.72
3.38
2.63
Do not distribute, quote, or cite
85
Draft Document
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Identification
Ref
Bi
Bi
Bi
Bu91
Bu91
Bu91
Bu91
Bu91
Bu91
Bu93
Bu93
Bu93
Bu93
Bu93
Bu93
Bu93
Bu93
Bu93
Bu95
Bu95
Bu95
Bu95
Bu95
Bu95
Bu95
Bu95
Bu95
Bu95
Bu95
Bu95
Bu95
Bu95
Bu95
Bu95
Bu95
Bu95
Bu95
Bu95
Bu95
Site3
3
27
23
4
4
4
4
4
4
SP2
N2
SP2
N2
N2
N2
SP2
SP2
SP2
ME2
ME3
NW
SJ1
SJ3
ME3
SJ1
SJ3
ME4
ME3
SJ1
SJ3
HD2
ME1
ME2
ME4
ME3
we
SJ1
HD2
ME2
Species"
NPK
NPK
NPK
BhS
BnT
FS
MS
RT
WS
BhS
BT
BT
BB
ChC
cc
FM
SD
WS
BhS
BhS
BhS
BhS
BhS
BB
ChC
ChC
CC
cc
cc
cc
FM
FM
FM
FM
FM
FM
FS
FS
FS
Type
lentic
lentic
lentic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lentic
lotic
lentic
lentic
lentic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
Model Parameters
EFb
0.58
0.87
1.21
0.63
0.63
0.63
0.63
0.63
0.63
0.18
1.26
0.18
1.26
1.26
1.26
0.18
0.18
0.18
0.37
0.10
0.20
0.26
0.29
0.10
0.26
0.29
0.12
0.10
0.26
0.29
0.15
0.90
0.37
0.12
0.10
0.40
0.26
0.15
0.37
CFC
1.71
1.71
1.71
1.82
1.45
1.41
1.71
2.44
1.41
1.82
1.45
1.45
1.71
1.71
1.92
2.00
2.00
1.41
1.82
1.82
1.82
1.82
1.82
1.71
1.71
1.71
1.92
1.92
1.92
1.92
2.00
2.00
2.00
2.00
2.00
2.00
1.41
1.41
1.41
T TplComPosite-d
1.98
1.98
1.98
1.21
2.42
1.47
2.21
2.04
1.53
1.21
2.42
2.42
1.19
1.19
1.63
2.35
2.23
1.53
1.21
1.21
1.21
1.21
1.21
1.43
1.19
1.19
1.63
1.63
1.63
1.63
2.35
2.35
2.35
2.35
2.35
2.35
1.47
1.47
1.47
Translated chronic water
criterion concentration
element
r1 e
^water
7.76
5.13
3.71
10.94
6.91
11.68
6.36
4.85
11.23
38.53
3.45
24.32
4.92
5.91
3.86
18.12
19.10
39.54
18.75
72.21
35.20
26.30
23.84
64.94
28.43
25.77
40.56
50.95
18.56
16.82
21.73
3.60
8.82
27.03
33.96
8.01
28.09
49.34
20.03
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86
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Identification
Ref
Bu95
Bu95
Bu95
Bu95
Bu95
Bu95
Bu95
Bu95
Bu95
Bu95
Bu95
Bu95
Bu95
Bu97
Bu97
Bu97
Bu97
Bu97
Bu97
Bu97
Ca
Ca
Fo
Fo
Fo
Fo
Fo
Fo
Fo
Fo
Fo
Fo
Fo
Fo
Fo
Fo
Fo
Fo
Fo
Site3
ME4
ME3
SJ3
ME3
ME4
ME3
SJ1
ME1
ME2
ME3
NW
SJ1
HD2
MUD2
MNP2
MUD2
WCP
CHI
MUD2
MNP3
DC
LC
CC-1A
CC-3A
CC-150
CC-350
CC-75
DC
HS
HS-3
LSV-2C
LSV-4
SFTC
CC-1A
CC-3A
CC-150
CC-350
CC-75
DC
Species"
FS
FS
FS
GnS
RSh
RSh
RSh
SD
SD
SD
SD
SD
Su
BhS
FM
FM
FM
GnS
GnS
SB
RT
RT
BnT
BnT
BnT
BnT
BnT
BnT
BnT
BnT
BnT
BnT
BnT
Sc
Sc
Sc
Sc
Sc
Sc
Type
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lentic
lotic
lentic
lotic
lotic
lentic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
lotic
Model Parameters
EFb
0.12
0.10
0.29
0.10
0.12
0.10
0.26
0.90
0.37
0.10
0.20
0.26
0.15
0.07
2.00
0.07
0.90
0.20
0.07
5.15
2.24
0.33
0.80
0.81
1.04
1.16
1.19
1.55
0.24
0.54
0.45
0.69
1.32
0.80
0.81
1.04
1.16
1.19
1.55
CFC
1.41
1.41
1.41
1.45
2.00
2.00
2.00
2.00
2.00
2.00
2.00
2.00
1.41
1.82
2.00
2.00
2.00
1.45
1.45
1.42
2.44
2.44
1.45
1.45
1.45
1.45
1.45
1.45
1.45
1.45
1.45
1.45
1.45
1.71
1.71
1.71
1.71
1.71
1.71
T TplComPosite-d
1.47
1.47
1.47
2.11
2.16
2.16
2.16
2.23
2.23
2.23
2.23
2.23
1.27
1.21
2.35
2.35
2.35
2.11
2.11
2.61
2.04
2.04
2.54
2.56
2.52
2.46
2.54
2.49
3.60
2.31
2.57
2.56
2.55
2.28
2.29
2.26
2.20
2.28
2.23
Translated chronic water
criterion concentration
element
r1 e
^water
61.38
77.10
25.46
51.93
29.38
36.90
13.44
3.80
9.30
35.79
17.45
13.04
57.17
J
98.08
1.62
46.12
3.58
25.39
70.54
0.80
1.37
9.39
5.18
5.09
4.00
3.67
3.48
2.72
11.96
8.48
9.16
5.92
3.11
4.87
4.79
3.77
3.46
3.28
2.56
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87
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Identification
Ref
Fo
Fo
Fo
Fo
Fo
Gr
Gr
HB
Le
Le
Le
Le
Le
Le
Le
Le
Le
Le
Le
Le
Le
Le
Le
Le
Le
Le
Sa87
Sa87
Sa87
Sa87
Sa87
Sa87
Sa93
Sa93
Sa93
Sa93
Sa93
Site3
HS
HS-3
LSV-2C
LSV-4
SFTC
17
17
LEMC
BA
BE
HR
BA
BE
HR
BA
BE
HR
BA
BE
HR
BA
BE
HR
BA
BE
HR
KP11
KP2
KP8
SLD
VP26
VW
GT4
GTS
SJR2
SJR3
GT4
Species"
Sc
Sc
Sc
Sc
Sc
FM
WS
CT
BB
BB
BB
CC
CC
CC
FM
FM
FM
GnS
GnS
GnS
WM
WM
WM
RSh
RSh
RSh
WM
WM
WM
WM
WM
WM
BgS
BgS
BgS
BgS
LMB
Type
lotic
lotic
lotic
lotic
lotic
lentic
lentic
lotic
lentic
lentic
lentic
lentic
lentic
lentic
lentic
lentic
lentic
lentic
lentic
lentic
lentic
lentic
lentic
lentic
lentic
lentic
lentic
lentic
lentic
lentic
lentic
lotic
lentic
lentic
lotic
lotic
lentic
Model Parameters
EFb
0.24
0.54
0.45
0.69
1.32
0.86
0.86
1.32
8.54
2.09
3.81
8.54
2.09
3.81
8.54
2.09
3.81
8.54
2.09
3.81
8.54
2.09
3.81
8.54
2.09
3.81
0.51
0.32
0.60
0.36
0.93
1.03
0.43
1.37
0.36
0.75
0.43
CFC
1.71
1.71
1.71
1.71
1.71
2.00
1.41
2.30
1.71
1.71
1.71
1.92
1.92
1.92
2.00
2.00
2.00
1.45
1.45
1.45
1.71
1.71
1.71
2.00
2.00
2.00
1.71
1.71
1.71
1.71
1.71
1.71
2.13
2.13
2.13
2.13
1.42
T TplComPosite-d
3.22
2.07
2.30
2.29
2.29
2.35
1.53
1.78
1.66
1.66
1.66
1.63
1.63
1.63
2.35
2.35
2.35
2.11
2.11
2.11
1.57
1.57
1.57
2.16
2.16
2.16
2.03
2.03
2.03
2.03
2.03
2.03
2.11
2.11
2.11
2.11
1.86
Translated chronic water
criterion concentration
element
r1 e
^water
11.25
7.98
8.62
5.57
2.93
3.75
8.18
2.83
0.62
2.55
1.40
0.57
2.33
1.28
0.38
1.55
0.85
0.58
2.38
1.30
0.66
2.70
1.48
0.41
1.69
0.92
8.62
13.79
7.24
12.13
4.66
4.24
7.90
2.47
9.44
4.52
13.47
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Identification
Ref
Sa93
Sa93
Sa93
Sa93
Sa93
Sa93
Sa93
St
St
Site3
GTS
SJR2
SJR3
GT4
GTS
SJR2
SJR3
M4720
M4720
Species"
LMB
LMB
LMB
WM
WM
WM
WM
BB
CC
Type
lentic
lotic
lotic
lentic
lentic
lotic
lotic
lentic
lentic
Model Parameters
EFb
1.37
0.36
0.75
0.43
1.37
0.36
0.75
0.10
0.10
CFC
1.42
2.13
1.42
1.71
1.71
1.71
1.71
1.71
1.92
T TplComPosite-d
1.86
1.86
1.86
1.93
1.93
1.93
1.93
1.66
1.63
Translated chronic water
criterion concentration
element
r1 e
^water
4.21
16.11
7.71
10.78
3.37
12.89
6.17
55.63
50.76
a- See Appendix L for description of abbreviations.
b- Geometric mean of the median enrichments functions (EF) for all available food types (algae,
detritus, and sediment). EF (L/g) = Cf00d/C water-
c- Taxa-specific conversion whole-body to egg ovary conversion factor (CF; dimensionless ratio).
d- Composite trophic transfer factor (TTFcomP°slte) product of TTF valuess for all trophic levels.
e- Translated water concentration corresponding to an egg-ovary criterion element of 15.2 mg Se/kg
dw, calculated by Equation 18.
1 -I
Cumulative
proportion
0
0
100
1 10
Water (jig/L)
Figure 11. Probability distribution of the water-column concentrations translated from the
egg-ovary criterion at lentic and lotic aquatic sites.
Dashed and dash-dot lines show the 20th percentiles of the lentic and lotic distributions,
respectively.
Table 13 summarizes the distributions of translated water concentration values. As
discussed in the Introduction and Background, the bioaccumulation potential of selenium
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depends on many different biogeochemical factors that characterize a particular aquatic system.
Uncertainty in the translation of the egg-ovary criterion concentration element to a water column
concentration element can be reduced by using site-specific data and information such as an EF
value calculated from site-specific measurements and a food-web model derived from a
biological assessment of the aquatic system. The basis for national water column criterion
element values, however, is constrained by the need to apply a single value to a large number of
aquatic systems, the available data used to derive the values, and the need to implement a variety
of simplifying assumptions which introduces uncertainty. Although the EPA utilized medians as
the statistical method of choice to characterize multiple measurements (except when estimating
the selenium concentration in paniculate material from algae, detritus, and sediment for the
reasons described above), the EPA selected the 20th percentile of the translated water column
values of each category (lotic and lentic) as the final water column criterion concentration
element. The EPA selected the 20* percentile to ensure adequate protection of the aquatic
species. The EPA examined the the performance of the 20* percentile water column values by
evaluating the protectiveness of the water column criterion element using binary classification
statistics. In this analysis EPA used an independent data set composed of measured
concentrations of selenium from 2,588 lotic and 596 lentic sites to complete a verification or
"ground-truthing" of the selected 20th percentile water column values to evaluate their
protectiveness, and found that the use of the 20th percentile water column values would prevent
potential exceedances of the fish tissue egg-ovary criterion element over 90% of the time, i.e.,
false negative conclusions regarding fish tissue exceedances would be minimized using the
selected 20* percentile water column value for the water column criterion element derivation.
However, states and tribes may choose to adopt a lower percentile value if there is reason to
believe that selenium bioaccumulation is greater and/or more variable in their waters or other
site-specific considerations. For this reason, the 10th and 5th percentile of the distribution of
translated water column concentrations are also presented in Table 13. States and tribes may
also chose to translate the egg-ovary criterion element to a water column concentration on a site-
specific basis.
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Table 13. Summary of water column criterion element concentration values
translated from the egg-ovary criterion element.
The 20th percentile values are the water column criterion element concentration values for the
national selenium criterion. All units |ig/L. This analysis is based on 50 lentic sites and 82 lotic
sites.
Median
20th percentile (final water-column criteria)
10th percentile
5th percentile
Lentic
2.7
1.3
0.7
0.6
Lotic
12.0
4.8
3.5
2.9
The distributions of translated chronic water column concentrations to which the 20*
percentile applies were derived using the medians of site-specific measurements. However, a site
would not attain its chronic water criterion element if only its median concentration attained;
rather its high-end concentrations would need to attain. Consequently, it cannot be inferred that
as many as 20% of sites that are at their tissue concentration would attain their chronic water
criterion element. Rather, the protectiveness of the water criteria are further described in Section
7.2.2.
4.2.6 Derivation of Averaging Period for Chronic Water Criterion Element
For setting averaging periods for aquatic life criteria, U.S. EPA (1995b) used the concept
that the criterion averaging period should be less than or equal to the "characteristic time"
describing the toxic speed of action. In the context of the waterborne direct toxicity of metals,
characteristic time = 1/k, where k is the first-order kinetic coefficient in a toxicokinetic model
fitted to the relationship between LCso and exposure duration.
In the context of selenium bioaccumulation in a single trophic level, k would the first-
order depuration coefficient, and 1/k would equal the time needed to depurate to a concentration
of 1/e times the initial concentration (where e=2.718). For depuration of two trophic levels
sequentially, invertebrates and fish, the characteristic time is likewise the time needed for c/c0
reach a value of 1/e.
For the first trophic level, the kinetics for algal bioaccumulation and depuration were
assumed to be rapid compared to the larger organisms as higher trophic levels; that is, the
characteristic time for algae was assumed to be negligible.
For the second trophic level, invertebrates, values for kiL2 are tabulated in elsewhere in
the document. A value of 0. I/day appears to be environmentally conservative, considerably
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higher than those for Lumbriculus, Asian clam, zebra mussel, but a bit lower than that for
copepods, which are very small in size. This corresponds to a characteristic time of 10 days.
For fish, the median depuration coefficient measured by Bertram and Brooks (1986) for
6-9 month-old (early adult) fathead minnows is applied, providing a kTL3 value of 0.02/day. This
corresponds to a characteristic time of 50 days. Because of the small size of adults of this
species, this represents faster kinetics than would likely be applicable to the salmonids and
centrarchids of greatest concern for selenium toxicity, consonant with the Newman and Mitz
(1988) inverse relationship between depuration rate and organism size. The striped bass k value
of Baines et al. (2002) is inapplicable here because it was measured in the early juvenile life
stage, a size that is too small to be relevant to reproductive impairment stemming from exposure
of adult females.
As shown in Appendix G, the characteristic time for the combined second and third
trophic levels (invertebrates and fish) is the approximate sum of the above two characteristic
times, or 60 days. The analysis of the protectiveness of a 30-day averaging period, shorter than
the characteristic time, was performed and is shown in Appendix G. That analysis demonstrated
that a 30-day averaging period for the chronic water criterion affords protection under all
conditions, and is therefore the duration recommended for the chronic water column criteria.
4.3 Intermittent-Exposure Water Criterion Element: Derivation from the Chronic Water
Criterion Element
Chapman et al. (2009) noted that selenium acute toxicity has been reported rarely in the
aquatic environment and that traditional methods for predicting effects based on direct exposure
to dissolved concentrations do not work well for selenium. Consequently, the available database
of acute toxicity LCsos for selenite and selenate are not useful for criteria purposes. As
demonstrated in Appendix G, the kinetics of selenium accumulation and depuration are
sufficiently slow that attainment of the water criterion concentration element by ambient 30-day
averages will protect sensitive aquatic life species even where concentrations exhibit a high
degree of variability.
To address intermittent exposures that could contribute to chronic effects of selenium due
to its bioaccumulative nature, EPA is providing an intermittent exposure water criterion
concentration element intended to limit cumulative exposure to selenium, which is derived from
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the chronic 30-day water criterion. To illustrate the concept, Figure 12 shows a possible
sequence of exposures over a 30-day period.
I
V.
£
c
•••iMnininilninlninin
10 19
Day
28
Figure 12. Illustration of intermittent spike exposure occurring for a certain percentage of
time (e.g., 10%) over a 30-day period, and background exposure occurring for the
remaining percentage of time (e.g., 90%).
The 30-day average concentration, Csoday, is given by:
"•"
~ Tint)
30 day = intJ
where CWis the intermittent spike concentration,/^ is the fraction of any 30-day period during
which elevated selenium concentrations occur, and Cbkgrndis the background concentration
occurring during the remaining time. Csodayis not to exceed the chronic criterion, WQCsoday. If
the intent is to apply a criterion element, WQC/ntto the intermittent spike concentrations, then
replacing C/nt with WQCJntand €30 day with WQCsodayVfr the above equation, and then solving for
WQCmt yields:
— Cbkgrnd(l — fint~)
fi
int
The above equation expresses the intermittent exposure water criterion element in terms
of the 30-day average chronic water criterion element, for a lentic or lotic system, as appropriate,
while accounting for the fraction in days of any 30-day period the intermittent spikes occur and
for the concentration background occurring during the remaining time. The reasonable worst-
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case assumption inherent in this approach is that selenium bioaccumulation is linear over a very
wide range of concentrations: that is, EFs and TTFs do not decrease significantly as
concentrations increase.
If the heights of three spikes in Figure 14 were to differ somewhat between each other,
then the intermittent criterion would apply to the arithmetic mean of the three. If the background
concentrations were to vary somewhat, then the arithmetic mean background would be input to
the equation. Nevertheless, the above approach is not intended for application to ordinary
smoothly varying concentrations. That situation is better addressed simply by applying the
chronic water criterion as a 30-day average for a lentic or lotic system, as appropriate.
Table 14 illustrates example values for the intermittent water criterion concentration
element. The bottom row of the lotic and lentic values and the right column are to emphasize that
WQC/ntis not an independent criterion element but a re-expression of the 30-day average water
criterion concentration element. WQCmt converges to WQCsoo^when the background
concentration is already at WQCsodayfx when the intermittent exposure is said to occur
throughout the 30-day period.
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Table 14. Representative Values of the Intermittent Water Criterion Concentration
Element.
Bkgrnd
Cone,
^bkgmd
(HS/L)
0
1
2
O
4.8
0
0.5
0.7
1
1.3
0.033
(1 day)
Fraction of Time,/^ in a 30-day period
0.05
(1.5 days)
0.1
(3 days)
0.2
(6 days)
0.5
(15 days)
1
(30 days)
Lotic Intermittent Criterion, WQQ^ (ug/L)
145.5
116.2
86.8
57.5
4.8
96
77
58
39
4.8
48
39
30
21
4.8
24
20
16
12
4.8
9.6
8.6
7.6
6.6
4.8
4.8
4.8
4.8
4.8
4.8
Lentic Intermittent Criterion, WQC^ (ug/L)
39.4
24.7
18.9
10.1
1.3
26
16.5
12.7
7.0
1.3
13 .
8.5
6.7
4.0
1.3
6.5
4.5
3.7
2.5
1.3
2.6
2.1
1.9
1.6
1.3
1.3
1.3
1.3
1.3
1.3
If the value offint, the intermittent exposure fraction of the month, is assigned a value less
than 1 day, then the intermittent criterion element value could exceed water concentrations that
have been shown to be acutely toxic to sensitive species in 2- or 4-day toxicity tests (compiled in
U.S. EPA 2004). Because the concentrations that would be acutely toxic in exposures of less
than 1 day might not be much greater than those observed to be toxic in 2-4 day exposures, the
intermittent fraction of the month should not be assigned a value less than 0.033, corresponding
to 1 day.
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5 National Criterion for Selenium in Fresh Waters
The available data indicate that freshwater aquatic life would be protected from the toxic
effects of selenium by applying the following four-part criterion:
1. The concentration of selenium in the eggs or ovaries offish does not exceed 15.2 mg/kg,
dry weight; l
2. The concentration of selenium (a) in whole-body offish does not exceed 8. 1 mg/kg dry
weight, or (b) in muscle tissue offish (skinless, boneless fillet) does not exceed 11.8
mg/kg dry weight; 2
3. The 30-day average concentration of selenium in water does not exceed 4.8 |ig/L in lotic
(flowing) waters and 1.3 |ig/L in lentic (standing) waters more than once in three years
on average;
4. The intermittent concentration of selenium in either a lentic or lotic water, as appropriate,
1,1 Tnjr\r 30-ay grn.nt ,1 • ,1
does not exceed WQCint = - - - - - more than once in three years on
Tint
average.3
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Table 15. 2014 External Peer Review Draft Freshwater Selenium Ambient Water
Quality Chronic Criterion for Aquatic Life.
Media
Type
Criterion
Element
Magnitude
Duration
Frequency
Fish Tissue
Egg/Ovary1
15.2mg/kg
Instantaneous
measurement5
Never to be
exceeded
Fish Whole
Body or
Muscle2
8.1 mg/kg
whole body
or
11. 8 mg/kg
muscle
(skinless,
boneless filet)
Instantaneous
measurement5
Never to be
exceeded
Water Column3
Monthly
Average
Exposure
1.3 |ig/L in
lentic aquatic
systems
4.8 jig/Linlotic
aquatic systems
30 days
Not more than
once in three
years on
average
Intermittent Exposure4
WQCint =
WQC30_day - Cbkgrnd(1 - f int)
f int
Number of days/month with
an elevated concentration
Not more than once in three
years on average
Overrides any whole-body, muscle, or water column elements when fish egg/ovary
concentrations are measured.
2 Overrides any water column element when both fish tissue and water concentrations are
measured.
3 Water column values are based on dissolved total selenium in water.
4 Where WQCso-dayis the water column monthly element, for either a lentic or lotic system, as
appropriate. Cbkgmd is the average background selenium concentration, and f;nt is the fraction of
any 30-day period during which elevated selenium concentrations occur, with f;nt assigned a
value >0.033 (corresponding to 1 day).
5 Instantaneous measurement. Fish tissue data provide point measurements that reflect integrative
accumulation of selenium over time and space in the fish at a given site. Selenium concentrations
in fish tissue are expected to change only gradually over time in response to environmental
fluctuations.
EPA recommends that states and tribes adopt into their water quality standards, is a
selenium criterion that includes all four elements, and expressing the four elements as a single
criterion composed of multiple parts, in a manner that explicitly affirms the primacy of the
whole-body or muscle elements over the water column element, and the egg-ovary element over
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any other element. The magnitude of the fish egg-ovary element is derived from analysis of the
available toxicity data. The magnitudes of the fish whole-body element and fish muscle elements
are derived from the egg-ovary element coupled with data on concentration ratios among tissues.
The magnitudes of the water column elements are derived from the egg-ovary element coupled
with bioaccumulation considerations. Inclusion of the fish whole-body or fish muscle element
into the selenium criterion ensures the protection of aquatic life when fish egg or ovary tissue
measurements are not available, and inclusion of the water column elements intothe selenium
criterion ensures protection when neither fish egg-ovary nor fish whole-body or muscle tissue
measurements are available.
To assure that the contribution of short-term exposures to the bioaccumulation risks is
accounted for in all situations, EPA is also recommending that the selenium intermittent
exposure element be included in the selenium criterion, as noted above. However, EPA is not
recommending a separate acute criterion derived from the results of toxicity tests having water-
only exposure, because selenium is bioaccumulative and toxicity primarily occurs through
dietary exposure. If there are rare instances where selenium sources could cause acute effects
without also exceeding the selenium chronic criterion outlined above, a pollution control
authority could establish a site-specific criterion to protect from those effects.
EPA recommends that when states implement the water quality criterion for selenium
under the NPDES permits program, states should establish additional procedures due to the
unique components of the selenium criterion expressions. Where states adopt the selenium water
column concentration criterion element values only for conducting reasonable potential (RP)
determinations and establishing water quality-based effluent limitations (WQBELS) per 40 CFR
122.44(d), existing implementation procedures used for other acute and chronic aquatic life
protection criteria would be appropriate. However, if states also decide to adopt the selenium fish
tissue criterion element values for NPDES permitting purposes, additional state WQS
implementation procedures (IPs) will be needed to determine the need for and development of
WQBELs necessary to ensure attainment of the fish tissue criterion element(s).
Protection of Downstream Waters
EPA regulations at 40 CFR 131.10(b) provide that "[i]n designating uses of a waterbody
and the appropriate criteria for those uses, the state shall take into consideration the water quality
standards of downstream waters and ensure that its water quality standards provide for the
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attainment and maintenance of the water quality standards of downstream waters." Especially in
cases where downstream waters are lentic waterbody types (e.g. lakes, impoundments), or harbor
more sensitive species, a selenium criterion more stringent than that required to protect in-stream
uses may be necessary in order to insure that water quality standards provide for the attainment
and maintenance of the water quality standards of downstream waters.
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6 Site-specific Criteria
All four elements of the selenium criterion can be modified to reflect site-specific
conditions where the scientific evidence indicates that different values will be protective of
aquatic life and provide for the attainment of designated uses.
Since the fish egg-ovary criterion element is based on toxicity data, a modification of that
element can be done by applying the Recalculation Procedure (U.S. EPA 2013a) to edit the
species toxicity database to reflect taxonomic relatedness to the site assemblage, while
recognizing tested surrogates for untested resident species.
However, species in the national data set that are not present at a site should not be
deleted from the data set if the species serves as a surrogate for other species known or expected
to be present at a site. Confidence in the applied tissue criterion element can be improved by
further toxicity testing offish species resident at the site. The most relevant testing would
measure the survival and occurrence of deformities in offspring of wild-caught female fish to
determine an ECio for selenium in the eggs or ovaries (e.g., following Janz and Muscatello
2008).
Using either the EPA national recommended egg-ovary, whole-body, or muscle criterion
concentration element or a site-specific egg-ovary, whole-body, or muscle criterion element,
translation of the fish tissue criterion to a water concentration can be performed in a manner that
accounts for site-specific conditions. Appendix I provides a step-wise process for deriving each
parameter used in Equation 18 to perform a site-specific translation. These steps include:
1. selection of a target fish species,
2. determining the primary food source for the target species,
3. determining the appropriate TTF values,
4. determining the appropriate EF value, and
5. determining the appropriate CF value.
Appendix I also provides information on how to obtain the site-specific information for each
step in the process. Other scientifically defensible translations, including traditional
Bioaccumulation Factors (BAFs), may also be appropriate.
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Where sensitive aquatic-dependent (e.g., bird) species are known to exist, states should
consider developing site-specific criteria based on data on such species.
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7 Effects Characterization
7.1 Fish
7.1.1 Principles for Using Studies for which ECmS Cannot Be Calculated
When the data from an acceptable chronic test met the conditions for logistic regression
analysis, the ECio. When data did not allow calculation of ECs but did allow calculation of
closely spaced NOECs and LOECs, then the NOEC was used to approximate the ECio. No
NOEC values were used in calculating the numeric value of the criterion.
When significant effects were observed at all treatment concentrations, such that no
treatment concentration was classified as a NOEC, then the chronic value was assigned as "less
than" (<) the lowest tested concentration. When no significant effects were observed at any
concentration, such that no treatment concentration was defined as an LOEC, then the chronic
value was assigned as "greater than" (>) the highest tested concentration.
A number of the chronic values in Sections 4.1.1 and 7.1.3 (reproductive effects) and in
Section 7.1.8 (nonreproductive effects) include a "greater than" (>) or "less than" (<) sign
because of an inability to resolve an exact value when all exposure concentrations of a study
yielded either too little or too much effect to provide a point estimate of a chronic value. The
decision to use chronic values with a "greater than" or "less than" sign in calculating an SMCV
followed a rule based on whether these values add relevant information to the mean, as described
below. None of these values were used in this assessment to derive the criteria values.
^^
Evaluation Approach
• Neither a low "greater than" value nor a high "less than" value were used to
calculate the SMCV;
• Both a low "less than" value and a high "greater than" value were included in the
SMCV calculation. However, none of these values were used in this assessment to
calculate the criteria values.
For example, a chronic value reported here as ">15 mg Se/kg" is ignored if the tentative
SMCV is 20 mg Se/kg. The ">15 mg Se/kg" value indicates that no significant effects were
observed at the study's highest tested concentration of 15 mg Se/kg. As this is consistent with
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what would be expected if the SMCV were 20 mg Se/kg, it provides no information to support
modifying the SMCV. However, a different study showing no effects at its highest tested
concentration and yielding the value ">25 mg Se/kg" is not consistent with an SMCV of 20 mg
Se/kg, and indicates that the ">25 mg Se/kg" value provides information for modifying the mean
upwards. Conversely, a chronic value reported here as "<15 mg Se/kg" indicates that significant
effects were observed even at the study's lowest tested concentration of 15 mg Se/kg. As this is
not consistent with a 20 mg Se/kg SMCV, it indicates the utility of the "<15 mg Se/kg"
information for modifying the SMCV downwards. On the other hand, a value reported here as
"<25 mg Se/kg" would not be used to recalculate a 20 mg Se/kg SMCV. The intent of the
approach is to use all quality information that is relevant and appropriate for calculating the
SMCVs.
7.1.2 Reproductive Effects in Catfish (Ictaluridae)
Some important families offish are not represented in the effects assessment, such as the
catfish family (Ictaluridae) . In their compilation of egg-ovary versus whole-body ratios,
Osmundson et al. (2007) found comparatively high concentrations of selenium in egg-ovary
compared to whole body in black bullhead, Ameiurus melas, which are related to the Ictaluridae.
This raises a question about the potential risks of reproductive effects in this species and possibly
in related Ictaluridae. In addition to this concern about how much selenium such species may
accumulate in their eggs, U.S. Fish and Wildlife Service (2005) has suggested that offspring of
channel catfish (Ictaluruspunctatus) and related species might be affected at unusually low egg
concentrations. This is based on results of a study in which adult female catfish were injected
with seleno-L-methionine (Doroshov et al. (1992b)). Effects were found in the offspring at egg
concentrations below levels observed in other studies in Section 4.1.2 and Appendix C. These
data were not included in derivation of the criteria because the injection route of exposure is not
an acceptable experimental protocol for studies used in criteria derivation due to its difference
from exposure routes in the environment (water column and diet).
In the absence of valid tests yielding an Ictaluridae ECio or chronic value, EPA evaluated
the potential vulnerability of the taxonomic group that includes catfish by examining
comparative fisheries observations of Ictaluridae and Centrarchidae sharing the same selenium-
contaminated water body. Crutchfield (2000) reports results of annual cove rotenone sampling
performed from 1982 to 1997 in Hyco Reservoir, North Carolina. The sampling was begun after
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centrarchid populations in this reservoir had collapsed due to the release of ash pond selenium
from a coal-fired power plant. The plant began operating a dry fly ash handling system in
January 1990, thereby eliminating the aquatic discharge of selenium; the sampling continued
through the recovery period.
Crutchfield (2000) reports abundance data (kg/ha) for 20 fish taxa, including four
Ictaluridae and three Centrarchidae. These data were examined to determine the relationship
between the Ictaluridae and the selenium-affected Centrarchidae populations. The correlation
matrix between annual measured abundance of the seven taxa is shown below in Table 16.
Correlation with the reciprocal of measured average concentrations of selenium in invertebrates
is also shown. Because the reciprocal of the selenium concentration is used, a positive correlation
means that abundance decreases as selenium concentration increases. Conversely, a negative
correlation means abundance decreases as selenium concentration decreases.
Table 16. Correlation matrix (values of r) for Ictaluridae and Centrarchidae
abundance and for selenium food chain contamination for the Hyco Reservoir data
reported by Crutchfield (2000).
Channel catfish
White catfish
Black bullhead
Ameiurus spp.
Bluegill
Largemouth bass
Pomoxis spp.
(crappie)
1 + Inverteb.
Se Cone.
Ictaluridae
Channel White Flat Ameiurus
catfish catfish bullhead spp.
1.00 -0.36 0.18 0.68
-0.36 1.00 0.02 -0.32
0.18 0.02 1.00 0.40
0.68 -0.32 0.40 1.00
0.08 -0.31 0.32 0.22
-0.33 -0.24 -0.08 -0.24
-0.08 -0.15 0.08 -0.05
-0.44 -0.06 -0.03 -0.31
Centrarchidae
Large- Pomoxis
mouth spp.
Bluegill bass (crappie)
0.08 -0.33 -0.08
-0.31 -0.24 -0.15
0.32 -0.08 0.08
0.22 -0.24 -0.05
1.00 0.78 0.76
0.78 1.00 0.78
0.76 0.78 1.00
0.80 0.92 0.69
1 + Inverteb.
Se Cone
-0.44
-0.06
-0.03
-0.31
0.80
0.92
0.69
1.00
The centrarchid abundances are well correlated with each other and are closely related to
selenium concentrations in the food chain with fish abundance decreasing as selenium
concentrations increase. Ictaluridae abundances, however, are unrelated either to the selenium-
sensitive centrarchid abundances or to the selenium concentrations in the food chain.
Observations of selenium contaminated Belews Lake accord with the above; Young et al. (2010)
indicate that out of as many as 29 resident species documented prior to contamination, only
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common carp, catfish, and fathead minnows remained after contamination. Based on field
observations, catfish and bullheads (Ictaluridae) thus appear to be less vulnerable than the fish
taxa most at risk (e.g., Centrarchidae, Salmonidae) in selenium-contaminated water bodies,
contrary to what might be suggested by the Doroshov et al. (1992) injection study.
7.1.3 Reproductive Studies Not Used in the Numeric Criterion Derivation
Pimephales promelas (fathead minnow)
GEI Associates (2008) collected fathead minnows from three sites in Colorado of
moderate to high selenium exposure and transported them to the laboratory for spawning and
subsequent assessment of embryo and larval development. Egg production, fertilization success,
embryo mortality and larval deformities from the offspring of the wild caught fish were
compared to offspring spawned from fathead minnows obtained from a commercial fish supplier.
Mean selenium concentrations in field-collected adult females ranged from 9.17 to 44.53 mg/kg
dw whole body; the mean selenium concentration in control females from the commercial
supplier was 2.86 mg/kg dw whole body. The response measurements for the embryo assessment
endpoints were variable and lacked a relationship with selenium exposure. Consequently, the
results of this study could not support a reliable estimate of an effect concentration, and chronic
values are not given in this document. A detailed summary of the GEI Associates (2008) study is
given in Appendix D. These values were not used in the criterion derivation.Salmonidae
Oncorhynchus clarki (cutthroat trout)
Kennedy et al. (2000) reported no significant differences in mortality and deformity in
eggs, larvae, and fry from wild-caught cutthroat trout between a reference and an exposed site
(Fording River, British Columbia, Canada). The observations were made on eggs reared in well
water from spawning age females collected from the two locations (N = 17 and 20, respectively)
and fertilized by one male collected at each site. The mean selenium content in eggs from fish
collected from the reference site was 4.6 mg/kg dw and from fish collected from the Fording
River was 21.2 mg/kg dw. The chronic value for eggs is >21.2 mg Se/kg dw. These values were
not used in the criterion derivation because they represent high "greater than" values, as
discussed above, and provide no additional important quantitative data for the analyses.
Hardy (2005) fed cutthroat trout experimental diets containing a range of
selenomethionine (0-10 mg/kg dw) for 124 weeks. No significant growth or survival effects were
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observed in the adult fish over the 124 weeks. The whole body concentration reached 12.5 mg/kg
dw selenium after 44 weeks. Embryo-larval observations (percent hatch and percent deformed)
were not related to whole body selenium concentrations in the spawning females (9.37 mg/kg
dw) fed the selenium-laden diet for 124 weeks. The concentration of selenium in eggs from these
females was 16.04 mg/kg dw. For this study the chronic value, an unbounded NOEC, is thus
>16.04 mg Se/kg dw in eggs. This value was not used in the criterion derivation.
Salvelinus fontinalis (brook trout)
Holm et al. (2005) collected spawning brook trout from streams with elevated selenium
contaminated by coal mining activity and from reference streams in 2000, 2001 and 2002.
Similar to procedures described by these authors for rainbow trout, above, fertilized eggs were
monitored in the laboratory for percent fertilization, deformities (craniofacial, finfold, and spinal
malformations), edema, and mortality. Embryos from the contaminated stream had on average a
higher frequency of craniofacial deformities than fry from the reference stream (7.9% for the
contaminated stream compared to 2.1% in the reference stream). Although this increased rate of
craniofacial deformities was calculated to be statistically significant when compared across sites,
the Abbott-adjusted effect is only 6% and is thus below the 10% effect represented by an ECio.
But more important, when comparing across adult females (the more reliable analysis for
selenium reproductive toxicity studies of this type, and the one used to obtain the related rainbow
trout ECio for these authors' studies), there is no apparent relationship between brook trout
craniofacial deformities and exposure across a broad range of concentrations, as illustrated in
Appendix C. An environmentally conservative estimate of the NOEC might be considered to be
the average concentration of selenium in eggs from the high exposure site (Luscar Creek), >7.78
mg Se/kg ww or >20.5 mg Se/kg dw using the 61.2% moisture content for rainbow trout eggs
cited above. However, the effect threshold appears to be substantially higher based on the
absence of any consistent concentration-response relationship up to the maximum observed egg
concentration of 18.9 mg Se/kg ww or 48.7 mg Se/kg dw, as shown in the Appendix C graphs.
Given the point estimate ECIO available for the related species, Salvelinus malma (Dolly
Varden, Section 4.1.1), the "greater than" chronic value for brook trout are not used to obtain the
GMCV, in accordance with the principles of Section 7.1.1.
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Lepomis machrochirus (bluegill)
Applicable chronic reproductive data for bluegill can be grouped by exposure type: field
and laboratory. In some field studies, chronic value estimates were "less than" fairly high
selenium concentrations (Bryson et al. 1984, 1985a; Gillespie and Baumann 1986). This low
resolution is due to the observed effect occurring at a single observed high exposure
concentration relative to a reference condition. In the Bryson et al. (1984, 1985a) and Gillespie
and Baumann (1986) studies, the artificially crossed progeny of females collected from a
selenium contaminated reservoir (Hyco Reservoir, Person County, NC) did not survive to swim-
up stage, irrespective of the origin of milt used for fertilization. Measured waterborne selenium
concentrations prior to the experiments ranged from 35 to 80 ug/L. The ovary tissue selenium
concentration associated with this high occurrence of mortality of hatched larvae was <30 mg/kg
dw tissue, as reported by Bryson et al. (1985a), and <46.30 mg/kg dw tissue, as reported by
Gillespie and Baumann (1986). In the case of the latter, nearly all swim-up larvae from the Hyco
Reservoir females were edematous, none of which survived to swim-up.
Bryson et al. (1985b) examined percent hatch and percent swim-up larvae from spawns
using fish collected from Hyco Reservoir and a control site. There were no differences in the
Hyco measurements relative to the control. The concentration of selenium in the liver of the
parental Hyco bluegill was 18.6 mg/kg dw. The chronic values for this embryo-larval
development test was >18.6 mg Se/kg dw liver. The high "less than" and low "greater than"
chronic values obtained from Bryson et al. (1984, 1985a, 1985b) and Gillespie and Baumann
(1986) were not used in the SMCV calculation because these values are consistent with and yet
provide no numeric basis for modifying the SMCV obtained from the ECios.
7.1.4 Salmo GMCV: EPA Re-analysis of a Key Study Used in Criterion Derivation
The lowest GMCV in the reproductive effects dataset is for Salmo; the calculated egg-
ovary criterion element is more sensitive to this value than to any other. However, several
reasonable ECios can be calculated from the Formation Environmental (2011) data for brown
trout. Because of the importance of this data for the numeric criterion calculation, EPA
conducted a careful and thorough reanalysis of the study data and subjected its reanalysis to
independent, external peer review (ERG 2012), to confirm the validity and scientific robustness
of the approach taken by EPA in the reanalysis and use of the reanalyzed data. Those
assessments are superseded by this document's reanalysis of a more complete enumeration of the
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deformity counts provided by AECOM (2012), provided in Appendix C. Below is a summary of
key issues considered in the EPA reanalysis of the data and EPA's conclusions regarding the
appropriate endpoint and effect concentration to use in the criteria derivation.
As described in detail in Appendix C, Formation Environmental (2011) evaluated
survival and deformities in the offspring of wild-caught brown trout having a range of exposure
levels. The evaluation of offspring had two key phases, hatch to swim-up, and swim-up to 15-
days post-swim-up. Based on its reanalysis of the data, EPA concludes that the best-supported
ECioS fall into the range, 15.91 - 21.16 mg Se/kg (dw egg). Uncertainties in the ECio appropriate
for this species stem from the observed high background deformity rates and by a lab accident
causing overflow loss of some organisms from several aquaria during the post-swim-up portion
of the test. This accident occurred when aquaria drainpipe filters became clogged with uneaten
food.
Deformities for the full test (hatch to 15-days post swim-up), calculated assuming all
overflow-missing individuals were deformed, yield the lowest of the above mentioned ECios,
15.91 mg Se/kg from Figure 13b, but the following factors may be noted:
a) High background deformity rates in unexposed, hatchery-reared fish (points with log
concentration <0.2) as shown in Figure 15a and b, increase the uncertainty in the 15.91
mg Se/kg EC 10 of Figure 13b.
b) The worst-case assumption that all individuals lost in the overflow were deformed
depresses the above ECio. An assumption that the lost individuals had the same frequency
of deformity as the others yields an ECio of 18.36 mg Se/kg (Figure 13a).
c) In Figure 13b, the fitted line's under-prediction of the fraction empirically observed to be
normal at 17.7 and 20.5 mg Se/kg (log ~ 1.3) suggests that the 15.91 mg Se/kg ECio may
be a low estimate. The under-prediction of the fraction normal in that key exposure range
is caused by TRAP applying a shallower slope so as to reduce its error in predicting the
fraction normal in the exposure range of >36 mg Se/kg, a range of less interest for
deformities because of the failure of all individuals to reach the swim-up stage at such
high exposures. When the combined survival and deformities are considered, and failing
to swim-up by the end of the test is equated with failing to survive (while applying the
same worst-case assumption that individuals lost in the overflow were dead, dying, or
deformed), the ECio rises to 20.65 mg Se/kg, as shown in Figure 15f Having no partial
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effects at exposures >26.8 mg Se/kg, allows a steep slope, which then allows an exact fit
to the responses at 17.7 and 20.5 mg Se/kg. It is for this reason that combining observed
effects, survival and deformities, has the unexpected result of increasing rather than
decreasing the ECio.
d) Survival and deformity ECios are very close in magnitude. Assuming individuals lost in
the overflow were dead or dying, the survival ECio for the full test is 16.79 mg Se/kg
(Figure 13d), only 6% higher than the corresponding deformity ECio of 15.91 mg Se/kg.
e) The assumption about organisms lost to the overflow affects the survival ECio as it did
the deformity ECio. If the health of missing organisms were assumed to be the same as
those remaining, the survival ECio would be 20.40 mg Se/kg (Figure 13c). The peer
review conducted by ERG (2012) did not provide a consensus on expectations of whether
less healthy organisms were more likely to have been lost in the overflow.
f) For combined survival and deformities, assuming the health of overflow-missing
organisms to be the same as those remaining, the ECio is 21.16 mg Se/kg (Figure 13e).
The use of the lowest of the above values (15.91 mg Se/kg) for setting the chronic value
for brown trout provides a greater margin of protection than would one of the higher
values. Were the Salmo GMCV set at the geometric mean of the above six values for the
test, 18.77 mg Se/kg, the FCV would be 17.3 mg Se/kg.
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Optimistic
Worst Case
E
o
•t
1.0
0.8
0.6
0.4
0.2
Q.
O
0.0
r.
-0.2 0.2 0.6 1.0 1.4 1.
log (mg Se/Kg egg dw)
1
~
-------
7.1.5 Influence of Curve-fitting on Calculation oi Lepomis GMCV
The Lepomis GMCV, the second lowest in the Species Sensitivity Distribution, has been
calculated to be 18.41 mg Se/kg, based on ECios from three studies with bluegill, 20.05 mg
Se/kg from Doroshov et al. (1992), 24.55 mg Se/kg from Coyle et al. (1993), and 12.68 mg
Se/kg from Hermanutz et al. (1992, 1996). The 12.68 value is low compared to the other studies
with bluegill or any other species. It stems from an environmentally conservative fitting of a
sigmoid curve to the data, as illustrated in Figure 14. This was the model fit having lowest error,
when measured vertically.
The dashed line of Figure 14 mimics an alternate orthogonal regression approach
effectively reduceing combined vertical and horizontal errors. It results in an ECio of 18.40 mg
Se/kg. Replacing 12.68 with 18.40 mg Se/kg in calculating the Lepomis GMCV would only
change the Lepomis chronic value from 18.41 to 20.84 mg Se/kg. This would change the FCV
from 15.2 to 15.6 mg Se/kg. The recommended FCV of 12.68 is based on the more conservative
curve-fitting approach.
1 ?0
= 10° -
1
^ 80
•o
*j
in
"TT fin
E
01
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Ovary Se |mg/kg dw)
Figure 14. Fitting the Hermanutz et al. (1992,1996) data to yield (a) the 12.68 mg Se/kg
ECio (dotted line) with TRAP measuring error vertically, and (b) a possible alternative
18.40 mg Se/kg ECio (dashed line), reducing horizontal error by running TRAP after
combining two points averaging 49.85% normal (absence of edema) into one point at their
geometric mean exposure of 24.56 mg Se/kg, and combining the two points averaging
19.3% normal into one at their geometric mean exposure, 24.45 mg Se/kg.
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Draft Document
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7.1.6 Impact of Number of Tested Species on Criterion Derivation
Many of the species used for testing the toxicity of selenium are those observed to be
affected at contaminated sites or otherwise suspected to be particularly sensitive. Six of the 8
minimum data requirements were met, and the other 2 (for planktonic and benthic crustaceans)
were waived (see section 3.7.1.2. Of the N=14 genera used for the calculation of the criterion,
nine are fish, which in general are more sensitive than invertebrates. Of the nine fish genera, five
are either salmonids or centrarchids. Had a broader array of expected insensitive taxa been
included, better reflecting the taxonomic composition of real-world sites, then the four most
sensitive genera would not likely change, but N would increase.
Nevertheless, the criterion calculation for selenium is not sensitive to the value of N.
Setting N=20 would only raise the criterion from 15.2 mg Se/kg to 16.0 mg Se/kg. Setting N=25
would raise the egg- ovary criterion element to 16.7 mg Se/kg. This insensitivity occurs because
the four lowest GMCVs are closely spaced, such that the calculated egg-ovary criterion element
is never distant from the lowest GMCV.
7.1.7 Conversions between Concentrations in Different Tissues
Chapman et al. (2009, 2010) note that risk characterization may start with selenium
concentrations in any environmental compartment, but uncertainty about potential adverse
effects is lowest when the concentrations in reproductive tissue are known. Because the egg-
ovary (EO) criterion element is derived from studies measuring the reproductive tissue
concentrations, it has greater certainty than the muscle (M) and whole-body (WB) criteria
element concentrations. As indicated by the information tabulated in Section 4.1.5, to obtain the
muscle criterion element concentration, conversion from EO to M involved dividing by the
EO/M ratios measured in each species except desert pupfish, where the EO/M ratio was the
measured desert pupfish EO/WB divided by the median generic fish M/WB ratio. As indicated in
Section 4.1.5, to obtain the whole-body criterion element value, conversion from EO to WB
involved dividing by EO/WB ratios measured in each taxa except Esox, Salvelinus, and
Oncorhynchus, where the EO/WB ratios were each of their own EO/M ratios times the median
generic fish M/WB ratio.
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7.1.8 Studies of Non-Reproductive Effects
This section presents laboratory-based dietary chronic exposure studies involving non-
reproductive endpoints. These studies do not involve effects on the offspring of exposed female
adults, and their results are not expressed as selenium concentrations in egg or ovary tissue.
Because selenium concentrations in whole body and muscle are generally lower than in egg and
ovary, with observed egg-ovary to whole-body ratios ranging from 1.3 to 7.4, and egg-ovary to
muscle ratios ranging from 1.0 to 5.8, whole-body and muscle effect concentrations cannot be
directly compared to egg-ovary effect concentrations. Non-reproductive effects were ultimately
determined to provide a less reliable basis for a criterion, in part because comparatively few of
such studies provided sigmoidal concentration-response curves.
V ^
Acipenseridae
^K 7
Acipenser transmontanus (white sturgeon)
Juvenile white sturgeon were exposed for 8 weeks to a series of 5 concentrations of
seleno-L-methionine added to an artificial diet (Tashjian et al. 2006). Survival was not affected
by selenium treatment with a mean survival rate of 99% across all groups. Fish fed the highest
three dietary treatments of selenium, 41.7, 89.8 and 191.1 mg Se/kg dw, exhibited significant
declines in growth assessed by body weight measurements. The ECio for reduction in body
weight is 15.08 mg Se/kg dw in whole body or 27.76 mg Se/kg dw muscle; the £€20 is 17.82 mg
Se/kg dw in whole body or 32.53 mg Se/kg dw muscle tissue. The criterion values derived in this
document that are based on reproductive endpoints are protective of the endpoint measured in
this non-reproductive study.
Cyprinidae
Pogonichthys macrolepidotus (Sacramento splittail)
Teh et al. (2004) exposed juvenile Sacramento splittail (7 months-old) to 8 levels of
dietary selenium, 0.4 (no added selenium), 0.7, 1.4, 2.7, 6.6, 12.6, 26.0, and 57.6 mg/kg.
Selenium was added to the diet via selenized yeast which was diluted with Torula yeast
(inactive) to attain the target levels. Mortality, growth, histopathology, deformities and selenium
content in muscle and liver were observed or measured after 5 and 9 months of exposure. The
appearance of deformities was the most sensitive endpoint. The authors determined the
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occurrence of deformities was higher in fish fed 6.6 and 12.6 mg Se/kg in their diet; however,
such pathology was examined for only 15 of the 120 individuals per treatment, and a consistent
concentration-response relationship did not occur (i.e., no deformities in the high concentration).
The lack of a concentration-response relationship for the incidence of deformities has also been
observed in another study. Crane et al. (1992) exposed a European species of perch, Perca
fluviatilis to three aqueous and dietary selenium treatments in experimental ponds for 288 days
up through spawning. Crane et al. (1992) found an increased occurrence of deformities in
embryos and larvae in the lowest selenium treatment relative to the control, but a decrease in the
middle treatment. No hatching occurred in the high treatment. Teh et al. (2004) proposed several
physiological mechanisms to explain the lack of a dose-response relationship, but it appears that
the underlying mechanism is not understood at this time. Toxicity tests with unusual dose-
response relationships are typically not considered for criteria derivation, but since another assay
(Crane et al. 1992) observed a similar relationship, the Teh et al. (2004) study with P.
macrolepidotus is included. Using prevalence of deformities as the endpoint, the NOEC, LOEC
and MATC (chronic value) in muscle tissue are 10.1, 15.1 and 12.34 mg Se/kg dw, respectively.
The critieron value in muscle tissue, based on the reproductive ECio, is 11.8 mg Se/kg dw.
Appendix C provides further details on the study results and an approximate estimate of their
relationship to egg-ovary and whole-body concentrations. Teh et al. (2004) is the only study in
which deformities developed in fish that were not exposed to selenium from their mothers'
ovaries. The selenium criteirion values derived based on reproductive endpoints are protective of
the endpoint measured in this non-reproductive study.
Pimephales promelas (fathead minnows)
Non-reproductive chronic values for fathead minnows were derived from two laboratory-
based studies. These studies (Bennett et al. 1986 and Dobbs et al. 1996) involved exposing algae
to selenium (either as sodium selenite or sodium selenate) in water, and subsequently feeding the
algae to rotifers which were in turn fed to fathead minnows. In the Bennett et al. (1986) study,
larval fathead minnows were fed control rotifers (cultured in chambers without selenium
containing algae) or selenium-contaminated rotifers (cultured in chambers with selenium
containing algae previously exposed to sodium selenite in the water) in three separate
experiments lasting 9 to 30 days. The different experiments were distinguished by 1) the day
selenium-laden rotifers were first fed; 2) the day selenium-laden rotifers were last fed; and 3) the
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age of larvae at experiment termination. The results from the three experiments reported by
Bennett et al. (1986) were conflicting. Larval growth was significantly reduced at larval whole-
body selenium concentrations of 43.0 mg Se/kg dw in the first experiment and 51.7 mg Se/kg dw
in the second experiment, but was slightly but not significantly reduced at 61.1 mg Se/kg dw in
the third experiment (see Appendix C). Following the approach of Section 7.1.1, the geometric
mean of these three values, 51.40 mg Se/kg dw, is the chronic value for this study.
Dobbs et al. (1996) used a test system similar that of Bennett et al (1986) (described
above). Larval fathead minnows were exposed to the same concentrations of sodium selenate in
the water as their prey (rotifers), but also received additional selenium from the consumption of
the selenium-contaminated rotifers. In this study, the fathead minnows did not grow well at
concentrations exceeding 108.1 jig Se/L in water, and they survived only to 11 days at selenium
concentrations equal to or greater than 393.0 |ig/L in the water (75 mg Se/kg dw in the diet, i.e.,
rotifers). The LOEC for retarded growth (larval fish dry weight) in this study was <73 mg Se/kg
dw tissue.
A third laboratory study, by Ogle and Knight (1989), examined the chronic effects of
elevated foodborne selenium on growth and reproduction of fathead minnows. Juvenile fathead
minnows were fed a purified diet mix spiked with inorganic and organic selenium in the
following percentages: 25 percent selenate, 50 percent selenite, and 25 percent seleno-L-
methionine. The pre-spawning exposure lasted 105 days using progeny of adult fathead minnows
originally obtained from the Columbia National Fishery Research Laboratory, as well as those
obtained from a commercial fish supplier. After the 105 day exposure period, a single male and
female pair from each of the respective treatment replicates were isolated and inspected for
spawning activity for 30 days following the first spawning event of that pair. There was no effect
from selenium on any of the reproductive parameters measured, including larval survival, at the
dietary concentrations tested (5.2 to 29.5 mg Se/kg dw food). Sub-samples of larvae from each
brood were maintained for 14 days post-hatch and exhibited >87.4 percent survival. The pre-
spawning adult fish fed a mean dietary level of 20.3 mg Se/kg dw exhibited a significant
reduction in growth compared to controls (16 percent reduction), whereas a nonsignificant
reduction in growth (7 percent) occurred in the fish fed 15.2 mg Se/kg dw. The chronic value, as
determined by the geometric mean of the NOEC and the LOEC measured at 98 days post-test
initiation, was 17.57 mg Se/kg expressed as the above dietary concentrations, and 5.961 mg
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Se/kg dw as fathead minnow whole-body tissue. The concentration-response relationship, as
indicated by the study data presented in Appendix D, was uniformly shallow, not resembling the
sharp sigmoidal function characteristic of most selenium response curves.
Since Ogle and Knight reported that food in the higher selenium concentrations remained
uneaten and fish were observed to reject the food containing the higher selenium concentrations,
the authors suggested that the decreased growth was caused by a reduced palatability of the
seleniferous food items, which contained unnatural percentages of inorganic selenium (Fan et al.
2002). This is a common observation also noted by Hilton and Hodson (1983) and Hilton et al.
(1980) and apparent in Coughlan and Velte (1989). It is here interpreted to be an artifact of
unrealistic spiking of the diet with inorganic selenium in this early experimental protocol. That
is, in the real world it is not expected that avoidance of food items that were unpalatable because
of excessive selenium would be either a mechanism by which selenium causes effects or a
mechanism by which organisms can avoid exposure. (See Janz et al. (2010) for a more complete
discussion of selenium's mechanism of toxicity.) Given the no observed effect on larval survival
and the apparent non-toxicological effect on growth in the Ogle and Knight study, a chronic
value for this study is not included.
Catostomidae
Xyrauchen texanus (razorback sucker)
Two non-reproductive endpoint studies have been done with the endangered razorback
sucker. In the first study, Beyers and Sodergren (200la) exposed larval razorback suckers for 28
days to a range of aqueous selenate concentrations (6.12, 25.4, 50.6, 98.9, and 190.6 ug/L) and
respectively fed them a range of selenium in their diet (rotifers containing <0.702, 1.35, 2.02,
4.63, and 8.24 mg/kg dw). Reflecting the lack of effects on survival and growth in any exposure,
the chronic value for this study, based on selenium measured in the larvae at the end of the test,
is >12.9 mg Se/kg dw.
In a second study, Beyers and Sodergren (200Ib) exposed larval razorback suckers to a
control water and three different site waters containing varying concentrations of selenium for 28
days. Two treatments were tested within each water type: fish fed rotifers cultured in the same
water type (site diet) and fish fed rotifers cultured in control water. There were no reductions in
survival or growth in fish exposed to both the site water and site diet compared to fish exposed to
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control water and control diet. There were, however, reductions in growth offish exposed to site
water/site food compared to the same site water and control food. The authors did not attribute
the effect on larval growth by the diet to selenium and cited several lines of evidence, including:
(1) there was not a dose-response relationship in the concentration of selenium in the food
(rotifers) and growth, nor in the concentration of selenium in the fish larvae and growth across
the three water types; and (2) water from the De Beque site promoted a significant reduction in
the growth offish exposed to site water/site food relative to site water/control food, but
contained low levels of selenium in the water (<1 ug/L) and in food (2.10 mg/kg dw) typically
lower than those that have been found to elicit effects. The chronic value for this study is >42 mg
Se/kg dw based on the whole body concentration of selenium in the larval razorback suckers
exposed to North Pond site water.
Two similar studies were conducted in 1996 and 1997 to determine effects of site water
and site food, both contaminated with selenium on the razorback sucker (Hamilton et al.
2001a,b; published later in a peer-reviewed journal in 2005, see Hamilton et al. 2005 a,b,c). Both
studies show marked effects of selenium on survival of razorback sucker larvae exposed to
contaminated food and to a lesser extent, contaminated water. Although the data convincingly
demonstrate effects to larval survival from exposure to contaminated food, interpretation of the
results in the context of chronic criterion derivation is complex because of inconsistencies
between: 1) levels of selenium in the food and larvae relative to larval survival; 2) the time to
larval mortality relative to selenium in the diet and selenium in the larvae; and 3) levels of other
inorganic contaminants in food and water (possible organic contaminants were not measured).
Summaries of each of these two studies as well as a third study with razorback suckers
(Hamilton et al. 2005d) are presented in Appendix D.
Due to the confounding results, lack of dose-response within and among related studies,
and the uncertainty of the effect of other inorganic contaminants on larval response to the various
dietary and waterborne treatments, the data from these three studies for razorback sucker
(Hamilton et al. 2001a,b; Hamilton et al. 2005d) have not been included. A more detailed
explanation of why these studies were not included is given in Appendix D. Because of the
vastly different results between the Beyers and Sodergren studies and Hamilton et al. studies and
the inability to resolve the differences, SMCV and GMCV were not calculated for the razorback
sucker.
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Catostomus latipinnis (flannelmouth sucker)
Beyers and Sodergren (200la) exposed flannelmouth sucker larvae to a range of aqueous
selenate concentrations (<1, 25.4, 50.6, 98.9, and 190.6 ug/L) and fed them a range of selenium
in their diet (rotifers containing <0.702, 1.35, 2.02, 4.63, and 8.24 mg/kg dw, respectively).
There were no survival or growth effects observed after the 28 day exposure. The chronic value
based on the concentration of selenium measured in the larvae exposed to the highest test
concentration was >10.2 mg Se/kg dw.
Salmonidae
Oncorhynchus tshawytscha (Chinook salmon)
Hamilton et al. (1990) conducted a 90-day growth and survival study with swim-up
larvae fed one of two different diets. The first diet consisted of Oregon moist™ pellets where
over half of the salmon meal was replaced with meal from selenium-laden mosquitofish
(Gambusia affinis) collected from the San Luis Drain, CA (SLD diet). The second diet was
prepared by replacing half the salmon meal in the Oregon moist™ pellets with meal from low-
selenium mosquitofish (i.e., the same relatively uncontaminated mosquitofish that were used in
the control diet) and spiked with seleno-DL-methionine (SeMe diet). Analysis of the trace
element composition in the two different diets indicated that while selenium was the most toxic
element in the SLD diet, concentrations of boron, chromium, iron and strontium in the high-
selenium mosquitofish replacement diet (SLD diet type) were slightly elevated compared to the
replacement diet. These trace elements were, however, only 1.2 (e.g., iron) to 2.0 times (e.g.,
chromium) higher in the SLD diet than the SeMe diet, which contained the following measured
concentrations (dry weight basis) in the food: 10 mg boron/kg, 2.8 mg chromium/kg, 776 mg
iron/kg, and 48.9 mg strontium/kg.
During the test, survival of control Chinook salmon larvae (consuming food at
approximately 3 mg Se/kg dw) was 99 percent up to 60 days post-test initiation. Between 60 and
90 days of exposure, however, the control survival declined to 66.7% in the SLD test and to
72.5% in the test using the SeMe diet, indicating compromised health. Therefore, only data
collected up to 60 days post-test initiation were considered for analysis. Nevertheless, there
remains the possibility that even at 60 days, the control organisms were not healthy, although
overt signs of stress did not appear until later.
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For the SeMe diet, regression analysis of the 60-day growth data yielded a whole-body
ECio of 7.355 mg Se/kg dw and an £€20 of 10.47 mg Se/kg dw. For the SLD diet, regression
analysis of the 60-day growth data yielded a whole-body ECio of 11.14 mg Se/kg dw and an
EC2o of 15.73 mg Se/kg dw. Note: The San Luis Drain mosquitofish (comprising the Chinook
salmon's SLD diet) were not tested for contaminants other than certain key elements. Because
the San Luis Drain receives irrigation drainage from the greater San Joaquin Valley, there is a
possibility that the SLD diet might have contained elevated levels of pesticides, possibly a
confounding factor, although the SLD diet was less toxic than the SeMe diet.
Oncorhynchus mykiss (rainbow trout)
Hilton and Hodson (1983) reared juvenile rainbow trout on either a high (25 percent) or
low (11 percent) available carbohydrate diet supplemented with sodium selenite for 16 weeks.
Body weights, feed:gain ratios, and total mortalities were followed throughout the exposure
every 28 days. Tissues (livers and kidneys) were extracted for selenium analysis after 16 weeks.
By the end of the exposure, fish fed diets (low carbohydrate and high carbohydrate) with the
highest selenium concentrations (11.4 and 11.8 mg Se/kg dw food, respectively) exhibited a 45
to 48 percent reduction in body weight (expressed as kg per 100 fish) compared to control fish.
The authors attributed such results to food avoidance. With only two dietary exposure
concentrations and a control, these data were not amenable to regression analysis. The MATC
for growth of juvenile rainbow trout relative to the concentrations of selenium in liver tissue of
trout reared on the high carbohydrate seleniferous dietary type is the geometric mean (GM) of
21.00 mg Se/kg dw liver (NOEC) and 71.7 mg Se/kg dw liver (LOEC), or 38.80 mg Se/kg dw
liver. The calculated MATC for the same group of experimental fish exposed to selenium in the
low carbohydrate diet is 43.5 mg Se/kg dw liver tissue, which is the same MATC for trout
exposed for an additional 4 weeks based on the occurrence of nephrocalcinosis in kidneys (see
Hicks et al. 1984; Appendix C).
Hilton et al. (1980) employed a similar test design to that of Hilton and Hodson (1983) to
examine the narrow window at which selenium changes from an essential nutrient to a toxicant
affecting juvenile rainbow trout. The food consisted of a casein-Torula yeast diet supplemented
with selenium as sodium selenite. As discussed previously for the Ogle and Knight (1989) study
with fathead minnow, this represents an unrealistic fraction of inorganic selenium in the diet. The
experiment lasted for 20 weeks. During this time, the trout were fed to satiation 3 to 4 times per
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day, 6 days per week, with one feeding on the seventh day. Organs (liver and kidney) and
carcasses were analyzed for selenium from fish sacrificed at 4 and 16 weeks. No gross
histopathological or physiological effects were detected in the fish, although trout raised on the
highest dietary level of selenium (13.06 mg Se/kg dw food) had a significantly lower body
weight (wet basis), a higher feed:gain ratio, and higher number of mortalities (10.7; expressed as
number per 10,000 fish days). The MATC for growth and survival of juvenile rainbow trout
relative to the final concentrations of selenium in liver tissue is the geometric mean of the NOEC
(40 mg Se/kg dw liver) and the LOEC (100 mg Se/kg dw liver), or 63.25 mg Se/kg dw, both of
which hinge on accepting dietary spiking entirely with inorganic selenium as an acceptable
experimental protocol.
The non-reproductive GMCV for Oncorhynchus (both rainbow trout and Chinook
salmon) is 9.052 mg Se/kg dw whole body based on ECio value derived from the Hamilton et al.
(1990) study with Chinook salmon. The NOEC values for the rainbow trout studies conducted by
Hilton and Hodson (1983), Hilton et al. (1980), and Hicks et al. (1984) were not used in the
GMCV calculation because of the large difference between the NOEC and the LOEC values. If
adult fish contained whole-body selenium concentrations equal to 9.052 mg Se/kg dw, their egg-
ovary concentrations would be estimated to be 21.5 mg Se/kg dw when translated using the
factor 2.37. The criterion values derived based on reproductive endpoints are protective of the
endpoint measured.
Moronidae
^
Morone saxitilis (striped bass)
A non-reproductive chronic value for selenium was determined from a laboratory dietary
exposure conducted using yearling striped bass (Coughlan and Velte 1989). During the
experiment, the bass were fed contaminated red shiners (38.6 mg Se/kg dw whole body) from
Belews Lake, NC (treated fish) or golden shiners with low levels of selenium (1.3 mg/kg dw
whole body) purchased from a commercial supplier (control fish). The test was conducted in soft
well water and lasted up to 80 days. During the experiment, all fish were fed to satiation 3 times
per day. Control fish grew well and behaved normally. Treated fish behaved lethargically, grew
poorly due to a significant reduction in appetite, and showed histological damage, all eventually
leading to the death of animals. The final selenium concentration in muscle of treated striped
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bass averaged from 16.2 to 18.5 mg/kg dw tissue (assuming 78.4 percent moisture content),
which was 3.4 to 3.6 times higher than the final selenium concentrations in control striped bass,
which averaged 5.10 mg/kg dw tissue. The chronic value for this species was determined to be
<16.2 mg Se/kg dw in muscle tissue.
Centrarchidae
Lepomis macrochirus (bluegill)
Bryson et al. (1985b) conducted juvenile survival toxicity tests using hatchery bluegill
and various forms of selenium spiked to an artificial diet as well as a diet consisting of
zooplankton collected from Hyco Reservoir. There was no effect on length or weight of the
juvenile bluegill after 60 days of exposure. The highest concentration of selenium measured in
whole body of the juveniles in these tests was in the seleno-DL-cysteine-2X treatment (3.74 mg
Se/kg dw).
Cleveland et al. (1993) performed a 90-day diet-only laboratory exposure in which
juvenile bluegill were fed a range of selenomethionine concentrations added to Oregon moist™
pellets. The authors observed no significant effects on survival, but did report a very small but
apparently statistically significant decrease in the condition factor, K, from 1.3 at four
concentrations between 1.0 and 4.7 mg Se/kg dw whole body, to 1.2 at the two concentrations
7.7 and 13.4 mg Se/kg dw whole body. The condition factor (weight x 105/length3) is intended to
reflect a fish's reserves. In contrast to the studies of Ogle and Knight (1989), Hilton and Hodson
(1983), and Hilton et al. (1989), which appear to have involved an inorganic selenium food
palatability problem, this study did not use inorganic selenium in the diet. Nevertheless, given
that the reduction in K (1.3 to 1.2) is slight and shows no increasing effect between 7.7 and 13.4
mg Se/kg dw, thus not yielding a sigmoidal concentration-response curve to support an ECio
calculation, the chronic value for this study was estimated at >13.4 mg Se/kg dw in whole body
tissue.
Data from Lemly (1993a) indicate that over-wintering fish may be more susceptible to
the effects of waterborne and dietary selenium due to increased sensitivity at low temperature.
The author exposed juvenile bluegill in the laboratory to a single elevated exposure level,
waterborne (1:1 selenite:selenate; nominal 5 jig Se/L) and foodborne (seleno-L-methionine in
TetraMin; nominal 5 mg Se/kg dw food) selenium for 180 days. Tests with a control and the
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treated fish were run at 4°C and 20°C with biological and selenium measurements made every 60
days. Survival and whole-body lipid content were unaffected at 20°C (whole-body selenium
concentrations equal to 6 mg/kg dw, the sole treatment exposure) when compared to control fish.
Thus, at 20°C the chronic value for juvenile bluegill exposed to waterborne and dietary selenium
based on survival was >6 mg/kg dw in whole-body tissue. Fish exposed to the combination low-
level waterborne and dietary selenium at 4°C exhibited significantly elevated mortality (40.4
percent) relative to controls (2.9 percent), and exhibited significantly greater oxygen
consumption and reduced lipid content, which are indicative of stress. At 4°C the chronic value
for juvenile bluegill exposed to waterborne and dietary selenium was <7.91 mg Se/kg dw in
whole body based on mortality and tissue measurements at the end of the test (180 days), and
5.85 mg Se/kg dw in whole body based on mortality at 180 days and tissue measurements at 60
days. The increase in the concentration of whole-body selenium between Day 60 and 180 at 4EC
was apparently due to reductions in body weight caused by loss of lipid (comparatively low in
selenium) while body burden in other tissues remained relatively constant. If this concentration
of selenium in tissues occurs in sensitive overwintering fish in nature, a concentration of 5.85
mg/kg dw (the selenium tissue concentration in the 4°C exposure after 60 days) in fish collected
during the summer or fall months could be considered a threshold concentration for the
selenium-sensitive fish during the winter months. Therefore, this study's chronic value for the
threshold concentration prior to winter stress is 5.85 mg Se/kg dw in whole body tissue.
Mclntyre et al. (2008) also investigated the toxicity of selenium to juvenile bluegill under
cold temperature conditions in the laboratory. Whereas relative to the control, Lemly (1993a)
tested only one exposure level, 5 mg Se/kg in the diet and 5 ug Se/L and one low temperature
regime, 4°C, Mclntyre et al. (2008) evaluated a range of diet and water concentrations, two types
of diet, and two low-temperature regimes. The goal of the study was to determine ECio and EC20
values for selenium exposure to juvenile bluegill in a 4°C and 9°C low-temperature regimes.
Three separate exposure systems were run concurrently for 182 days. Two systems exposed
juvenile bluegill to a series of six aqueous and dietary selenium treatments and a control; one
exposure system (ESI) with a cold temperature regime (4°C), and one (ESS) with a cool
temperature regime (9°C), both using a yeast-worm-fish food chain bioaccumulation system.
That is, graded levels of selenized-yeast in ESI and ESS were fed to the oligochaete,
Lumbriculus variegatus, which in turn was fed to bluegill. The third exposure system (ES2) used
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diet and exposure conditions similar to Lemly's 4°C treatment, i.e., nominal 5 jig Se/L in the
water and nominal 5 mg Se/kg dw food (seleno-L-methionine in TetraMin). The cold
temperature regime for ESI and ES2 was 20°C for the first 30 days of exposure, and then
decreased 2°C/week until it reached 4°C (test day 79) at which point temperature was maintained
until test termination (test day 182). The cool temperature regime (ESS) was similar except when
the temperature reached 9°C (test day 65), it was maintained until test termination (test day 182).
At the end of the 182 day exposure in the ES2 (with Lemly's diet and temperature), the
bluegill accumulated an average (geometric mean) whole body concentration of 9.99 mg/kg dw
with no meaningful mortality in the treatment or control. Significant mortality of juvenile
bluegill was observed in the two highest treatments in the cold (ESI) and cool (ESS)
Lumbriculiis-fed tests. No effects on body weight or condition factor were observed. The ECio
and EC20 values for the cold treatment (ESI) are 9.27 and 9.78 mg Se/kg dw in whole body,
respectively. The ECio and £€20 values for the cool treatment (ESS) are slightly higher at 14.00
and 14.64 mg Se/kg dw in whole body, respectively.
The design and the results of the Mclntyre et al. (2008) study have similarities and
differences with Lemly (1993a), as presented in detail with comparisons and contrasts in
Appendix C. Both studies found juvenile bluegill were more sensitive in a cold-temperature
regime than in a cool (Mclntyre et al.) or a warm regime (Lemly). The effect levels determined
for the cold temperature regime differed by a factor of 1.58 (ESI of Mclntyre et al., 9.27 mg
Se/kg; Lemly, 5.85 mg Se/kg), a difference rather typical of chronic studies conducted in
different laboratories using different fish populations (Delos 2001) and similar to the 1.51 factor
difference between two ECios of Hamilton et al. (1990) for chinook salmon. As these two cold-
temperature juvenile-survival lab studies are far more similar than they are different, their results
were combined per the standard Guidelines procedure to determine the non-reproductive SMCVs
for bluegill. These SMCVs were determined separately for two temperature conditions for
bluegill, 4°C and 9°C. The SMCV for 4°C is 8.15 mg Se/kg dw whole body, based on three
chronic values: (a) the Lemly (1993a) concentration prior to winter stress (5.85 mg Se/kg dw
whole body), (b) the Mclntyre et al. (2008) ESI ECio (9.27 mg Se/kg dw whole body), and (c)
the Mclntyre et al. (2008) ES2 NOEC (>9.992 mg Se/kg dw whole body). This value is not less
than the reproductive endpoint-based whole-body criterion concentration of 8.13 mg Se/kg dw.
The SMCV for 9°C is 14.00 mg Se/kg dw whole body, based on the Mclntyre et al. (2008) ES3
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. The studies of Bryson et al (1985b) and Cleveland et al. (1993) were not conducted at cold
temperature and were thus not used for these SMCV calculations.
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Summary of Studies with Non-Reproductive Effects.
Table 17.
Exposed.)
Freshwater Chronic Values from Acceptable Tests - Non-Reproductive Endpoints (Parental Females Not
Species
Acipenser
trammontanus
white sturgeon
Pogonichthys
macrolepidotus
Sacramento splittail
Pimephales
promelas
fathead minnow
Reference
Tashjian et al.
2006
Teh et al.
2004
Bennett et al.
1986
Exposure
route and
duration
dietary (lab)
8 weeks
dietary (lab)
9 months
dietary (lab)
9 to 19 days
7
Selenium form
seleno-L-
methionine in
artificial diet
selenized-yeast
algae exposed to
selenite then fed
to rotifers which
were fed to fish
Toxicological
endpoint
ECio juvenile growth
/
EC2pxTuvenile growth
NOEC
LOEC
MATC juvenile
deformities (juvenile
exposure only)
Chronic value for
larval growth
Chronic value,
mg/kg dwa
15.08WB
27.76 M
17.82WB
32.53 M
10.1 M
15. 1M
12.34 M
51.40WB
SMCV
mg/kg dw
EC 10
15.1 WB
27.8 M
EC2o
17.8 WB
32.5 M
10.1 M
15. 1M
12.3 M
51.40WB
GMCV
mg/kg dw
15.1 WB
27.8 M
10.1 M
15. 1M
12.3 M
51.40WB
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Species
Pimephales
promelas
fathead minnow
Xyrauchen texanus
razorback sucker
Xyrauchen texanus
razorback sucker
Catostomus
latipinnis
flannelmouth sucker
Reference
Dobbs et al.
1996
Beyers and
Sodegren
200 la
Beyers and
Sodegren
2001b
Beyers and
Sodegren
200 la
Exposure
route and
duration
dietary and
waterborne
(lab)
8 days
dietary and
waterborne
(lab)
28 days
dietary and
waterborne
(lab)
28 days
dietary and
waterborn^r
(lab)
28 days
Selenium form
algae exposed to
selenate in water
then fed to
rotifers which
were fed to fish
water: selenate;
diet: algae
exposed to
selenate in water
then fed to
rotifers which
were fed to fish
water: site
waters;
diet: algae /
exposed to site
water then fed to
rotifers which
were fed to fish
water: selenate;
diet: algae
exposed to
selenate in water
then fed to
rotifers which
were fed to fish
Toxicological
endpoint
LOEC for larval fish
dry weight after 8 d
NOEC for survival
and growth
NOEC for survival
and growth
NOEC for survival
and growth
Chronic value,
mg/kg dwa
<73 WBb
>12.9WBb
>42 WBb
>10.2WB
SMCV
mg/kg dw
69.83 M
see text
>10.2WB
GMCV
mg/kg dw
69.83 M
>10.2WB
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Species
Oncorhynchus
tshawytscha
chinook salmon
Oncorhynchus
mykiss
rainbow trout
Oncorhynchus
mykiss
rainbow trout
Morone saxitilis
striped bass
Reference
Hamilton et
al. 1990
Hilton and
Hodson 1983;
Hicks et al.
1984
Hilton et al.
1980
Coughlan and
Velte 1989
Exposure
route and
duration
dietary (lab)
60 days
dietary (lab)
60 days
dietary (lab)
16 weeks
dietary (lab)
20 weeks
dietary (lab)
80 days
Selenium form
mosquitofish
spiked with
seleno-DL-
methionine
mosquitofish
spiked with SLD
diet
sodium selenite
in food
preparation
sodium selenite
in food
preparation
Se-laden shiners
from Belews
Lake, NC
Toxicological
endpoint
ECio for juvenile
growth
EC 20 for juvenile
growth „
ECio for juvenile
growth
EC 20 for juvenile
growth
juvenile growth
NOEC
LOEC
MATC
juv. survival &
growth
NOEC
LOEC
MATC
LOEC for survival
of yearling bass
Chronic value,
mg/kg dwa
7.355 WB
10.47 WB
/^
11.14WB
15.73 WB
21 Liver
7 1.7 Liver
3 8. 80 Liver
40 Liver
100 Liver
63. 25 Liver
<16.2MC
SMCV
mg/kg dw
ECio
9.052 WB
EC 20
12.83 WB
NOAEC
28.98 L
LOAEC
84.68 L
MATC
49.52 L
<16.2M
GMCV
mg/kg dw
ECio
9.052 WB
<16.2M
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Species
Lepomis
macrochirus
bluegill
Lepomis
macrochirus
bluegill
Reference
Lemly 1993 a
Mclntyre et al.
2008
Exposure
route and
duration
dietary and
waterborne
(lab)
180 days
20 to 4°C
dietary and
waterborne
(lab)
180 days
20°C
dietary and
waterborne
(lab)
182 days
20 to 4°C
(ESI)
dietary and
waterborne
(lab)
182 days
20 to 9°Cy
(ESS) /_
dietary and
waterborne
(lab)
182 days
20 to 4°C
(ES2)
Selenium form
diet: seleno-L-
methionine
water: 1:1
selenate:selenite
diet: seleno-L-
methionine
water: 1:1
selenate:selenite
diet:
Lumbriculus fed
selenized-yeast
water: 1:1
selenate:selenite
diet:
Lumbriculus fed
selenized-yeast
water: 1:1
selenate:selenite
diet: seleno-L-
methionine
water: 1:1
selenate:selenite
Toxicological
endpoint
LOEC for juvenile
mortality at 4°C
Threshold prior to
"winter stress"
NOEC for juvenile
mortality at 20°G//
/
/
ECiojuv. survival
ESI
EC2ojuv. survival
ESI
ECiojuv. survival
ESS
EC2ojuv. survival
ESS
NOECjuv. surv.
ES2
Chronic value,
mg/kg dwa
<7.91 WB
/
/
5.85 WB
>6.0 WB
9.27 WB
9.78 WB
14.00 WB
14.64 WB
>9.992 WB
SMCV
mg/kg dw
4°C
ECio-
NOAEC
8.15WB
4°C
EC20-
LOAEC
8.80WB
9°C ECio
14.0 WB
9°C EC20
14.6 WB
GMCV
mg/kg dw
8.15 WB
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Species
Lepomis
macrochirus
bluegill
Lepomis
macrochirus
bluegill
Reference
Bryson et al.
1985b
Cleveland et
al. 1993
Exposure
route and
duration
dietary (lab)
60 days
dietary (lab)
90 days
Selenium form
seleno-DL-
cysteine
seleno-L-
methionine
Toxicological
endpoint
NOEC for juvenile
growth
NOEC for juvenile
survival
Chronic value,
mg/kg dwa
>3.74WBb
>13.4WBb
SMCV
mg/kg dw
GMCV
mg/kg dw
a All chronic values reported in this table are based on the measured concentration of selenium in whole body (WB), muscle (M) or
liver (L) tissues.
Chronic value not used in SMCV calculation (see text).
Tissue value converted from ww to dw. See Appendix C for conversion
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7.1.9 Comparison of Fish Chronic Reproductive Effects and Chronic Non-Reproductive
Effects
A chonic criterion element of 15.2 mg/kg dw in the egg/ovary addresses the toxic effect
identified by the Chapman et al. (2009, 2010) expert workshop to be of greatest concern, and is
expected to be protective of non-reproductive endpoints such as juvenile survival and growth.
The egg/ovary chronic criterion element can be compared to the most sensitive non-reproductive
SMCV, cold-stressed (4°C) bluegill, using a conversion from selenium in egg/ovary to whole
body tissue. For bluegill the EO/WB ratio of 2.13 is based on n=27 observations compiled from
four different sources (Coyle et al. 1993; Doroshov et al. 1992; Hermanutz et al. 1996; and
Osmundson et al. 2007) and discussed in Section 4.1.5. These yielded a good correlation
between the concentrations of selenium in egg/ovary relative to whole body. Using an egg-ovary
to whole-body ratio of 2.13, a concentration of 15.2 mg/kg dw in the egg-ovary of bluegills
converts to a whole-body selenium concentration of 7.14 mg/kg dw, which is below the 4°C non-
reproductive SMCV for bluegill of 8.15 mg/kg dw. Equivalently, the same whole-body to ovary-
egg conversion factor of 2.13 can be used to convert the non-reproductive bluegill SMCV of
8.15 mg/kg Se whole-body to an ovary/egg concentration of 17.36 mg Se/kg dw, which would be
protected by the reproductive egg/ovary criterion of 15.2 mg/kg.
The bluegill cold-stressed non-reproductive SMCV is also protected by the reproductive
effect-based whole-body criterion of 8.13 mg Se/kg. Figure 15 shows the non-reproductive effect
whole-body GMCVs compared to the whole-body criterion.
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Ol
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12-80 mg Se/kg dw WB. If juvenile mortality occurred at concentrations lower than those found
to induce larval deformities and at concentrations as low as Lemly (1993a) reported in the lab
(EC/io = 7.91 mg Se/kg WB), then centrarchids would likely not have been present in Belews
Lake. The observations of Lemly (1993b) are evidence that larval deformity, not juvenile
mortality, is the more sensitive endpoint.
The Crutchfield and Person (2000) predictions and field observations of recovery of
bluegill at Hyco Reservoir likewise suggest that significant mortality was unlikely to be
occurring at the concentrations Lemly (1993a) reported to cause substantial mortality. During a
time period over which Crutchfield (2000) indicated dietary invertebrate concentrations
exceeded 20 mg Se/kg dw, Crutchfield and Person (2000) indicated that bluegill population
growth occurred at rates predicted to be natural for the unimpaired species. In contrast, if the
Lemly (1993a) lab EC/io of 7.91 mg Se/kg dw whole-body were applicable to this field situation,
the mortality associated with the resulting bluegill whole-body concentrations (25 mg Se/kg dw
whole-body, assuming a trophic transfer factor of 1.27) would have prevented any recovery.
Selenium-induced cold temperature loss of lipid and body condition, a non-reproductive
sublethal effect that Lemly (1993a) observed to accompany juvenile mortality in the laboratory
(but which Mclntyre et al. (2008) did not observe in a similar study) has also not generally been
corroborated by field evidence (Janz 2008). Several studies have measured growth and energy
storage indicators in juvenile fish just prior to and just after winter at reference sites and sites
with elevated selenium in northern Canada (Bennett and Janz 2007a, b; Kelly and Janz 2008;
Driedger et al 2009; Weber et al. 2008). The growth (length, weight, condition factor, muscle
RNA:DNA ratio, muscle protein) and energy storage (whole body lipids, whole body
triglycerides, liver triglycerides, liver glycogen) indicators for five fish species (northern pike,
burbot, fathead minnow, creek chub, white sucker) measured just after winter were similar or
greater than those measured just before winter at the selenium exposed sites. The slimy sculpin
did show a decrease in whole body triglycerides but the reduction was similar at exposed and
reference sites.
In contrast to the non-reproductive effects, the reproductive effects show more clear-cut
concentration-response relationships (11 of the 19 reproductive chronic values are specific ECs,
whereas only 5 of the 19 non-reproductive chronic values), are more readily reproducible, and
are better corroborated by field observations. Reproductive effects represent the endpoint of
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greatest concern (Chapman et al. 2009, 2010); all non-reproductive GMCVs are protected by a
criterion derived from the reproductive GMCVs. The reproductive endpoint data, expressed
relative to selenium concentrations in fish eggs and ovaries, thus provide a more reliable and
protective basis for the criterion. Because the data set used to derive the criterion is comprised
primarily of the aquatic species considered most sensitive to selenium (salmonids and
centrarchids) and because the criteria are designed to protect 95% of the genera, the criterion of
15.2 mg/kg dw ovary/egg should be protective of aquatic populations offish and invertebrates.
7.2 Water
7.2.1 Validation of Translation Equation for Developing Water Column Concentrations
The EPA evaluated the efficacy of the equation used to translate the egg-ovary criterion
element to a water column concentration. The EPA's translation equation is given as:
C,
C
water
"egg-ovary
'TTFcomposltexEFxCF (Equation 18)
Because fish bioaccumulate selenium over a relatively long time period, single measurements of
selenium in fish tissue are likely to be less variable and a better representation of selenium loads
to the aquatic system than single measurements of selenium in the water column. Thus the EPA
used a validation approach based on fish tissue measurements rather than single water
measurements.
The EPA solved Equation 18 for egg-ovary concentration yielding:
Cegg.ovxy = Cwater x TTFcompos'tex EF x CF (Equation 19)
The EPA then used Equation 19 to calculate the predicted concentration of selenium in the eggs
and ovaries offish from all spatially and temporally relevant measurements in the water column.
The EPA then compared those predicted values to the measured concentration in the fish.
The EPA searched its selenium database for measurements of selenium in fish tissue
taken from aquatic sites with a previously calculated EF value. Identified tissue measurements
from other than eggs or ovaries were converted to equivalent egg-ovary concentrations using
species-specific conversion factors as described previously. For each tissue measurement, the
EPA searched its selenium database again for water column measurements that were taken from
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the same aquatic site and within one year of the tissue measurement. If more than one water
column measurement was matched to a tissue measurement, the median water column
measurement was used. For each matched pair of tissue and water measurements, the EPA
identified appropriate species-specific TTF and CF values as described previously, and the EF
value from the site the samples were taken. The EPA then used Equation 19 to calculate the
predicted egg-ovary concentration from the observed water column concentration. Finally, the
EPA compared the predicted egg-ovary concentrations with the observed egg-ovary
concentrations.
The EPA identified 300 tissue measurements associated with one or more water column
measurements. A predicted egg-ovary concentration was calculated for each water column
concentration as described above. Figure 16 shows all 300 predicted egg-ovary concentrations
plotted against the measured egg-ovary concentrations. Because both the predicted and observed
selenium concentrations exhibited substantial heteroscedasticity (the variability of one variable is
unequal across the range of values of a second variable that predicts it), they are plotted and
analyzed on a log scale. The predicted and measured concentrations is highly correlated
(r = 0.84, t(298) = 26.28, P < 10'80 ).
1000.0
Observed egg-ovary
concentration (mg/kg
dw)
100.0 •
10.0 •
1.0 -
0.1
0.1 1.0 10.0 100.0 1000.0
Predicted egg-ovary concentration (mg/kg dw)
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Figure 16. Scatter plot of predicted versus measured concentrations of selenium in fish.
Dashed line shows unity y = x line.
Although there is a strong correlation between predicted and observed water
concentration values, there is some scatter around the hypothesized y = x unity line. Dispersion
around the unity line is likely attributable to several sources of uncertainty. Potential sources of
uncertainty include small sample sizes, temporal or spatial variability in selenium exposure, and
local variability in aquatic food webs. The EPA limited its analysis to only those aquatic sites
with at least two particulate measurements available to calculate its EF value and with at least
one of them from algae or detritus. Nevertheless, only one or two measurements of algae and/or
detritus were available for 41 of the 69 aquatic sites evaluated. Although such a restriction
reduces uncertainty when applying Equation 19 to available data, the EPA believes that two
particulate measurements are only marginally sufficient. Another potential source of uncertainty
is the frequent absence of site-specific information about the types and proportions of organisms
ingested by fish. In most cases, the EPA estimated the type and proportion of prey organisms
using general knowledge of the fish species and aquatic system location. Notwithstanding the
limitations in available data, the EPA concludes from this analysis that Equation 18 provides a
reasonable translation of the egg-ovary criterion to a site-specific water concentration.
7.2.2 Evaluating the Protectiveness of the Final Water-Column Criterion Values
To evaluate the protectiveness of the water column criterion values, the EPA used field
data to compare attainment or exceedance of the egg-ovary criterion element to attainment or
exceedance of the water column criterion element values at aquatic sites with measurements of
selenium in both tissue and water. The EPA identified fish tissue measurements in its database of
available selenium measurements, and then searched for matched measurements in water defined
as those measurements collected at the same aquatic site within one year of the tissue
measurement. Because water measurements represent the concentration of selenium at a specific
location and point in time whereas tissue measurements represent the accumulated exposure to
selenium over a larger geographic area and time period, the EPA assessed tissue attainment or
exceedance by comparing the egg-ovary criterion element to each single tissue sample, and
assessed water column attainment or exceedance by comparing the water column criterion
element value to a water column concentration that was estimated from at least 3 matched water
measurements. Measurements of selenium in tissue other than eggs or ovaries were converted to
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an equivalent egg-ovary concentration using the same method that was used for validation of
Equation 18 described above.
The final selenium water criterion values are expressed as a 30-day average not to exceed
more than once in 3 years on average. To assess attainment or exceedance using water
concentration measurements that were randomly sampled at a site over two years (between one
year before and one year after the site-matched tissue sample was collected), the EPA adjusted
the water concentrations matched to each tissue measurement by assuming a lognormal
distribution and calculating the 95* percentile. For tissue measurements with 6 or more matched
water-column measurements, the EPA calculated the 95th percentile using the standard deviation
of the measurements matched with the tissue measurement. For tissue measurements with fewer
than 6 matching water measurements, the EPA calculated the water column standard deviation
for each matched tissue measurement, calculated the average standard deviation, and then used
the average standard deviation to calculate the 95* percentile water column concentration for
each tissue measurement. Because the average water column standard deviation for lentic aquatic
systems was significantly smaller than for lotic aquatic systems (t(in) = 2.32, P < 0.05), the EPA
applied the average standard deviation from lentic aquatic systems to water measurements from
lentic aquatic systems, and applied the average standard deviation from lotic aquatic systems to
water measurements taken from lotic aquatic systems.
The EPA used these data to assess attainment or exceedance of 140 instances in lentic
aquatic systems and 688 instances in lotic aquatic systems. Although such a binary classification
scheme does not consider the degree to which measurements are above or below a criterion,
water quality standards are usually implemented as a binary decision (a water body either attains
or exceeds criteria) and thus is a useful tool to evaluate the performance of the water column
criterion element concentrations. Table 18 and Table 19 summarize the results of this binary
classification.
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Table 18. Comparison of criterion attainment using the water column and egg-ovary
concentration values in lentic aquatic systems.
Water concentration greater
than water criterion element
Water concentration less than
water criterion element
Tissue concentration
greater than tissue
criterion element
96 (69%)
10 (7%)
Tissue concentration less
than tissue criterion
element
16(11%)
13 (15%)
Table 19. Comparison of selenium criterion attainment using the water column and
egg-ovary concentration values in lotic aquatic systems.
Water concentration greater
than water criterion element
Water concentration less than
water criterion element
Tissue concentration
greater than tissue
criterion element
248 (36%)
52 (8%)
Tissue concentration less
than tissue criterion
element
206 (30%)
182(26%)
The EPA used these binary classifications tables to calculate the binary classification
statistics specificity, sensitivity, positive prediction value, negative prediction value, and
accuracy. Sensitivity is the probability that the water column concentration will exceed the water
column criterion element when the egg-ovary concentration is exceeding the egg-ovary citerion
element. Specificity is the probability that the water column concentration will attain (be equal to
or less than) the water column criterion element when the egg-ovary concentration is attaining
(equal to or less than ) the egg-ovary criterion element. Positive prediction value is the
probability that the egg-ovary concentration will exceed the egg-ovary criterion element when
the water column concentration is exceeding the water column criterion element. Negative
prediction value is the probability that the egg-ovary concentration will attain the egg-ovary
criterion element when the water concentration is attaining the water criterion element. Accuracy
is the probability that any assessment decision will be correctly categorized. Finally,
environmental protectiveness indictates the percent of time/measurements that meeting the 30-
day water column criterion element would be expected to protect against any fish egg-ovary
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criterion element exceeedances, which are 93 and 92% for lentic and lotic systems, respectively.
The environmental protectiveness value indicates that false negative conclusions regarding fish
tissue exceedances would be minimized, and would occur less than 10% of the time, if the
selected 20th percentile water column value for the water column criterion element is not
exceeded. The binary classification statistics are shown in Table 20.
Table 20. Binary classification statistics for lentic and lotic aquatic systems.
Sensitivity
Specificity
Positive prediction value
Negative prediction value
Accuracy
Environmental protectiveness
Lentic
0.91
0.53
0.86
0.64
0.81
0.93
Lotic
0.83
0.47
0.55
0.78
0.62
0.92
These binary classification statistics indicate that the chronic water criterion element
values are highly sensitive to exceedance of the egg-ovary criterion element (that is, when the
egg-ovary criterion element is exceeded, it is highly likely that the water column criterion
element will also be exceeded). Specificity, positive prediction value, negative prediction value,
and accuracy are all within a reasonable range of values (see appendix H for further explanation
and details on the binary classification statistics). The EPA concludes from these analyses that
the lentic and lotic water column criterion element values are adequately protective of aquatic
life.
^
7.2.3 Uncertainty in Bioaccumulation of Total Dissolved Selenium
Geochemicalform of selenium. The form of selenium in water (selenate, selenite, or
organoselenium) determines how readily selenium enters aquatic food webs and cycles through
particulate matter, consumer organisms, and predators. Inorganic selenium forms (e.g., selenate
and selenite) have relatively limited bioavailability compared with organo-selenium forms.
Typically, inorganic selenium released to aquatic systems is reduced and biotransformed to
organo-selenium compounds (Bowie et al. 1996). Organoselenium and selenite are more
bioavailable than selenate and thus may bioaccumulate to a greater extent (Besser et al. 1993;
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Rosetta and Knight 1995). Thus variability and uncertainty in the form of selenium released to a
waterbody contributes to uncertainty in anticipated effects for a given water body.
7.3 Protection of Threatened or Endangered Species
The chronic toxicity dataset for selenium contains toxicity data for two Federally-listed
endangered species, Cyprinodon macularius (desert pupfish) and Oncorhynchus mykiss (listed as
steelhead, indicating anadromous individuals, but herein called rainbow trout, implying non-
anadromous individuals).
Desert pupfish, Cyprinodon macularius,whh a chronic value estimated to be >27 mg
Se/kg dw egg, is not among the most sensitive species. Its chronic value of >27 mg Se/kg dw
egg, is substantially above the chronic egg-ovary criterion element of 15.2 mg Se/kg dw.
Oncorhynchus mykiss has a SMCV of 21.1 mg Se/kg dw egg, and is in the fourth most
sensitive genus. The dataset contains multiple studies with cutthroat trout (Oncorhynchus clarkf)
some subspecies of which are Federally listed as threatened. The SMCV for cutthroat trout is
24.06 mg Se/kg dw egg. Both of these chronic values for Oncorhynchus species are greater than
the chronic egg-ovary criterion element.
The dataset also contains toxicity information for Salvelinus malma (Dolly Varden)
which is not threatened or endangered, but is so closely related to the threatened Salvelinus
conjluentus (bull trout) that it can hybridize with that species, producing fertile offspring (Baxter
et al. 1997). Dolly Varden is the least sensitive fish species for which information is available,
with SMCV of 56 mg Se/kg dw egg. Salvelinus fontinalis, brook trout, can also hybridize with
bull trout, but the offspring are sterile, suggesting that it is less closely related. With the available
study of brook trout, although Section 7.1.3 conservatively sets the NOEC at >20.5 mg Se/kg dw
egg, which was the average concentration at the Holm et al. (2005) high-exposure site, the
concentration-response information for the offspring of individual females, presented in
Appendix C, suggests that its ECio could be substantially higher, possibly as high as that for
Dolly Varden.
The criterion of 15.2 mg Se/kg (dw) egg-ovary element is below all of the above
mentioned chronic values for threatened and endangered (or closely related) species. However,
because other threatened or endangered species might be more sensitive, if relevant new
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information becomes available in the future, it should be considered in state- or site-specific
criteria calculations.
7.4 Aquatic-Dependent Wildlife is Beyond the Scope of this Aquatic Criteria Derivation.
AWQC that are developed by the EPA typically focus directly on aquatic life, not
aquatic-dependent wildlife such as birds. As presented by Campbell (2011), EPA recognizes that
selenium effects on aquatic-dependent wildlife are also of concern but considers them beyond
the scope of this national criterion update. In the interest of providing updated guidance to
protect against the known risks of selenium exposure to fish, EPA decided to focus its analyses
on updating the existing selenium criterion for freshwater aquatic life based on the latest science
evidence. EPA plans, in the future, to consider the effects of selenium on aquatic-dependent
wildlife, potentially in the form of criteria expanded to address aquatic-dependent wildlife. When
translated to a water concentration, a criterion protective of aquatic-dependent wildlife may be
more stringent or possibly less stringent, than the values provided for aquatic life in this 2014
criteria document. This is likely because data indicate that selenium does not significantly
biomagnify moving up the food chain except in specific ecosystems with mollusk-based food-
webs, unlike bioaccumulative chemicals such as mercury. The single largest step in tissue
selenium accumulation in aquatic environments occurs at the base of the food web where algae
and other microorganisms accumulate selenium from water (Orr et al. 2012; Stewart et al. 2010).
Mollusks such as mussels and clams accumulate selenium to a much greater extent than
planktonic crustaceans and insects due to higher ingestion rates of both particulate-bound (algae)
and dissolved selenium from the water column through filter feeding, as well as the lower rate at
which they eliminate selenium (Luoma and Rainbow 2005). Thus, aquatic-dependent wildlife
criteria for species that feed primarily on mollusks would be expected to have lower values than
the 2014 selenium criterion found in this document. The criteria values for aquatic-dependent
wildlife would be expected to depend on the aquatic systems, species, and food webs considered.
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vxEPA
United States Office of Water EPA 822-P-14-001
Environmental Protection 4304T May 2014
Agency
External Peer Review Draft
Aquatic Life Ambient Water Quality
Criterion for
Selenium - Freshwater
2014
(Appendices A-K)
U.S. Environmental Protection Agency
Office of Water
Office of Science and Technology
Washington, B.C.
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LIST OF APENDICIES
APPENDIX A: Selenium Chemistry 1
Inorganic Selenium 2
Organoselenium 4
Departure from Thermodynamic Equilibrium 5
Physical Distribution of Species in Surface Water 5
APPENDIX B: Conversions 1
Conversion of wet to dry tissue weight 2
Methodology 2
Derivation of egg-ovary to whole-body conversion factors 3
CF values calculated directly from whole-body and egg-ovary selenium measurements.. 5
Muscle to egg-ovary conversion factors 17
Muscle to whole-body correction factor 34
Derivation of Trophic Transfer Function values 45
Methodology 45
TTF values from physiological coefficients 47
TTF values from field data 53
APPENDIX C: Summaries of Chronic Studies Considered For Criteria Derivation 1
APPENDIX D: Other Data 1
Selenite 2
Selenate 2
Other Data -Endangered Species 13
Other Data- Chronic Studies with Fish Species 18
Other Data- Chronic Studies with Invertebrate Species 32
Other Data- Field Study West Virginia Impoundments 35
APPENDIX E:Toxicity of Selenium to Aquatic Plants 1
Selenite 2
Selenate 2
APPENDIX F: Unused Data 1
APPENDIX G: Supplementary information on Selenium Bioaccumulation in Aquatic Animals 1
1.0 Effects of Growth Rate on the Accumulation of Selenium in Fish 2
2.0 Analysis of the Relative Contribution of Aqueous and Dietary Uptake on the Bioaccumulation
of Selenium 3
3.0 Kinetics of Accumulation and Depuration: Averaging Period 4
3.1 Background 4
3.2 Approach for Modeling Effects of Time-Variable Se Concentrations 5
3.2.1 Model Results 9
3.2.1.1 Steady concentrations at the water criterion concentration 9
3.2.1.2 Normally distributed 1-day spikes having CV=0.12, uniformly separated by
29 days at zero concentration 11
3.2.1.3 Log-normally distributed 1-day spikes, with log standard deviation = 0.20,
uniformly separated by 29 days at zero concentration 12
3.2.1.4 Log-normally distributed, smoothly variable concentrations, continuous
rather than intermittent exposure 13
3.2.2 Summary of Scenario Results 14
3.2.3 Conclusion 14
APPENDIX H: Binary Classification Statistics 1
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Sensitivity, Specificity, and Related Statistics in Applying Water-Column Criteria 2
APPENDIX I: Site Specific Criteria 1
1.0 Translating the concentration of selenium in tissue to a concentration in water 2
1.1 Relating the Concentration of Selenium in Tissue and Water using the mechanistic modeling
approach 4
1.2 Steps for deriving a site-specific water concentration value from the egg-ovary FCV 9
1.2.1 Identify the appropriate target fish species 9
1.2.1.1 When fish are present 9
1.2.1.2 When fish are absent 12
1.2.2 Model the food-web of the targeted fish species 13
1.2.3 Identify appropriate TTF values 14
1.2.3.1 Select the appropriate TTF values from a list of EPA-derived values 14
1.2.3.2 Deriving TTF values from existing data 16
1.2.3.3 Deriving TTF values by conducting additional studies 17
1.2.3.4 Extrapolating TTF values from existing values 18
1.2.4 Determine the appropriate FF value 18
1.2.4.1 Deriving a site-specific EF value from field measurements 18
1.2.4.2 Deriving an appropriate FF value from existing data 19
1.2.4.3 Extrapolating from FF values of similar waters 19
1.2.5 Determine the appropriate CF value 20
1.2.5.1 Selecting the appropriate CF value from the list of values that were used to
derive EPA's recommended water criteria concentration values 20
1.2.5.2 Deriving a CF value from existing data 21
1.2.5.3 Deriving a CF value by conducting additional studies 21
1.2.5.4 Extrapolating the CF value from the list of values that were used to derive
EPA's recommended water criteria concentration values 22
1.2.6 Translate the selenium egg-ovary FCV into a site-specific water concentration
value using Equation 18 22
1.3 Managing uncertainty using the mechanistic modeling approach 22
1.4 Example calculations 23
1.4.1 Example 1 23
1.4.2 Example 2 24
1.4.3 Example3 25
1.4.4 Example 4 26
1.5.5 Example 5 27
1.5.6 Example 6 28
1.5.7 Example 7a 29
Derivation of a site specific water column criterion for a river impacted by selenium.
29
1.5.7 Example 7b 29
Derivation of a site specific water column criterion for a lake impacted by selenium. 29
2.0 Translating the concentration of selenium in tissue to a concentration in water using
Bioaccumulation Factors (BAF) 30
2.1 Summary of the BAF approach 30
2.2 Managing uncertainty using the BAF approach 31
3.0 Comparison of Mechanistic Bioaccumulation Modeling and BAF approaches 33
APPENDIX J: Analytical Methods for measuring Selenium 1
General considerations when measuring concentrations of selenium 2
Analytical methods recommended for measuring selenium in water 3
American Public Health Standard Method 3114 B 3
EPA Method 200.8 4
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EPA Method 200.9 4
Analytical methods available for measuring selenium in fish tissue 5
Strong acid digestion 6
Dry-ashing digestion 6
APPENDIX K: Abbreviations 1
Reference and site abbreviations 2
Reference and site abbreviations 6
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APPENDIX A: Selenium Chemistry
A-l Draft Document
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Selenium in aquatic ecosystems exists in a broad range of oxidation states: (+ VI) in selenates (HSeO4~,
SeO42") and selenic acid (H2SeO4), (+ IV) in selenites (HSeO3~, SeO32") and selenous acid (H2SeO3), 0 in
elemental selenium, and (-II) in selenides (Se2~, HSe"), hydrogen selenide (H2Se), and organic selenides
(R2Se). Selenium also shows some tendency to form catenated species like organic diselenides (RseSeR).
Within the normal physiological pH range and the reduction potential range permitted by water, only Se,
SeO32", HSeO3", and SeO42" can exist at thermodynamic equilibrium (Milne 1998). While ionic reactions
are expected to be rapid in water, oxidation-reduction reactions may be slow, and the possibility exists for
the formation of HSe" in living systems and some environments where anoxic conditions arise. The
parallel behavior of comparable species of sulfur and selenium in living systems has often been observed,
but it is important to recognize that their chemical characteristics are different in many ways. For
instance, selenate is comparable to chromate in oxidizing strength and far stronger than sulfate [£°(SeO42~
/H2SeO3) = 1.15 V; £°(Cr2O727Cr3+) = 1.33V; £°(SO42'/H2SO3) = 0.200V (standard potentials in acid
solution: Weast 1969)], whereas selenide is a much stronger reducing agent than sulfide [£°(Se/H2Se) = -
0.36 V;£°[S/H2S]= 0.14V)].
Inorganic Selenium
Selenate usually predominates in well-aerated surface waters, especially those with alkaline conditions. In
spite of its oxidizing strength, selenate (SeO42~) exhibits considerable kinetic stability in the presence of
reducing agents (Cotton and Wilkinson 1988). The radius of SeO42" is comparable to that of SO42"(Frausto
da Silva and Williams 1991), and uptake by cells is expected to take place via the same ion channels or
permeases for both anions. Competition between sulfate and selenate uptake has been observed in many
species: algae (Riedel and Sanders 1996), aquatic plants (Bailey et al., 1995), crustaceans (Ogle and
Knight 1996), fungi (Gharieb et al. 1995), HeLa cells (Yan and Frenkel 1994), and wheat (Richter and
Bergmann 1993). Reduced selenate bioconcentration with increasing sulfate concentration has been
demonstrated in Daphnia magna (Hansen et al. 1993). A significant inverse relationship was shown to
exist between acute selenate toxicity to aquatic organisms and ambient sulfate concentrations (Brix et al.
200la). Competition with selenate has also been observed for phosphate in green algae (Riedel and
Sanders 1996), and with chromate and tungstate in anaerobic bacteria (Oremland et al. 1989).
Selenous acid species (HSeO3~ and SeO32") can predominate in solution under the moderately oxidizing
conditions encountered in oxygenated waters. Between pH 3.5 and 9.0 biselenite ion is the predominant
ion in water, and at pH values below 7.0, selenites are rapidly reduced to elemental selenium under mildly
reducing conditions (Faust 1981), situations that are common in bottom sediments.
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Most selenite salts are less soluble than the corresponding selenates. The extremely low solubility of
ferric selenite Fe2(SeO3)3 (Ks= 2.0 ± 1.7 x 10'31), and of the basic ferric selenite Fe2(OH)4SeO3 (Ks = 10"
61 7), is important to the environmental cycling of selenium. Selenites also form stable adsorption
complexes with ferric oxides, forming complexes of even lower solubility than the ferric selenites. Under
certain conditions, selenite (in contrast to selenate) seems to be completely adsorbed in high amounts by
ferric hydroxide and, to a lesser extent, by aluminum hydroxide (Faust 1981). Coprecipitation techniques
have been applied for preconcentration of selenium in natural waters, using iron (III) hydroxides, which
coprecipitates selectively the selenite, but not the selenate, species in river and sea waters (Yoshii et al.
1977). Alum and iron coagulation precipitation can be used in water treatment processes to remove
selenite (Clifford et al. 1986). The low levels of selenium in ocean waters have been attributed to the
adsorption of selenite by the oxides of metals, such as iron and manganese (National Academy of
Sciences 1976).
Relative to selenate, selenite is more reactive because of its polar character, resulting from the asymmetric
electron density of the ion, its basicity (attraction to bond with proton), and its nucleophilicity (attraction
to bond to a nucleus using the lone pair electrons of the ion). No evidence has yet been presented to show
that HSeO3" or SeO32" is taken up intact into the cell interior. Evidence indicates that selenite is reduced
rapidly, even before uptake in some cases, making it difficult to distinguish between uptake and metabolic
processes (Milne 1998). Freshwater phytoplankton process selenate and selenite by different mechanisms,
leading to different concentrations within the cell, and the concentrations attained are affected by various
chemical and biological factors in the environment (Riedel et al. 1991). These authors suggested that
selenate is transported into the cell by a biological process with low affinity, whereas selenite appears to
be largely physically adsorbed. Contradictory evidence suggesting that selenite uptake is enzymatically
mediated was found with marine phytoplankton (Baines and Fisher 2001). Experimental results
supporting the hypothesis that separate accumulation mechanisms for selenate and selenite are present in
D. magna have been published (Maier et al. 1993). However, while some organisms appear to absorb
selenite nonspecifically, specific transport systems exist in other species. Sulfate competition is
insignificant in the aquatic plant Ruppia maritima (Bailey et al. 1995), and specific uptake systems have
been demonstrated in some soft line microorganisms (Heider and Boeck 1993). Selenite uptake in green
algae, unlike selenate, is increased substantially at lower pH values, a property that represents another
difference between these two anions (Riedel and Sanders 1996). The uptake of inorganic selenium
species, selenate and selenite, by the green alga Chlamydomonas reinhardtii (Dang) was examined as a
function of pH over the range 5 to 9, and in media with varying concentrations of major ions and nutrients
using 75Se as a radiotracer. Little difference was noted in the uptake of selenate as a function of pH, with
A-3 Draft Document
-------
the maximum uptake found at pH 8; however, selenite uptake increased substantially at the lower pH
values. Differences in speciation are suggested to be the cause of these differences. Selenate exists as the
divalent ion SeO42" over the range of pH tested; whereas monovalent biselenite ion HSeO3" is prevelant at
these pH values. At the low end of the pH range, neutral selenous acid may also play a role.
Elemental selenium is not measurably soluble in water. It has been reported that elemental selenium is
slowly metabolized by several bacteria (Bacon and Ingledew 1989), and the translocation of elemental
selenium into the soft tissue of the marine mollusk Macoma balthica has been reported (Luoma et al.
1992). The bioavailability of elemental selenium to M. balthica was assessed by feeding the organisms
75Se-labeled sediments in which the elemental selenium was precipitated by microbial dissimilatory
reduction. A 22% absorption efficiency of particulate elemental selenium was observed. In view of the
insolubility of elemental selenium, uptake may be preceded by air oxidation, or in reducing environments
thiols may facilitate the solubilization (Amaratunga and Milne 1994). Elemental selenium can be the
dominant fraction in sediments (Zawislanski and McGrath 1998).
Selenium is reduced to hydrogen selenide, H2Se, or other selenides at relatively low redox potentials.
Hydrogen selenide by itself is not expected to exist in the aquatic environment since the Se°/H2Se couple
falls even below the FrYH2 couple. Aqueous solutions of H2Se are actually unstable in air due to its
decomposition into elemental selenium and water. Under moderately reducing conditions, heavy metals
are precipitated as the selenides, which have extremely low solubilities. The following are log Ks values
of some heavy metal selenides of environmental interest: -11.5 (Mn2+), -26.0 (Fe2+), -60.8 (Cu+), -48.1
(Cu2+), -29.4 (Zn2+), -35.2 (Cd2+), and -64.5 (Hg2+). The precipitation of selenium as heavy metal
selenides can be an important factor affecting the cycling of the element in soils and natural waters.
Organoselenium
Organic selenides (conventionally treated as Se(-II) species) in variable concentrations, usually in the
form of free and combined selenomethionine and selenocysteine, are also present in natural surface
waters (Fisher and Reinfelder 1991). Dissolved organic selenides may be an important source of selenium
for phytoplankton cells, because they can account for -80% of the dissolved selenium in open ocean
surface waters, and for a significant fraction in many other environments as well (Cutter 1989; Cutter and
Cutter 1995). Dissolved organoselenium levels of 14.2%, 65% and 66% were measured in samples (one
meter depth) from Hyco Reservoir, NC; Robinson Impoundment, SC; and Catfish Lake, NC; respectively
(Cutter 1986). The Hyco Reservoir organoselenium was identified as being protein bound.
A-4 Draft Document
-------
Organoselenium concentrations were found to range from 10.4% (58.7 ug/L) to 53.7% (1.02 ug/L) of the
total selenium present in Lake Creek and Benton Lake, MT surface waters (Zhang and Moore 1996).
Organoselenium quite often is measured as the difference between total dissolved selenium and the sum
of selenite plus selenate, and is therefore not typically characterized. Much more work is needed in the
area of specific identification and characterization of the nature of the organic selenides present in aquatic
ecosystems. Organoselenium form(s) are much more bioavailable and probably play a very important role
in selenium ecotoxic effects (e.g. Besser et al., 1993; Rosetta and Knight 1995).
Departure from Thermodynamic Equilibrium
In the highly dynamic natural waters, there is often a departure from thermodynamic equilibrium. In the
thermodymanic models, kinetic barriers to equilibrium and biological processes are not adequately
considered, and the speciation of selenium in oxidized natural waters is not accurately predicted. Selenate
is usually the predominate form in solution; however, selenite and Organoselenium can both exist at
concentrations higher than predicted (Faust 1981; Luoma et al. 1997). Bioaccumulation by
microorganisms, bioproduction and release of Organoselenium, and mineralization of particulate selenium
forms contribute to the disequilibrium.
Physical Distribution of Species in Surface Water
The physical distribution of various selenium species in surface waters is regulated by:
• sorption to or incorporation in suspended particulate matter (SPM), and
• complexation with inorganic and/or organic colloidal material, such as (FeO OH)n and humic
substances (dissolved organic matter, DOM).
Both sorption to SPM and complexation with colloidal matter reduces the bioavailability of the selenium
species. The average fraction of selenium associated with the particulate phase (0.45um filtration) as
determined from eleven different studies of various surface waters was found to be 16% (0-39% range) of
the total selenium, i.e., an average operationally defined dissolved selenium level of 84% (Table A-l). In
the James River, VA, the dissolved inorganic and organic selenium was found to be 77% and 70%
associated with colloidal matter, respectively (Takayangi and Wong 1984). A study of lake ecosystems in
Finland (Wang et al. 1995) found that 52% of the dissolved selenium was associated with humic
substances, and in a similar speciation study of Finnish stream waters, Lahermo et al. (1998) determined
that 36% of the selenium was complexed with humic matter. Hence, in various waterbodies physical
A-5 Draft Document
-------
distribution as well as chemical speciation of selenium must be considered in relationship to
bioavailability and aquatic toxicity.
Until recently, the organic selenium fraction has been routinely measured as the difference between total
dissolved selenium and the sum of selenite and selenate. Unfortunately, the calculation of this important
selenium fraction in water as the difference between the total and measurable inorganic fractions has not
permitted this fraction to be fully characterized. New techniques are currently being developed which
should help the specific identification and characterization of the nature of the organic selenides present
in aquatic systems. This work is particularly important because portions of the organic selenium fraction
(e.g., selenomethionine) of total dissolved selenium in water have been shown to be much more
bioavailable than the other forms of selenium, and therefore this work is also important for understanding
the manifestation of selenium ecotoxic effects.
Table A-l. Particulate and dissolved selenium as a function of total selenium in freshwater and
marine aquatic ecoystems
Reference
Cutter 1989
Cutter 1986
Tanizaki et al. 1992
Luomaetal. 1992
Cumbie and VanHorn, 1978
GLEC 1997
Wangetal. 1995
Lahermoetal. 1998
Hamilton et al. 200 la,b
Hamilton et al. 200 la,b
Hamilton et al. 200 la,b
Nakamoto and Hassler 1992
Nakamoto and Hassler 1992
Welsh and Maughan 1994
Welsh and Maughan 1994
Welsh and Maughan 1994
Welsh and Maughan 1994
Welsh and Maughan 1994
Welsh and Maughan 1994
Waterbody
Carquinezitist, CA
Hyco Reservoir, NC
Japanese Rivers
San Francisco Bay, CA
Belews Lake, NC
Unnamed Stream, Albright, WV
Finnish Lakes
Finnish Streams
Adobe Creek, Fruita, CO
North Pond, Fruita, CO
Fish Ponds, Fruita, CO
Merced River, CA
Salt Slough, CA
Cibola Lake, CA
Hart Mine Marsh, Blythe, CA
Colorado River, Blythe, CA
Palo Verda Oxbow Lake, CA
Palo Verda Oufall Drain, CA
Pretty Water Lake, CA
Particulate Se
(% of Total)
20-40
0
16
22-31
8
4
10
8
18
0
7
0
4
39
6
11
33
0
21
Fraction
dissolved, fd
0.6-0.8
1
0.84
0.69-0.78
0.92
0.96
0.9
0.92
0.82
1
0.93
1
0.96
0.62
0.94
0.89
0.67
1
0.79
A-6 Draft Document
-------
APPENDIX B: Conversions
B-l Draft Document
-------
Conversion of wet to dry tissue weight
Methodology
Conversion factors (CF) derived from selenium measurements were calculated using concentrations
expressed as dry weights ((ig/g dry weight). The majority of tissue and whole-body selenium
concentrations were reported as dry weights. Measurements reported as wet weight were converted to
equivalent dry weights using available percent moisture data for the relevant species and tissue type.
Species-specific percent moisture data for muscle tissue were available for bluegill (Gillespie and
Baumann 1986; Nakamoto and Hassler 1992), rainbow trout (Seiler and Skorupa 2001), and for a
composite average of nine fish species (May et al. 2000). Species specific percent moisture data for
ovaries were available for bluegill (Gillespie and Baumann 1986; Nakamoto and Hassler 1992), fathead
minnow (GEI Associates 2008; Rickwood et al. 2008), and rainbow trout (Seiler and Skorupa 2001).
Species-specific % moisture data for whole-body tissues were available for bluegill (USGS NCBP).
Measurements reported as wet weight were converted to equivalent dry weights using available percent
moisture data for the relevant species and tissue type. If percent moisture data were unavailable for a fish
species, percent moisture data for a similar species (i.e., same genus or, if unavailable, same family) were
used. Table B-l lists percent moisture by tissue type, species, data source, and the target species and
study for which the % moisture data were used to convert from wet to dry weight.
Table B-l. Percent moisture, by species and tissue type
% Moisture Data Source
Species
Study
% Moisture by Tissue
Whole-
body
Muscle
Ovary
Conversion Applied to
Species
Study
Used in derivation of FCV
Rainbow trout
Rainbow trout
Fathead
minnow
Bluegill
Seiler & Skorupa
2001
Seiler & Skorupa
2001
Average of GEI
Assoc. 2008;
Rickwood et al. 2008
Average of Gillespie
& Baumann 1986
and Nakamoto &
Hassler 1992
61.20
61.20
75.30
76.00
Rainbow trout
Brook trout
Fathead minnow
Bluegill
Holm et al. 2005
Holm et al. 2005
Schultz and
Hermanutz 1990
Hermanutz et al.
1996
B-2 Draft Document
-------
Avg of 9 spp
May et al. 2000
78.4
Striped bass
Coughlan and
Velte 1989
Used in conversion of FCV in egg/ovary to whole-body Se concentrations
Bluegill
Bluegill
Bluegill
Rainbow trout
Rainbow trout
Rainbow trout
Rainbow trout
Rainbow trout
Rainbow trout
USGS NCBP
May et al. 2000
Average of Gillespie
&Baumann 1986
and Nakamoto &
Hassler 1992
May et al. 2000
Seiler & Skorupa
2001
May et al. 2000
Seiler & Skorupa
2001
May et al. 2000
Seiler & Skorupa
2001
74.80
80.09
77.54
77.54
77.54
76.00
61.20
61.20
61.20
Bluegill
Bluegill
Bluegill
Brook Trout
Brook Trout
Rainbow Trout
Rainbow Trout
Rainbow Trout
Rainbow Trout
Hermanutz et al.
1996
Hermanutz et al.
1996
Hermanutz et al.
1996
Holm et al. 2005
Holm et al. 2005
Holm et al. 2005
Holm et al. 2005
Casey & Siwik
2000
Casey & Siwik
2000
Derivation of tissue conversion factors
Methodology
EPA used a mechanistic bioaccumulation modeling approach to derive a mathematical relationship
between the concentration of selenium in water to the concentration of selenium in the eggs and ovaries
offish. This approach characterizes selenium bioaccumulation as a series of steps representing the phase
transformation of selenium from dissolved to particulate form, and then the trophic transfer of selenium
through aquatic food webs to invertebrates and fish. The final step in this process is the transfer of
selenium into eggs and ovary tissue.
Equation 1 quantitatively models the transfer of selenium through each environmental compartment as a
series of site-specific and species-specific parameters. The parameter CFin Equation 1 represents the
species-specific proportion of selenium in egg or ovary tissue relative to the average concentration of
selenium in all body tissues and is given as:
c.
CF =
'egg-ovary
r
whole-body
(Equation 1)
B-3 Draft Document
-------
where
CF = Whole-body to egg-ovary conversion factor (dimensionless ratio).
Cegg-ovary = Selenium concentration in the eggs or ovaries offish (|ig/g dw)
Cwhoie-body = Selenium concentration in the whole body offish (|ig/g dw).
EPA derived species-specific conversion factor (CF) values using the same methods that were used to
derive species-specific TTF values from field data. EPA obtained matched pairs of selenium
measurements in the whole-body and eggs and/or ovaries offish from published scientific literature.
When both egg and ovary measurements were reported, EPA used the average. EPA first confirmed a
statistical relationship between egg-ovary and whole body selenium for each species using ordinary least
squares (OLS) linear regression. If the regression resulted in a statistically significant (P<0.05) positive
slope, EPA calculated the ratio of the egg-ovary to whole body selenium concentration for each matched
pair of measurements and used the median as the CF value for that species.
EPA derived CF values from selenium measurements in units of (ig/g dry weight. The majority of tissue
and whole body selenium concentrations were reported as dry weights. Measurements reported as wet
weight were converted to equivalent dry weights using available percent moisture data for the relevant
species and tissue type. If percent moisture data were unavailable for a fish species, percent moisture data
for a similar species (i.e., same genus or, if unavailable, same family) were used. A listing of percent
moisture concentrations by species and target tissue are provided in the bottom portion of Table B-l.
Studies that reported selenium measurements in egg-ovary tissue did not always report matched pairs of
selenium measurements in the whole-body. However, several of these studies did report selenium
measurements in muscle tissue. When matched pairs of selenium concentrations from egg-ovary and
muscle tissue were available, EPA calculated a species specific egg-ovary to muscle conversion factor
following the procedures used to calculate the egg-ovary to whole-body conversion factor. The species-
specific egg-ovary to muscle conversion factors were then converted to egg-ovary to whole-body
conversion factors by calculating and applying a single muscle to whole-body conversion factor.
The EPA developed species-specific egg-ovary to muscle and muscle to whole-body correction factors
following the procedure described for whole-body to egg-ovary conversion factors. The EPA obtained
matched pairs of selenium measurements in the whole-body and muscle filets and matched pairs of
selenium measurements in muscle filets and whole-body from published scientific literature. EPA first
confirmed a statistical relationship between the two tissue types for each species using OLS linear
regression. If the regression resulted in a significant fit with a positive slope, the EPA calculated the ratio
B-4 Draft Document
-------
of each matched pair of measurements and then calculated the median ratio. Because the number of
matched pairs of selenium measurements from muscle and whole-body were limited, the EPA took the
median of all the species-specific muscle to whole-body conversion factors to derive a single muscle to
whole-body conversion factor, and applied that conversion factor to the species-specific muscle to egg-
ovary conversion factors to derive species specific egg-ovary to whole-body conversion factors.The EPA
then used the average of the species-specific median ratios as the correction factor for all uncorrected CF
values.
CF values calculated directly from whole-body and egg-ovary selenium measurements
Cwhoie-body = Selenium concentration
Cegg = Selenium concentration
Covary = Selenium concentration
(—egg-ovary ~ Average selenium conc(
. . egg-owy
Katio —
whole-body
in all tissues ((ig/g dw)
in eggs ((ig/g dw)
in ovary tissue ((ig/g dw)
;ntration in eggs and ovari
[egg
4-C1 ^
ovary
2
,••
)
Black bullhead (Ameiurus melas)
Study
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
CP (
whole-body ^ess ^
5.30
4.80
5.50
4.90
9.60
7.60
7.30
6.60
8.60
2.00
5.30
-i
' ovary
64.30
35.40
52.80
56.00
42.80
38.70
37.30
34.30
26.40
56.70
64.30
^ess-ovary
64.30
35.40
52.80
56.00
42.80
38.70
37.30
34.30
26.40
56.70
64.30
Ratio
12.13
7.38
9.60
11.43
4.46
5.09
5.11
5.20
3.07
28.35
12.13
B-5 Draft Document
-------
^ess-ovary
60 n
40 -
20 -
Median ratio: 6.29
R2:
F:
df:
P:
0.37
4.67
8
0.063
20 40
/-<
^whole-body
60
Not used because P > 0.05 and negative
slope.
B-6 Draft Document
-------
Bluegill (Lepomis macrochirus)
Study
CP 1
whole-body ^egg '
Coyleetal. 1993
Coyleetal. 1993
Coyleetal. 1993
Coyleetal. 1993
Coyleetal. 1993
Doroshov et al
Doroshov et al
Doroshov et al
Doroshov et al
. 1992
. 1992
. 1992
. 1992
Hermanutz et al. 1996
Hermanutz et al. 1996
Hermanutz et al. 1996
Hermanutz et al. 1996
Hermanutz et al. 1996
Hermanutz et al. 1996
Hermanutz et al. 1996
Hermanutz et al. 1996
Hermanutz et al. 1996
Hermanutz et al. 1996
Hermanutz et al. 1996
Hermanutz et al. 1996
Hermanutz et al. 1996
Hermanutz et al. 1996
Hermanutz et al. 1996
Hermanutz et al. 1996
Hermanutz et al. 1996
Osmundson et
60 i
40 -
*- egg-ovary
20 -
n .
al. 2007
o
.
0
-------
Bluehead sucker (Catostomus discobolus)
Study
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
10 -I
C 5
'-egg-ovary
o -
C
Brown trout
Study
o x°
J/
o
5
^-whole-body
(Salmo truttd)
NewFields 2009
NewFields 2009
NewFields 2009
NewFields 2009
NewFields 2009
NewFields 2009
NewFields 2009
NewFields 2009
NewFields 2009
NewFields 2009
NewFields 2009
NewFields 2009
NewFields 2009
NewFields 2009
NewFields 2009
NewFields 2009
NewFields 2009
whole-body
1.30
2.00
2.10
2.20
2.40
3.90
5.60
10
whole-body
3.60
4.10
3.70
4.30
3.00
3.10
2.70
2.50
8.90
13.80
17.90
13.60
17.20
6.70
9.60
22.60
7.20
c c
'—egg '—ovary
2.40
4.20
3.70
4.00
4.10
7.10
8.10
Median ratio:
R2:
F:
df:
P:
C C
*-egg '-ovary
0.80
0.90
0.80
0.90
1.20
1.20
1.00
1.00
12.80
40.30
36.00
26.80
26.90
18.60
17.70
38.80
13.20
^egg-ovary
2.40
4.20
3.70
4.00
4.10
7.10
8.10
1.82
0.95
88.9
5
<0.001
^egg-ovary
0.80
0.90
0.80
0.90
1.20
1.20
1.00
1.00
12.80
40.30
36.00
26.80
26.90
18.60
17.70
38.80
13.20
Ratio
1.85
2.10
1.76
1.82
1.71
1.82
1.45
Ratio
0.22
0.22
0.22
0.21
0.40
0.39
0.37
0.40
1.44
2.92
2.01
1.97
1.56
2.78
1.84
1.72
1.83
B-8 Draft Document
-------
Brown trout (Salmo truttd)
NewFields 2009
NewFields 2009
NewFields 2009
NewFields 2009
NewFields 2009
NewFields 2009
NewFields 2009
NewFields 2009
NewFields 2009
NewFields 2009
NewFields 2009
NewFields 2009
NewFields 2009
NewFields 2009
NewFields 2009
NewFields 2009
NewFields 2009
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
9.20
13.20
8.60
11.30
20.00
8.40
5.60
6.70
5.90
6.00
7.00
5.60
4.70
7.20
9.20
5.50
8.50
4.60
4.30
5.00
5.50
13.40
20.50
12.50
11.20
28.10
12.80
8.40
8.50
8.40
9.10
7.50
6.60
6.90
6.20
14.00
6.90
9.50
1.20
37.80
35.60
32.50
13.40
20.50
12.50
11.20
28.10
12.80
8.40
8.50
8.40
9.10
7.50
6.60
6.90
6.20
14.00
6.90
9.50
1.20
37.80
35.60
32.50
1.46
1.55
1.45
0.99
1.41
1.52
1.50
1.27
1.42
1.52
1.07
1.18
1.47
0.86
1.52
1.25
1.12
0.26
8.79
7.12
5.91
50
25
'egg-ovary
25
/-<
^whole-body
50
Median ratio: 1.45
R2:
F:
df:
P:
0.47
31.3
36
<0.001
B-9 Draft Document
-------
Channel catfish (Ictaluruspunctatus)
Study
Osmundson
Osmundson
Osmundson
Osmundson
Cp
whole-body ^eg!
et
et
et
et
al.
al.
al.
al.
2007
2007
2007
2007
3
3
2
4
.40
.30
.60
.00
2 ^ovarv
29
21
13
30
Cegg-ovary Ratio
.50
.10
.70
.30
29
21
13
30
.50
.10
.70
.30
8
6
5
7
.68
.39
.27
.58
'egg-ovary
30 -
20 -
10 -
Median ratio: 6.98
R2: 0.82
F: 9.1
df: 2
P: 0.099
Not used because P > 0.05.
10 20
^whole-body
30
Common carp (Cyprinus carpio)
Study
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Cp
whole-body ^egi
et
et
et
et
et
al.
al.
al.
al.
al.
2007
2007
2007
2007
2007
6
4
11
23
4
.30
.80
.70
.10
.10
a ^ovarv
12.
9.
16.
27.
9.
^ess-ovarv
10
40
30
30
90
12
9
16
27
9
.10
.40
.30
.30
.90
Ratio
1.
1.
1.
1.
2.
92
96
39
18
41
30 n
20
'egg-ovary
10 -
Median ratio: 1.92
0 10 20
/-<
^whole-body
30
R2:
F:
df:
P:
0.96
584.8
3
<0.001
B-10 Draft Document
-------
Cutthroat trout (Oncorhynchus clarkii)
Study
Hardy 2005
Hardy 2005
Hardy 2005
Hardy 2005
Hardy 2005
Hardy 2005
Hardy 2005
Hardy 2005
Hardy 2005
Hardy 2005
20 -,
15 -
egg-ovary
5 -
n -
Cp
whole-body ^egg
0
2
2
6
1
4
5
9
11
5
0
o
o
0°
70
60
80
40
20
60
90
10
40
60
1
3
5
18
1
7
6
5
5
16
^ovarv
00
80
50
00
60
80
60
10
20
00
Median ratio:
R2:
F:
df:
P:
^egg-ovary
1
3
5
18
1
7
6
5
5
16
1.45
0.11
1.01
8
0.406
00
80
50
00
60
80
60
10
20
00
Ratio
1
1
1
2
1
1
1
0
0
2
43
46
96
81
33
70
12
56
46
86
12
Not used because P > 0.05.
whole-body
B-11 Draft Document
-------
Flannelmouth sucker (Catostomus latipinnis)
Study
Cp
whole-body ^es
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
8 -I
*" egg-ovary
0 -
(
2007
2007
2007
2007
2007
2007
2007
o ->
^f
3
2
3
3
3
4
4
00
60
10
10
50
40
50
g ^ovarv
4
4
5
4
5
6
6
Median ratio
R
F
^egg-ovary
00
10
90
30
70
20
20
2
df:
)
4
^whole-body
8
P
4
4
5
4
5
6
6
1.41
0.65
9.2
5
0.021
00
10
90
30
70
20
20
Ratio
1.33
1.58
1.90
1.39
1.63
1.41
1.38
Green sunfish (Lepomis cyanellus)
Study
^whole-body *-*es
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
2007
2007
2007
2007
2007
2007
2007
2007
2007
2007
2007
2007
2007
2007
22
8
15
4
5
4
3
11
6
9
9
6
7
7
80
80
40
80
70
40
80
90
40
50
10
20
00
70
g ^ovarv
27
10
21
7
8
6
6
18
12
13
15
10
11
12
^egg-ovary
40
20
80
00
90
40
40
10
30
80
20
80
70
60
27
10
21
7
8
6
6
18
12
13
15
10
11
12
40
20
80
00
90
40
40
10
30
80
20
80
70
60
Ratio
1.20
1.16
1.42
1.46
1.56
1.45
1.68
1.52
1.92
1.45
1.67
1.74
1.67
1.64
B-12 Draft Document
-------
Green sunfish (Lepomis cyanellus)
Study
Osmundson et
Osmundson et
Osmundson et
Osmundson et
Osmundson et
Osmundson et
Osmundson et
Osmundson et
Osmundson et
Osmundson et
Osmundson et
Osmundson et
Osmundson et
Osmundson et
Osmundson et
Osmundson et
Osmundson et
Osmundson et
Osmundson et
Osmundson et
Osmundson et
Osmundson et
Osmundson et
Osmundson et
30 -i
20 -
^-•egg-ovary
10 •
n .
al. 2007
al. 2007
al. 2007
al. 2007
al. 2007
al. 2007
al. 2007
al. 2007
al. 2007
al. 2007
al. 2007
al. 2007
al. 2007
al. 2007
al. 2007
al. 2007
al. 2007
al. 2007
al. 2007
al. 2007
al. 2007
al. 2007
al. 2007
al. 2007
o/
M^
^whole-body *^e
6.20
10.20
9.70
9.90
7.20
9.00
9.70
8.90
9.80
9.90
10.30
5.30
10.10
11.80
3.30
4.00
4.30
3.70
6.20
3.50
4.40
5.60
4.90
4.40
>ft
/^
X
*g ^ ovary
10.00
13.90
15.20
14.70
8.80
12.90
13.10
11.50
13.20
11.60
7.50
8.10
13.20
14.00
5.20
5.80
4.10
4.90
9.50
4.80
5.60
10.10
7.50
5.90
Median ratio:
R2:
F:
df:
P:
^ egg-ovary
10.00
13.90
15.20
14.70
8.80
12.90
13.10
11.50
13.20
11.60
7.50
8.10
13.20
14.00
5.20
5.80
4.10
4.90
9.50
4.80
5.60
10.10
7.50
5.90
1.45
0.87
240.0
36
< 0.001
Ratio
1.61
1.36
1.57
1.48
1.22
1.43
1.35
1.29
1.35
1.17
0.73
1.53
1.31
1.19
1.58
1.45
0.95
1.32
1.53
1.37
1.27
1.80
1.53
1.34
0 10 20
/~<
^whole-body
30
B-13 Draft Document
-------
Roundtail chub (Gila robusta)
Study
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Cp
whole-body ^egi
4.10
5.30
6.40
6.80
5.50
6.60
8.40
a ^ ovary
7.90
10.80
15.20
14.10
10.60
18.00
17.80
^ egg-ovary
7.90
10.80
15.20
14.10
10.60
18.00
17.80
Ratio
1.93
2.04
2.38
2.07
1.93
2.73
2.12
20
10
'egg-ovary
10
~<
whole-body
20
Median ratio: 2.07
R2:
F:
df:
P:
0.80
20.4
5
0.004
Smallmouth bass (Micropterus dolomieu)
Study
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
C/~<
whole-body ^egi
4.20
5.50
5.40
7.80
5.10
4.90
g ^ ovary *-
6.00
8.00
6.50
11.00
7.10
8.80
' egg-ovary K3.ll 0
6.00
8.00
6.50
11.00
7.10
8.80
1.43
1.45
1.20
1.41
1.39
1.80
12 n
c,.
6 -
gg-ovary
Median ratio: 1.42
R2:
F:
df:
P:
0.73
10.6
4
0.026
12
'whole-body
B-14 Draft Document
-------
White sucker (Catostomus commersonii)
Study
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Cp
whole-body ^egi
3.80
4.20
3.30
4.50
6.30
6.80
11.00
12.70
5.70
3.90
3.80
9.90
5.30
10.70
5.90
7.00
6.40
6.30
5.30
6.20
5.60
8.80
8.70
11.40
10.70
8.40
7.00
7.50
10.30
6.70
2.10
1.80
3.20
2.30
3.10
3.00
2.80
2.50
3.40
2.80
a ^ ovary
6.20
6.20
5.20
6.50
7.70
5.80
10.90
11.20
9.40
5.40
5.10
10.40
10.40
11.00
11.70
11.60
9.40
10.20
7.30
8.90
10.50
10.20
8.10
9.50
10.70
8.30
12.00
6.10
6.10
11.30
2.60
3.60
4.40
4.40
4.80
4.30
4.10
3.80
3.60
3.80
^egg-ovary
6.20
6.20
5.20
6.50
7.70
5.80
10.90
11.20
9.40
5.40
5.10
10.40
10.40
11.00
11.70
11.60
9.40
10.20
7.30
8.90
10.50
10.20
8.10
9.50
10.70
8.30
12.00
6.10
6.10
11.30
2.60
3.60
4.40
4.40
4.80
4.30
4.10
3.80
3.60
3.80
Ratio
1.63
1.48
1.58
1.44
1.22
0.85
0.99
0.88
1.65
1.38
1.34
1.05
1.96
1.03
1.98
1.66
1.47
1.62
1.38
1.44
1.88
1.16
0.93
0.83
1.00
0.99
1.71
0.81
0.59
1.69
1.24
2.00
1.38
1.91
1.55
1.43
1.46
1.52
1.06
1.36
B-15 Draft Document
-------
15 -\
10 -
*" egg-ova ly
5 -
O O
5 10
/-<
^whole-body
Median ratio: 1.41
R2: 0.54
F: 45.4
df: 38
P: < 0.001
15
Table B-2. Summary of whole-body to egg-ovary conversion factors (CF) from matched pairs of
whole-body and egg-ovary measurements.
Common name
bluegill
bluehead sucker
brown trout
common carp
flannelmouth sucker
green sunfish
roundtail chub
smallmouth bass
white sucker
Scientific name
Lepomis macrochirus
Catostomus discobolus
Salmo trutta
Cyprinus carpio
Catostomus latipinnis
Lepomis cyanellus
Gila robusta
Micropterus dolomieu
Catostomus commersonii
Median ratio (CF)
2.13
1.82
1.45
1.92
1.41
1.45
2.07
1.42
1.41
B-16 Draft Document
-------
Muscle to egg-ovary conversion factors
^ ovary
-'egg-ovary
Ratio
= Selenium concentration in muscle tissue only (|ig/g dw)
= Selenium concentration in eggs ((ig/g dw)
= Selenium concentration in ovary tissue ((ig/g dw)
(C +C
A 1 • j. j.- • J egg ovary
= Average selenium concentration in eggs and ovaries
C
egg-ovary
C
muscle
Black bullhead
Study
(Ameiurus melas)
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
80 •
60 •
C 40 -
'"egg-ovary
20 •
0 -
C
o
0
246
r
'"muscle
^muscle '"egg ^
3.40
3.90
4.30
4.70
5.70
7.40
7.50
7.80
7.80
9.20
o
o
8 10
Median
-^
^ ovary
64.30
35.40
52.80
56.00
42.80
38.70
37.30
34.30
26.40
56.70
ratio:
R2:
F:
df:
P:
Not used because P >
slope.
*^ egg-ovary
64.30
35.40
52.80
56.00
42.80
38.70
37.30
34.30
26.40
56.70
6.84
0.17
1.65
8
0.250
Ratio
18.91
9.08
12.28
11.91
7.51
5.23
4.97
4.40
3.38
6.16
0.05 and negative
B-17 Draft Document
-------
Bluegill (Lepomis macrochirus)
Study
Brysonetal. 1984
Brysonetal. 1985a(pt. 1)
Brysonetal. 1985a(pt. 1)
Brysonetal. 1985a(pt. 2)
Doroshov et al.
Doroshov et al.
Doroshov et al.
Doroshov et al.
Hermanutz et al
Hermanutz et al
Hermanutz et al
Hermanutz et al
Hermanutz et al
Hermanutz et al
Hermanutz et al
Hermanutz et al
Hermanutz et al
Hermanutz et al
Hermanutz et al
Hermanutz et al
Hermanutz et al
Hermanutz et al
Hermanutz et al
Hermanutz et al
Hermanutz et al
Hermanutz et al
Hermanutz et al
Hermanutz et al
1992
1992
1992
1992
. 1996
. 1996
. 1996
. 1996
. 1996
. 1996
. 1996
. 1996
. 1996
. 1996
. 1996
. 1996
. 1996
. 1996
. 1996
. 1996
. 1996
. 1996
. 1996
. 1996
Osmundson et al. 2007
60 -I
40 -
c
egg-ovary
20 -
n .
o
o ^X^
0 ^X
§ .x*^
qjgrtP
«P ° °
W
\J vfrp j| j jj
0 20 40 60
r
^muscle
Cp
muscle ^es
84.0
59.0
2.7
25.0
1.5
5.8
10.4
23.6
1.6
8.5
14
2.1
20.6
1.9
3.5
17.6
12.5
2.3
6.9
44.9
39.8
5.3
12.5
7.8
3.2
6.1
18.7
15.1
12.9
^
^ o
80 100
g ^ovarv
49.0
30.0
2.2
9.1
2.8
8.3
19.5
38.4
2.0
18.8
15.5
0.3
16.7
4.4
8.4
29.0
24.5
3.2
10.3
42.1
55.0
7.0
26.0
14.9
4.4
7.9
16.3
15.9
9.7
Median ratio:
R2:
F:
df:
P:
^egg-ovary
49.0
30.0
2.2
9.1
2.8
8.3
19.5
38.4
2.0
18.8
15.5
0.3
16.7
4.4
8.4
29.0
24.5
3.2
10.3
42.1
55.0
7.0
26.0
14.9
4.4
7.9
16.3
15.9
9.7
1.38
0.65
50.37
27
<0.001
Ratio
0.58
0.51
0.81
0.36
1.87
1.43
1.88
1.63
1.25
2.21
1.11
0.14
0.81
2.32
2.40
1.65
1.96
1.39
1.49
0.94
1.38
1.32
2.08
1.91
1.38
1.30
0.87
1.05
0.75
B-18 Draft Document
-------
Bluehead sucker (Catostomus discobolus)
Study
muscle
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
egg-ovary | Oa*t^^^
1 o
0 5 10
r
^muscle
\^ egg ^ ovary ^ egg-ovary Iv3.ll 0
1.5
2.3
2.5
2.7
3.1
5.2
8.6
2.4 2.4
4.2 4.2
3.7 3.7
4 4
4.1 4.1
7.1 7.1
8.1 8.1
Median ratio: 1.48
R2: 0.91
F: 47.70
df: 5
P: <0.001
1.60
1.83
1.48
1.48
1.32
1.37
0.94
Brook trout (Salvelinusfontinalis)
Study
Holm et al. 2005
Holm et al. 2005
Holm et al. 2005
Holm et al. 2005
Holm et al. 2005
Holm et al. 2005
Holm et al. 2005
Holm et al. 2005
Holm et al. 2005
Holm et al. 2005
Holm et al. 2005
Holm et al. 2005
Holm et al. 2005
Holm et al. 2005
Holm et al. 2005
Holm et al. 2005
Holm et al. 2005
muscle
2
1
2
2
2
5
9
10
11
11
12
15
16
19
20
23
34
\^ egg ^ ovary ^ egg-ovary Kail 0
.80
.40
.20
.00
.20
.00
.70
.50
.20
.40
.30
.90
.50
.60
.40
.40
.70
1.50
2.50
3.40
4.70
2.90
5.60
9.90
15.40
12.80
14.80
12.20
12.40
13.20
15.50
15.30
25.40
32.50
1.50
2.50
3.40
4.70
2.90
5.60
9.90
15.40
12.80
14.80
12.20
12.40
13.20
15.50
15.30
25.40
32.50
0.54
1.79
1.55
2.35
1.32
1.12
1.02
1.47
1.14
1.30
0.99
0.78
0.80
0.79
0.75
1.09
0.94
B-19 Draft Document
-------
40 n
egg-ovary
20 -
Median ratio: 1.09
R2:
F:
df:
P:
0.91
152.3
15
< 0.001
0 20
Brown trout
Study
^ muscle
(Salmo trutta)
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
40
CP P P
muscle *^egg ^ovarv *^ egg-ovary
3.2 - 1.2 1.2
3.6 - 37.8 37.8
4 - 35.6 35.6
6.3 - 32.5 32.5
Ratio
0.38
10.50
8.90
5.16
40 -I
30 -
egg-ovary
10 -
c
O -
o v'
^/^ o
'
Q j
5 10
r
^^muscle
Median ratio: 7.03
R2: 0.17
F: 0.40
df: 2
P: 0.71
Not used because P > 0.05.
B-20 Draft Document
-------
Channel catfish (Ictalurispunctatus)
Study
muscle
Ratio
Dsmundson et al. 2007 3.4 - 29.5
Dsmundson et al. 2007 3.6 - 21.1
Dsmundson et al. 2007 3.7 - 13.7
Dsmundson et al. 2007 5.3 - 30.3
40 -
30 -
C 20 -
'"egg-ovary
10
X
0
"I "I
0 5 10
C
'"muscle
Median ratio:
R2:
F:
df:
P:
Not used because P >
29.5
21.1
13.7
30.3
5.79
0.20
0.49
2
0.67
0.05.
8.68
5.86
3.70
5.72
Common carp
Study
(Cyprinus carpio)
CP P P
muscle '"ess '"ovarv '"ess-ovarv
3arcia-Hernandez 2000 4.6 - 1.8 1.8
Dsmundson et al. 2007 7.8 - 12.1 12.1
Dsmundson et al. 2007 8.2 - 9.4 9.4
Dsmundson et al. 2007 20 - 16.3 16.3
Dsmundson et al. 2007 24.2 - 27.3 27.3
Dsmundson et al. 2007 6.6 - 9.9 9.9
30
20 -
c
^ egg-ovary
10
0
•
^^^^
^^^^ o
o ^"^
QO^
o
? r i
0 10 20 30
C ,
'"muscle
Median ratio: 1.14
R2: 0.84
F: 21.7
df: 4
P: 0.007
Ratio
0.39
1.55
1.15
0.82
1.13
1.50
B-21 Draft Document
-------
Cutthroat trout (Oncorhynchus clarkii)
Study
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Kennedy et al. 2000
Kennedy et al. 2000
Kennedy et al. 2000
Kennedy et al. 2000
Kennedy et al. 2000
Kennedy et al. 2000
Kennedy et al. 2000
Kennedy et al. 2000
Kennedy et al. 2000
Kennedy et al. 2000
Cp
muscle ^es
6.80
4.20
3.00
4.90
4.50
4.00
5.00
5.00
5.00
8.40
8.30
7.00
6.60
8.40
9.80
8.50
16.00
7.00
8.00
7.00
7.00
9.00
7.00
7.00
8.00
9.80
7.00
9.00
7.00
8.00
10.00
41.30
15.30
14.10
12.50
13.70
14.30
9.50
9.40
8.70
9.50
!g ^ ovary
28.20
47.80
22.00
9.80
8.20
7.00
10.00
10.00
8.00
16.20
18.30
14.30
14.30
14.70
16.40
15.90
20.00
14.00
19.00
14.00
14.00
16.00
13.00
14.00
14.00
20.20
22.00
16.00
12.00
13.00
14.00
75.40 66.80
58.40 31.60
30.60 31.40
20.20 18.50
19.40 19.50
16.20 16.20
16.10 19.30
14.40 22.00
13.20 17.00
12.60 13.60
^egg-ovary
28.20
47.80
22.00
9.80
8.20
7.00
10.00
10.00
8.00
16.20
18.30
14.30
14.30
14.70
16.40
15.90
20.00
14.00
19.00
14.00
14.00
16.00
13.00
14.00
14.00
20.20
22.00
16.00
12.00
13.00
14.00
71.10
45.00
31.00
19.35
19.45
16.20
17.70
18.20
15.10
13.10
Ratio
4.15
11.38
7.33
2.00
1.82
1.75
2.00
2.00
1.60
1.93
2.20
2.04
2.17
1.75
1.67
1.87
1.25
2.00
2.38
2.00
2.00
1.78
1.86
2.00
1.75
2.06
3.14
1.78
1.71
1.63
1.40
1.72
2.94
2.20
1.55
1.42
1.13
1.86
1.94
1.74
1.38
B-22 Draft Document
-------
Cutthroat trout (Oncorhynchus clarkii)
Study
Kennedy et al.
Kennedy et al.
Kennedy et al.
Kennedy et al.
Kennedy et al.
Kennedy et al.
2000
2000
2000
2000
2000
2000
Rudolph et al. 2007
Rudolph et al. 2007
Rudolph et al. 2007
Rudolph et al. 2007
Rudolph et al. 2007
Rudolph et al. 2007
Rudolph et al. 2007
Rudolph et al. 2007
Rudolph et al. 2007
Rudolph et al. 2007
Rudolph et al. 2007
Rudolph et al. 2007
Rudolph et al. 2007
Rudolph et al. 2007
Rudolph et al. 2007
Rudolph et al. 2007
Rudolph et al. 2007
Rudolph et al. 2007
Rudolph et al. 2007
Rudolph et al. 2007
Rudolph et al. 2007
Rudolph et al. 2007
150 •
100 •
^ egg-ovary
50 -
n -
0
0
muscle
10.20
10.70
6.60
9.70
10.90
6.90
7.70
8.20
8.00
8.10
6.60
8.50
7.20
7.30
7.60
8.70
8.20
7.90
7.60
11.80
40.40
46.10
50.40
34.70
39.00
35.40
11.30
13.40
c c
^egg ^ovarv
12.30 14.50
10.50 20.60
9.90 21.50
9.10 13.20
8.50 13.40
13.20 20.30
13.90
12.50
15.00
14.90
15.20
12.90
12.30
16.70
13.10
15.60
13.90
15.10
12.30
16.10
86.30
121.00
140.00
51.00
65.30
46.80
16.90
20.60
Median ratio:
R2:
F:
df:
P:
^egg-ovary
13.40
15.55
15.70
11.15
10.95
16.75
13.90
12.50
15.00
14.90
15.20
12.90
12.30
16.70
13.10
15.60
13.90
15.10
12.30
16.10
86.30
121.00
140.00
51.00
65.30
46.80
16.90
20.60
1.81
0.82
308.3
67
< 0.001
Ratio
1.31
1.45
2.38
1.15
1.00
2.43
1.81
1.52
1.88
1.84
2.30
1.52
1.71
2.29
1.72
1.79
1.70
1.91
1.62
1.36
2.14
2.62
2.78
1.47
1.67
1.32
1.50
1.54
50
100
150
*^mu
iscle
B-23 Draft Document
-------
Dolly varden (Salvelinus malma)
Study
Colder 2009
Colder 2009
Colder 2009
Colder 2009
Colder 2009
Colder 2009
Colder 2009
Colder 2009
Colder 2009
Colder 2009
Colder 2009
Colder 2009
Colder 2009
Colder 2009
Colder 2009
Colder 2009
Colder 2009
200 -I
150 -
100 -
»- egg-ovary
50 •
muscle
73
45
107
97
114
115
79
9
3
5
2
4
6
55
58
39
50
o
.00
.90
.00
.20
.00
.00
.60
.90
.40
.30
.80
.90
.60
.70
.30
.50
.50
*^egg
92
40
107
102
124
185
112
7
12
9
5
10
11
65
51
60
56
^ovarv
30
70
00
00
00
00
00
00
10
60
40
50
00
80
90
50
60
Median ratio:
/>
R2:
F:
df:
P:
*^egg-ov
92
40
107
102
124
185
112
7
12
9
5
10
11
65
51
60
56
1.26
0.90
140.3
15
ary
.30
.70
.00
.00
.00
.00
.00
.00
.10
.60
.40
.50
.00
.80
.90
.50
.60
Ratio
1
0
1
1
1
1
1
0
3
1
1
2
1
1
0
1
1
26
89
00
05
09
61
41
71
56
81
93
14
67
18
89
53
12
< 0.001
0 50 100 150 200
Flannelmouth
Study
muscle
sucker (Catostomus latipinnis)
^muscle
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
3.6
3.8
4.1
4.6
5.2
6.2
7.3
^egg
^ ovary
4
4
5
4
5
6
6
^egg-ovary
0
1
9
3
7
2
2
4.0
4.1
5.9
4.3
5.7
6.2
6.2
Ratio
1
1
1
0
1
1
0
11
08
44
93
10
00
85
B-24 Draft Document
-------
c
'"egg-ovary
10 -
5 -
0 -
°O^S>^>
^°
^^^^^^_^^^^^_,^^^^^_^^^^^_l
0 5 10
r
'"muscle
Median ratio: 1.08
R2: 0.58
F: 6.92
df: 5
P: 0.036
Green sunfish (Lepomis cyanellus)
Study
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
muscle
err
*^egg ^ovarv ^egg-ovary
etal. 2007 28.1 - 27.4 27.4
etal. 2007 12.9 - 10.2 10.2
etal. 2007 21.9 - 21.8 21.8
et al. 2007
et al. 2007
et al. 2007
et al. 2007
5-77
6.1 - 8.9 8.9
5.2 - 6.4 6.4
5.1 - 6.4 6.4
etal. 2007 15.7 - 18.1 18.1
etal. 2007 10.1 - 12.3 12.3
et al. 2007 1
1.5 - 13.8 13.8
etal. 2007 10.5 - 15.2 15.2
et al. 2007
et al. 2007
et al. 2007
et al. 2007
et al. 2007
7.2 - 10.8 10.8
9.3 - 11.7 11.7
7.7 - 12.6 12.6
6 - 10 10
12 - 13.9 13.9
etal. 2007 12.1 - 15.2 15.2
etal. 2007 12.5 - 14.7 14.7
et al. 2007
et al. 2007 1
7.5 - 8.8 8.8
1.3 - 12.9 12.9
etal. 2007 13.6 - 13.1 13.1
etal. 2007 13.2 - 11.5 11.5
etal. 2007 12.4 - 13.2 13.2
etal. 2007 12.5 - 11.6 11.6
et al. 2007
et al. 2007
et al. 2007 1
8.6 - 7.5 7.5
5.3 - 8.1 8.1
1.9 - 13.2 13.2
etal. 2007 13.6 - 14 14
Ratio
0.98
0.79
1.00
1.40
1.46
1.23
1.25
1.15
1.22
1.20
1.45
1.50
1.26
1.64
1.67
1.16
1.26
1.18
1.17
1.14
0.96
0.87
1.06
0.93
0.87
1.53
1.11
1.03
B-25 Draft Document
-------
Green sunfish (Lepomis cyanellus)
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
30 -j
20
egg-ovary
10
0 -
^^
<&
™™™™™^^
) 10 20
r
^muscle
S"
30
3.8
4.2
4.1
4.2
5.7
4.4
3.5
5.5
5
4.3
5.2
5.8
4.1
4.9
9.5
4.8
5.6
10.1
7.5
5.9
Median ratio:
R2:
F:
df:
P:
5.2
5.8
4.1
4.9
9.5
4.8
5.6
10.1
7.5
5.9
1.21
0.89
281.4
36
<0.001
1.37
1.38
1.00
1.17
1.67
1.09
1.60
1.84
1.50
1.37
Mountain whitefish (Prosopium williamsoni)
Study
Golder 2005
Golder 2005
Golder 2005
Golder 2005
Golder 2005
Golder 2005
Golder 2005
Golder 2005
Golder 2005
Golder 2005
Golder 2005
Golder 2005
Golder 2005
Golder 2005
Golder 2005
Golder 2005
Golder 2005
muscle
3
3
3
4
3
3
5
5
5
7
7
5
7
3
4
4
5
ce£
.60
.70
.10
.20
.90
.50
.20
.00
.20
.60
.20
.50
.80
.70
.70
.40
.70
c
26.90
25.80
20.00
19.30
19.20
23.20
38.00
41.00
32.00
34.00
32.00
40.00
39.70
20.30
22.40
28.90
30.10
^egg-ovary
26.90
25.80
20.00
19.30
19.20
23.20
38.00
41.00
32.00
34.00
32.00
40.00
39.70
20.30
22.40
28.90
30.10
Ratio
7.47
6.97
6.45
4.60
4.92
6.63
7.31
8.20
6.15
4.47
4.44
7.27
5.09
5.49
4.77
6.57
5.28
B-26 Draft Document
-------
Mountain whitefish (Prosopium williamsoni)
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
Colder 2005
50 i
r 25 •
^ egg-ovary
0 •
(
J^°
£°
) 25
^muscle
4.00
10.00
4.90
7.60
6.10
6.80
5.00
6.60
5.00
4.80
50
-
-
-
-
-
-
-
-
-
-
Median
31.50
35.20
26.70
26.80
29.70
41.10
29.00
34.50
36.30
28.90
ratio:
R2:
F:
df:
P:
31.50
35.20
26.70
26.80
29.70
41.10
29.00
34.50
36.30
28.90
5.80
0.33
12.4
25
<0.001
7.88
3.52
5.45
3.53
4.87
6.04
5.80
5.23
7.26
6.02
Northern pike (Esox lucius)
Study
Muscatello et al. 2006
Muscatello et al. 2006
Muscatello et al. 2006
Muscatello et al. 2006
Muscatello et al. 2006
Muscatello et al. 2006
Muscatello et al. 2006
Muscatello et al. 2006
Muscatello et al. 2006
Muscatello et al. 2006
Muscatello et al. 2006
Muscatello et al. 2006
Muscatello et al. 2006
Muscatello et al. 2006
Cp
muscle ^e
0.90
1.90
2.60
1.30
1.00
17.00
16.50
16.50
2.00
2.00
1.30
2.50
1.30
47.80
<
3.50
2.70
3.40
3.70
2.70
43.20
24.50
26.10
3.40
4.10
4.10
4.10
3.40
48.20
^ ovary
-
-
-
-
-
-
-
-
-
-
-
-
-
-
^egg-ovary
3.50
2.70
3.40
3.70
2.70
43.20
24.50
26.10
3.40
4.10
4.10
4.10
3.40
48.20
Ratio
3.89
1.42
1.31
2.85
2.70
2.54
1.48
1.58
1.70
2.05
3.15
1.64
2.62
1.01
B-27 Draft Document
-------
r
t>0 -
30 -
*^ egg-ovary
4
0
x
O ./r
*/
y^
/
s
30
^muscle
X
o
60
Median ratio:
R2:
F:
df:
P:
1.88
0.83
58.9
12
<0.001
Rainbow trout (Oncorhynchus mykiss)
Study
Casey
Casey
Casey
Casey
Casey
Casey
Casey
Casey
Casey
Casey
Casey
Casey
Casey
Casey
Casey
Casey
Casey
Casey
Casey
Casey
Casey
Casey
Casey
Casey
Casey
Casey
Casey
Casey
and
and
and
and
and
and
and
and
and
and
and
and
and
and
and
and
and
and
and
and
and
and
and
and
and
and
and
and
Siwik
Siwik
Siwik
Siwik
Siwik
Siwik
Siwik
Siwik
Siwik
Siwik
Siwik
Siwik
Siwik
Siwik
Siwik
Siwik
Siwik
Siwik
Siwik
Siwik
Siwik
Siwik
Siwik
Siwik
Siwik
Siwik
Siwik
Siwik
2000
2000
2000
2000
2000
2000
2000
2000
2000
2000
2000
2000
2000
2000
2000
2000
2000
2000
2000
2000
2000
2000
2000
2000
2000
2000
2000
2000
^muscle *^e
4.10
3.80
2.60
3.30
2.30
2.80
2.30
2.80
3.00
4.90
1.50
2.60
4.60
4.60
3.60
2.40
3.70
2.70
0.70
0.60
0.60
28.60
30.90
32.40
28.00
31.70
29.50
30.10
*g ^ ovary
11.60
10.10
0.10
4.90
3.60
5.30
3.70
6.40
5.20
6.80
3.60
6.90
6.90
6.40
5.50
10.50
7.60
4.10
1.10
0.90
1.30
56.30
56.00
71.50
61.30
54.50
56.80
57.90
^ egg-ovary
11.60
10.10
0.10
4.90
3.60
5.30
3.70
6.40
5.20
6.80
3.60
6.90
6.90
6.40
5.50
10.50
7.60
4.10
1.10
0.90
1.30
56.30
56.00
71.50
61.30
54.50
56.80
57.90
Ratio
2
2
0
83
66
04
1.48
1
1
1
2
1
1
57
89
61
29
73
39
2.40
2
1
1
1
4
2
1
1
1
2
1
1
2
2
1
1
1
65
50
39
53
38
05
52
57
50
17
97
81
21
19
72
93
92
B-28 Draft Document
-------
Rainbow trout (Oncorhynchus mykiss)
Casey and Siwik 2000 29.90 64.70
Casey and Siwik 2000 32.80 46.60
Casey and Siwik 2000 3 1 .40 56.50
Casey and Siwik 2000 32.00 67.50
Casey and Siwik 2000 35.70 59.40
Casey and Siwik 2000 24.60 48.70
Casey and Siwik 2000 30.30 69.10
Casey and Siwik 2000 25.70 43.50
Casey and Siwik 2000 35.00 58.10
Casey and Siwik 2000 33.80 59.20
Casey and Siwik 2000 28.70 55.00
Casey and Siwik 2000 25.80 49.00
Holm et al. 2005 1.70 1.00
Holm et al. 2005 1.60 3.50
Holm et al. 2005 1.30 4.60
Holm et al. 2005 4.00 12.80
Holm et al. 2005 4.30 17.10
Holm et al. 2005 8.50 17.50
Holm et al. 2005 7.40 29.70
80 -,
«o/ Median ratio:
r 40 -
^ egg-ovary
m®
J%° R2:
/ F
0 / df:
o/ P:
f
64.70 2.16
46.60 1.42
56.50 1.80
67.50 2.11
59.40 1.66
48.70 1.98
69.10 2.28
43.50 1.69
58.10 1.66
59.20 1.75
55.00 1.92
49.00 1.90
1.00 0.59
3.50 2.19
4.60 3.54
12.80 3.20
17.10 3.98
17.50 2.06
29.70 4.01
1.92
0.96
990.0
45
<0. 001
0 40 80
^muscle
B-29 Draft Document
-------
Razorback sucker (Xyrauchen texanus)
Study
Hamilton et al. 2005 a,b,c
Hamilton et al. 2005 a,b,c
Hamilton et al. 2005 a,b,c
Hamilton et al. 2005 a,b,c
Hamilton et al. 2005 a,b,c
Hamilton et al. 2005 a,b,c
Hamilton et al. 2005 a,b,c
Hamilton et al. 2005 a,b,c
Hamilton et al. 2005 a,b,c
Hamilton et al. 2005 a,b,c
Waddell and May 1995 a
Waddell and May 1995 a
Waddell and May 1995 a
50 -,
/)
T 1
0 25
muscle
a Data from this study labeled above with
Cp
muscle ^e
6.30
15.60
29.20
5.10
5.80
13.50
16.20
6.00
12.50
18.00
4.40
7.10
32.00
1
50
*g ^ovarv
6.50 7.00
46.50 30.60
37.80 45.50
6.00
5.90
27.50
42.10
5.10
10.00
12.90
3.70
4.70
10.60
Median ratio:
R2:
F:
df:
P:
^egg-ovary
6.75
38.55
41.65
6.00
5.90
27.50
42.10
5.10
10.00
12.90
3.70
4.70
10.60
1.12
0.61
12.5
8
0.008
Ratio
1.07
2.47
1.43
1.18
1.02
2.04
2.60
0.85
0.80
0.72
X
X
X
'x's' were excluded because results appeared atypical.
Roundtail chub (Gila robustd)
Study
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Cp
muscle ^e
4.3
5
6.2
6.9
7
7.3
9.8
*g ^ovarv
7.9
10.8
15.2
-
10.6
18
17.8
^egg-ovary
7.9
10.8
15.2
14.1
10.6
18
17.8
Ratio
1.84
2.16
2.45
2.04
1.51
2.47
1.82
B-30 Draft Document
-------
30 -I
20
C °.»^^
'"egg-ovary ^^r
10 J x»^ 0
0
0 \ r , ,
0 5 10 15
r
'"muscle
Median ratio: 2.04
R2: 0.62
F: 8.27
df: 5
P: 0.026
Smallmouth bass (Micropterus dolomieu)
^tllfly ^muscle ^ess ^ovarv ^egg-ovarv
Osmundson et al. 2007 3.7 - 6.0 6.0
Osmundson et al. 2007 6.5 - 8.0 8.0
Osmundson et al. 2007 6.9 - 6.5 6.5
Osmundson et al. 2007 11 11
Osmundson et al. 2007 5.5 - 7.1 7.1
Osmundson et al. 2007 7.7 - 8.8 8.8
15 -I
c^""
*"egg-ovary O***^**^
5 -
0 + r , ,
0 5 10 15
r
'"muscle
Median ratio: 1.19
R2: 0.85
F: 23.5
df: 4
P: 0.006
Ratio
1.62
1.23
0.94
1.00
1.29
1.14
B-31 Draft Document
-------
White Sucker (Catostomus commersonii)
Study
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Cp
muscle ^eg!
2.9
4.8
3.7
3.7
8.4
9.4
15.5
23.6
9.4
6.1
4.6
12.3
9.2
9.4
9.4
10.5
11.4
9.6
9.3
9.8
10.5
11.1
12.1
12.8
16.0
12.1
9.0
10.6
12.6
11.6
2.8
2.5
4.3
3.5
4.3
3.1
3.6
3.0
4.1
3.6
Z ^ ovary
6.2
6.2
5.2
6.5
7.7
5.8
10.9
11.2
9.4
5.4
5.1
10.4
10.4
11
11.7
11.6
9.4
10.2
7.3
8.9
10.5
10.2
8.1
9.5
10.7
8.3
12
6.1
6.1
11.3
2.6
3.6
4.4
4.4
4.8
4.3
4.1
3.8
3.6
3.8
^egg-ovary
6.2
6.2
5.2
6.5
7.7
5.8
10.9
11.2
9.4
5.4
5.1
10.4
10.4
11
11.7
11.6
9.4
10.2
7.3
8.9
10.5
10.2
8.1
9.5
10.7
8.3
12
6.1
6.1
11.3
2.6
3.6
4.4
4.4
4.8
4.3
4.1
3.8
3.6
3.8
Ratio
2.14
1.29
1.41
1.76
0.92
0.62
0.70
0.47
1.00
0.89
1.11
0.85
1.13
1.17
1.24
1.10
0.82
1.06
0.78
0.91
1.00
0.92
0.67
0.74
0.67
0.69
1.33
0.58
0.48
0.97
0.93
1.44
1.02
1.26
1.12
1.39
1.14
1.27
0.88
1.06
B-32 Draft Document
-------
C 10 •
'"egg-ovary
20 25
Median ratio: 1.00
R2:
F:
df:
0.59
53.92
38
< 0.001
Table B-3. Summary of muscle to egg-ovary conversion factors
Common name
bluegill
bluehead sucker
brook trout
common carp
cutthroat trout
dolly varden
flannelmouth sucker
green sunfish
mountain whitefish
northern pike
rainbow trout
razorback sucker
roundtail chub
smallmouth bass
white sucker
Scientific name
Lepomis macrochirus
Catostomus discobolus
Cyprinus carpio
Oncorhynchus clarkii
Catostomus latipinnis
Lepomis cyanellus
Oncorhynchus mykiss
Gila robusta
Micropterus dolomieu
Catostomus commersonii
Median ratio
1.38
1.48
1.09
1.14
1.81
1.26
1.08
1.21
5.80
1.88
1.92
1.12
2.04
1.19
1.00
B-33 Draft Document
-------
Muscle to whole-body correction factor
Cwhoie-body = Selenium concentration
/">
^muscle
Ratio
Black bullhead
Study
= Selenium concentration
r
muscle
r
whole-body
(Ameiurus melas)
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
15 -|
10 -
muscle
5 -
0 -
(
o
°GO °
o°o°
) 5 10
whole-body
in all tissues ((ig/g dw)
in muscle tissue
whole-body
5.30
4.80
5.50
4.90
9.60
7.60
7.30
6.60
8.60
2.00
only ((ig/g dw)
Cmuscie Ratio
3.40
3.90
4.30
4.70
5.70
7.40
7.50
7.80
7.80
9.20
0.64
0.81
0.78
0.96
0.59
0.97
1.03
1.18
0.91
4.60
Median ratio: 0.93
R2: 0.00
F: 0.03
df: 8
P: 0.973
15 Not used because P > 0.05.
Bluegill (Lepomis macrochirus)
Study
Doroshov et al. 1992
Doroshov et al. 1992
Doroshov et al. 1992
Doroshov et al. 1992
Hermanutz et al
Hermanutz et al
Hermanutz et al
1996
1996
1996
whole-bodv
1.60
5.50
9.30
19.30
1.50
18.10
1.90
Cmuscie Ratio
1.50
5.80
10.40
23.60
2.10
20.60
1.90
0.94
1.05
1.12
1.22
1.40
1.14
1.00
B-34 Draft Document
-------
Bluegill (Lepomis macrochirus)
Hermanutzetal. 1996 2.80 3.50
Hermanutzetal. 1996 12.30 17.60
Hermanutzetal. 1996 9.40 12.50
Hermanutzetal. 1996 1.50 2.30
Hermanutzetal. 1996 4.90 6.90
Hermanutzetal. 1996 21.00 44.90
Hermanutzetal. 1996 24.30 39.80
Hermanutzetal. 1996 2.70 3.40
Hermanutzetal. 1996 5.00 5.30
Hermanutzetal. 1996 9.50 12.50
Hermanutzetal. 1996 6.60 7.80
Hermanutzetal. 1996 1.80 3.20
Hermanutzetal. 1996 4.20 6.10
Hermanutzetal. 1996 10.30 18.70
Hermanutzetal. 1996 13.80 15.10
Osmundson et al. 2007 8.80 12.90
50 -
^muscle ^
O
X Median ratio:
R2:
F:
,,,
df:
-t 1 1
0 25 50
/-<
^whole-body
1.25
1.43
1.33
1.53
1.41
2.14
1.64
1.26
1.06
1.32
1.18
1.78
1.45
1.82
1.09
1.47
1.32
0.89
172.2
21
< 0.001
B-35 Draft Document
-------
Bluehead sucker (Catostomus discobolus)
Study
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
^whole-bodv ^muscle JVallO
1.30
2.00
2.10
2.20
2.40
3.90
5.60
1.50
2.30
2.50
2.70
3.10
5.20
8.60
1.15
1.15
1.19
1.23
1.29
1.33
1.54
10 n
5 -
10
^whole-
ratio: 1.23
R2:
F:
df:
P:
0.99
682.9
5
<0.001
body
Brown trout (Salmo trutta)
Study
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
^whole-bodv ^muscle JvJltlO
4.60
4.30
5.00
5.50
3.20
3.60
4.00
6.30
0.70
0.84
0.80
1.15
muscle
4 -
4
^whole-body
Median ratio: 0.82
R2: 0.78
F: 7.2
df: 2
P: 0.122
Not used because P > 0.05.
B-36 Draft Document
-------
Channel catfish (Ictaluruspunctatus)
Study
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
^whole-bodv ^muscle JvatlO
3.40
3.30
2.60
4.00
3.40
3.60
3.70
5.30
1.00
1.09
1.42
1.33
6 -
c , 3 •
^muscle
0 Median r
T ' ' 1NUU US^U U^V^CIU
036
/-<
^whole-body
atio: 1.21
R2: 0.49
F: 2.0
df: 2
P: 0.338
seP>0.05.
Common carp (Cyprinus carpio)
Study Cwi,0ie.boiiv Cmuscie
Osmundson et al. 2007 6.30 7
Osmundson et al. 2007 4.80 8
Osmundson et al. 2007 1 1 .70 20
Osmundson et al. 2007 23. 10 24
Osmundson et al. 2007 4.10 6
Ratio
80 1
20 1
00 1
20 1
60 1
24
71
71
05
61
30 n
20 -
10
Median ratio: 1.61
10 20
/-<
^whole-body
30
R2:
F:
df:
P:
0.85
17.6
3
0.017
B-37 Draft Document
-------
Flannelmouth sucker (Catostomus latipinnis)
Study
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
10 -i
C 5-1
^muscle |
o J
0
^whole-bodv ^muscle JVallO
2007
2007
2007
2007
2007
2007
2007
o
^
5 10
^whole body
3.0 3.6 1.20
2.6 3.8 1.46
3.1 4.1 1.32
3.1 4.6 1.48
3.5 5.2 1.49
4.4 6.2 1.41
4.5 7.3 1.62
Median ratio: 1.46
R2: 0.91
F: 50.1
df: 5
P: <0.001
Green sunfish (Lepomis cyanellus)
Study
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
Osmundson et al.
^whole-body ^muscle IvallO
2007
2007
2007
2007
2007
2007
2007
2007
2007
2007
2007
2007
2007
2007
2007
2007
2007
22.80 28.10 1.23
8.80 12.90 1.47
15.40 21.90 1.42
4.80 5.00 1.04
5.70 6.10 1.07
4.40 5.20 1.18
3.80 5.10 1.34
11.90 15.70 1.32
6.40 10.10 1.58
9.50 11.50 1.21
9.10 10.50 1.15
6.20 7.20 1.16
7.00 9.30 1.33
7.70 7.70 1.00
6.20 6.00 0.97
10.20 12.00 1.18
9.70 12.10 1.25
B-38 Draft Document
-------
Green sunfish (Lepomis cyanellus)
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
9.90
7.20
9.00
9.70
8.90
9.80
9.90
10.30
5.30
10.10
11.80
3.30
4.00
4.30
3.70
6.20
3.50
4.40
5.60
4.90
4.40
8.00
7.90
6.40
8.70
8.30
6.10
5.60
18.10
9.40
12.20
5.30
7.30
9.30
6.80
7.50
12.50
7.50
11.30
13.60
13.20
12.40
12.50
8.60
5.30
11.90
13.60
3.80
4.20
4.10
4.20
5.70
4.40
3.50
5.50
5.00
4.30
10.10
11.90
11.10
11.80
11.00
7.10
6.70
26.40
9.60
16.70
8.10
10.60
14.20
11.30
12.80
1.26
1.04
1.26
1.40
1.48
1.27
1.26
0.83
1.00
1.18
1.15
1.15
1.05
0.95
1.14
0.92
1.26
0.80
0.98
1.02
0.98
1.26
1.51
1.73
1.36
1.33
1.16
1.20
1.46
1.02
1.37
1.53
1.45
1.53
1.66
1.71
B-39 Draft Document
-------
Green sunfish (Lepomis cyanellus)
30 -
C , 15 •
*^ muscle
0 -
C
J/°
jr
&P*
15
/-<
^whole-body
Median ratio:
R2
F:
df:
P:
30
1.23
: 0.91
501.6
51
< 0.001
Roundtail chub (Gila robustd)
Study
Osmundson et
Osmundson et
Osmundson et
Osmundson et
Osmundson et
Osmundson et
Osmundson et
al. 2007
al. 2007
al. 2007
al. 2007
al. 2007
al. 2007
al. 2007
^whole-bodv ^muscle
4.10 4.30
5.30 5.00
6.40 6.20
6.80 6.90
5.50 7.00
6.60 7.30
8.40 9.80
Ratio
1
0
0
1
1
1
1
05
94
97
01
27
11
17
10 n
^muscle
Median ratio: 1.05
5
hole-body
10
R2:
F:
df:
P:
0.86
29.6
5
0.002
B-40 Draft Document
-------
Smallmouth bass (Micropterus dolomieu)
Study
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
12 -i
C(~\
muscle
o -
c
et al. 2007
et al. 2007
et al. 2007
et al. 2007
et al. 2007
et al. 2007
et al. 2007
whole-bodv
4.10
5.30
6.40
6.80
5.50
6.60
8.40
muscle
4.30
5.00
6.20
6.90
7.00
7.30
9.80
/ Median ratio:
o /
/Q
o
6
whole-body
12
R2
F:
df:
P:
Ratio
1.05
0.94
0.97
1.01
1.27
1.11
1.17
1.23
: 0.83
20.2
4
0.008
White sucker (Catostomus commersonii)
Study
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
Osmundson
et al. 2007
et al. 2007
et al. 2007
et al. 2007
et al. 2007
et al. 2007
et al. 2007
et al. 2007
et al. 2007
et al. 2007
et al. 2007
et al. 2007
et al. 2007
et al. 2007
et al. 2007
et al. 2007
et al. 2007
et al. 2007
whole-bodv
3.80
4.20
3.30
4.50
6.30
6.80
11.00
12.70
5.70
3.90
3.80
9.90
5.30
10.70
5.90
7.00
6.40
6.30
muscle
2.90
4.80
3.70
3.70
8.40
9.40
15.50
23.60
9.40
6.10
4.60
12.30
9.20
9.40
9.40
10.50
11.40
9.60
Ratio
0.76
1.14
1.12
0.82
1.33
1.38
1.41
1.86
1.65
1.56
1.21
1.24
1.74
0.88
1.59
1.50
1.78
1.52
B-41 Draft Document
-------
White sucker (Catostomus commersonii)
Study
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
Osmundson et al. 2007
whole-bodv
5.30
6.20
5.60
8.80
8.70
11.40
10.70
8.40
7.00
7.50
10.30
6.70
2.10
1.80
3.20
2.30
3.10
3.00
2.80
2.50
3.40
2.80
3.10
5.50
7.00
7.30
2.40
2.70
2.70
2.60
19.60
9.80
8.70
8.70
9.10
13.40
3.10
2.40
2.10
3.20
2.80
Cmuscie Ratio
9.30
9.80
10.50
11.10
12.10
12.80
16.00
12.10
9.00
10.60
12.60
11.60
2.80
2.50
4.30
3.50
4.30
3.10
3.60
3.00
4.10
3.60
5.60
6.30
9.10
8.50
3.00
4.40
3.20
1.60
28.10
12.10
11.80
12.60
12.30
18.00
2.80
3.20
3.10
4.30
3.40
1.75
1.58
1.88
1.26
1.39
1.12
1.50
1.44
1.29
1.41
1.22
1.73
1.33
1.39
1.34
1.52
1.39
1.03
1.29
1.20
1.21
1.29
1.81
1.15
1.30
1.16
1.25
1.63
1.19
0.62
1.43
1.23
1.36
1.45
1.35
1.34
0.90
1.33
1.48
1.34
1.21
B-42 Draft Document
-------
White sucker (Catostomus commersonii)
Study
^whole-body ^muscle rvatlO
30 -,
20 -
^muscle
10 -
0 10 20
^whole-body
30
Median ratio: 1.34
R2: 0.91
F: 561.3
df: 57
P: < 0.001
Table B-4. Muscle to whole-body correction factor
Common name
Bluegill
Bluehead sucker
Common carp
Flannelmouth sucker
Green sunfish
Roundtail chub
Smallmouth bass
White sucker
Scientific name
Lepomis macrochirus
Catostomus discobolus
Cyprinus carpio
Catostomus latipinnis
Lepomis cyanellus
Gila robusta
Micropterus dolomieu
Catostomus commersonii
Median muscle to whole-body correction factor
Median ratio
1.32
1.23
1.61
1.46
1.23
1.05
1.23
1.34
1.27
B-43 Draft Document
-------
Table B-5. Final whole-body to egg-ovary conversion factors (CF)
Common name
Species
Bluegill
Bluehead sucker
Brook trout
Brown trout
Common carp
Cutthroat trout
Dolly varden
Flannelmouth sucker
Green sunfish
Mountain whitefish
Northern pike
Rainbow trout
Razorback sucker
Roundtail chub
Smallmouth bass
White sucker
Median ratio
\\^ egg-ovary ^whole-body)
2.13
1.82
1.45
1.92
1.41
1.45
2.07
1.42
1.41
Median ratio
V ^egg-ovary ^muscle)
1.09
1.81
1.26
5.80
1.88
1.92
1.12
Muscle to
whole-body
correction
factor
1.27
1.27
1.27
1.27
1.27
1.27
1.27
Final CF
values
2.13
1.82
1.38
1.45
1.92
2.30
1.61
1.41
1.45
7.39
2.39
2.44
1.42
2.07
1.42
1.41
Genus
Catostomus
Esox
Lepomis
Micropterus
Oncorhynchus
1.41
2.39
1.79
1.42
2.37
Family
Catostomidae
Centrarchidae
Cyprinidae
Salmonidae
1.41
1.45
2.00
1.96
B-44 Draft Document
-------
Common name
Order
Perciformes
Median ratio
V ^egg-ovary ^whole-body)
Median ratio
\\^ egg-ovary ^muscle)
Muscle to
whole-body
correction
factor
Final CF
values
1.45
Class
Actinopterygii
1.71
Derivation of Trophic Transfer Function values
Methodology
Taxa specific trophic transfer functions (TTF) to quantify the degree of biomagnification across a given
trophic level were calculated from either physiological parameters measured in laboratory studies or from
field measurements of paired selenium concentrations in consumer species and their food. TTFs from
both approaches were used to calculate translated water concentrations; however, when TTF data of
similar quality are available from both approached, as was the case with bluegill, field-derived TTF data
are used.
Physiological data consisted of assimilation efficiencies (AE), measured as either a percentage or a
proportion, ingestion rates (IR), measured as grams of Se per grams of food consumed per day, and efflux
rate constant (ke), measured as I/day. All available data were collected for a particular species, and then
the TTF for that species was calculated using the equation:
TTF =
AE xIR
Where AE, IR, and Ke were estimated as the median value of all available data for that parameter for that
species.
B-45 Draft Document
-------
The majority of TTF were calculated using paired whole-body Se measurements from organisms
collected at the same site in the field. TTFs for trophic level 2 organisms were determined using the
equation:
rTL2
T12 _ ^tissue
~ £TL2
Where Cj^d equals the average Se concentration in particulate matter, defined as the average of Caigae,
Cdetntus, and CSediment. Of the three types of particulate matter potentially assumed by TL2 organisms (e.g.,
the majority of invertebrates), Csediment correlated relatively poorly to Q^fue, when compared to Caigae and
Cdetntus- In order to minimize potentially erroneous TTF calculations based solely on sediment Se
concentrations, while note completely discounting the importance of organic matter in sediments as a
potential food source, Csediment was included in CpartlCuiate calculations only when either Caigae or Cdetntus data
were also available.
TTFs for trophic level 3 organisms were determined using the equation:
7TFTL3 = tissue
Cfood
Where Cj^d equaled the average whole-body Se concentration in invertebrates collected at the same site
as their potential predator species. The majority of trophic level 3 organisms were fish species, but
damselflies and dragonflies of the order Odonata are also trophic level 3 organisms, and TTFTL3 values
were calculated for those species as well.
For all field derived data used to determine TTFs, EPA first confirmed a statistical relationship between
whole-body selenium concentrations for each species and its food using OLS linear regression. If the
regression resulted in a statistically significant (P<0.05) positive slope, EPA calculated the TTF as the
median ratio of the paired concentration data.
B-46 Draft Document
-------
TTF values from physiological coefficients
AE(%) =
TTF
Assimilation efficiency
= Ingestion rate
= Efflux rate constant
AExIR
Invertebrates:
Baltic macoma (Macoma balthica)
Physiological Parameters
AE (%)
ke (d'1) TTF Study
22.5
91.0
84.0
95.0
78.0
74.0
92.3
58.0
85.8
64.9
90.4
Median Values and TTF
84.0 0.27a
Luomaetal. 1992
Luomaetal. 1992
Luomaetal. 1992
Luomaetal. 1992
0.03 Reinfelder etal. 1997
0.03 Reinfelder etal. 1997
Schleckat et al. 2002
Schleckat et al. 2002
Schleckat et al. 2002
Schleckat et al. 2002
Schleckat et al. 2002
0.03 7.56
Value taken from Mtilus edulis
Short-necked clam (Ruditapes philippinarum)
Physiological Parameters
AE (%) IR(g g1 d'1)
TTF Study
70.0
52.0
Median Values and TTF
61.0 0.27a
0.013
0.013
0.013 12.67
Zhang etal. 1990
Zhang etal. 1990
' Value taken from Mytilus edulis
B-47 Draft Document
-------
Quahog (Mercenaria mercenaria)
Physiological Parameters
AE (%) IR(g g'1 d'1)
ke (d'1) TTF Study
100.1
92.0
Median Values and TTF
96.1 0.27a
0.01
0.01 25.93
Reinfelder and Fisher
1994
Reinfelder et al. 1997
' Value taken from Mytilus edulis
Eastern Oyster (Crassostrea virginica)
Physiological Parameters
AE (%) IR(g g1 d'1)
ke
TTF
Study
105.4
70.0
Median Values and TTF
87.7 0.27a
0.005
0.070
0.038
Okazaki and Panietz
1981
Reinfelder and Fisher
1994
Reinfelder et al. 1997
6.31
' Value taken from Mytilus edulis
Common mussel (Mytilus edulis)
Physiological Parameters
AE(%) IRfeg'd1) ke(d1) TTF
86.0 0.02
75.0 0.05
60.7
48.0
13.7
55.1
55.8
71.9
71.5
27.9
84.4
81.0
79.4
63.0 0.037
61.5 0.05
Study
Reinfelder etal. 1997
Reinfelder etal. 1997
Wang and Fisher 1996
Wang and Fisher 1996
Wang and Fisher 1996
Wang and Fisher 1996
Wang and Fisher 1996
Wang and Fisher 1996
Wang and Fisher 1996
Wang and Fisher 1996
Wang and Fisher 1996
Wang and Fisher 1996
Wang and Fisher 1996
Wang and Fisher 1996
Wang and Fisher 1996
B-48 Draft Document
-------
Physiological Parameters
AE (%) IR(g g'1 d'1) ke
69.0
81.0
82.0
72.0
78.0
76.0
71.0
33.9
27.5
0.27
Median Values and TTF
71.3 0.27
(d1)
0.027
0.022
0.020
0.018
0.055
0.065
0.058
0.022
0.026
0.019
0.026
TTF Study
Wang and Fisher 1996
Wang and Fisher 1997
Wang and Fisher 1997
Wang and Fisher 1997
Wangetal. 1995
Wangetal. 1995
Wangetal. 1995
Wangetal. 1996
Wangetal. 1996
Wangetal. 1996
Wangetal. 1996
Wangetal. 1996
Wangetal. 1996
Wangetal. 1996
7.30
Asian clam (Corbicula flumined)
Physiological Parameters
AE (%) IR(g g1 d'1) ke
55.0 0.05
Median Values and TTF
55.0 0.05
(d1)
0.006
0.006
TTF Study
Lee et al. 2006
4.58
Zebra mussel (Dreissena polymorphd)
Physiological Parameters
AE (%) IR(g g'1 d'1) ke
18.0
24.0
46.0
40.0
41.0
7.7
23.0
28.0
0.40
(d1)
TTF Study
Roditi and Fisher 1999
Roditi and Fisher 1999
Roditi and Fisher 1999
Roditi and Fisher 1999
Roditi and Fisher 1999
Roditi and Fisher 1999
Roditi and Fisher 1999
Roditi and Fisher 1999
Roditi and Fisher 1999
B-49 Draft Document
-------
0.026
Median Values and TTF
26.0 0.40 0.026
Roditi and Fisher 1999
4.00
Water flea (Daphnia magnd)
Physiological Parameters
AE (%) IR(g g1 d'1) ke (dj)
0.08
0.34
57.9
43.0
39.8
33.0
41.4
41.5
38.0
24.5
0.101
0.12
0.131
0.134
0.108
0.112
Median Values and TTF
40.6 0.21 0.12
TTF Study
Goulet et al. 2007
Goulet et al. 2007
Yu and Wang 2002b
Yu and Wang 2002b
Yu and Wang 2002b
Yu and Wang 2002b
Yu and Wang 2002b
Yu and Wang 2002b
Yu and Wang 2002b
Yu and Wang 2002b
Yu and Wang 2002b
Yu and Wang 2002b
Yu and Wang 2002b
Yu and Wang 2002b
Yu and Wang 2002b
Yu and Wang 2002b
0.74
Copepod (Temora longicornis)
Physiological Parameters
AE (%) IR(g g1 d'1) ke (dj)
55.0 0.42 0.115
Median Values and TTF
55.0 0.42 0.115
TTF Study
Wang and Fisher 1998
2.01
Copepod (Small, unidentified)
Physiological Parameters
AE (%) IR(g g'1 d'1) ke (d'1)
50.0 0.42 0.155
Median Values and TTF
50.0 0.42 0.155
TTF Study
Schlekatetal.2004
1.35
B-50 Draft Document
-------
Copepod (Large, unidentified)
Physiological Parameters
AE (%) IR(g g1 d'1) ke (dj)
52.0 0.42 0.155
Median Values and TTF
50.0 0.42 0.155
TTF Study
Schlekatetal.2004
1.41
Blackworm (Lumbriculus variegatus)
Physiological Parameters
AE (%) IR(g g1 d'1) ke (dj)
0.009
0.006
24.0 0.067 0.013
9.0 0.067 0.009
Median Values and TTF
16.5 0.067 0.0086
TTF Study
Riedel and Cole 2001
Riedel and Cole 2001
Riedel and Cole 2001
Riedel and Cole 2001
1.29
Mayfly (Centroptilum triangulifer)
Physiological Parameters
AE (%) IR(g g'1 d'1) ke (d'1)
38.0 0.72 0.25
40.0 0.72 0.19
Median Values and TTF
39.0 0.72 0.22
TTF Study
Riedel and Cole 2001
Riedel and Cole 2001
1.28
Vertebrates:
Bluegill (Lepomis macrochirusf
Physiological Parameters
AE (%) IR(g g'1 d'1) ke (d1)
34.0
22.0
24.0
36.0
30.0
32.0
43.0
40.0
TTF Study
Besseretal. 1993
Besseretal. 1993
Besseretal. 1993
Besseretal. 1993
Besseretal. 1993
Besseretal. 1993
Besseretal. 1993
Besseretal. 1993
B-51 Draft Document
-------
37.0 0.041
0.031
0.034
36.0 0.031
0.038
0.038
0.008
0.042
Median Values and TTF
35.0 0.025 0.036 1.156
a
Besseretal. 1993
Besseretal. 1993
Besseretal. 1993
Besseretal. 1993
Besseretal. 1993
Besseretal. 1993
Whitledge and Haywood
2000
Whitledge and Haywood
2000
3 Not used because of availability of acceptable field-based TTF data
Fathead Minnow (Pimephales promelas)
Physiological Parameters
AE(%) IRfeg'd1) ke(d1) TTF
50.0
0.050
0.029
0.019
0.3
0.014
0.013
0.016
0.012
0.026
0.018
0.025
Median Values and TTF
50.0 0.050 0.0185 1.35
Study
Presser and Luoma 2010
Bertram and Brooks 1986
Bertram and Brooks 1986
Bertram and Brooks 1986
Bertram and Brooks 1986
Bertram and Brooks 1986
Bertram and Brooks 1986
Bertram and Brooks 1986
Bertram and Brooks 1986
Bertram and Brooks 1986
Bertram and Brooks 1986
Bertram and Brooks 1986
Striped Bass (Morone saxatilis)
Physiological Parameters
AE(%) IRfeg'd1) ke(d1) TTF
33 0.17 0.09
42 0.5 0.08
0.12
Study
Baines et al. 2002
Baines et al. 2002
Buckel and Stoner 2004
B-52 Draft Document
-------
0.16
0.11
0.08
Median Values and TTF
37.5 0.335
Buckel and Stoner 2004
Buckel and Stoner 2004
Buckel and Stoner 2004
0.085 1.48
TTF values from field data
Invertebrates:
c
^ -inve
Selenium concentration in algae (mg/kg)
Selenium concentration in detritus (mg/kg)
Selenium concentration in sediment (mg/kg)
Selenium concentration in invertebrate tissue (mg/kg)
Ratio
rtVClttgC SC
('invert
('part
1C111U111 CU11CC111
.IclUUll 111 \Jc
uutuitac mcuciicu
V 3
/
Scuds (Amphipoda)
Study
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Lambing etal. 1994
Saikietal. 1993
Saikietal. 1993
Saikietal. 1993
Saikietal. 1993
Saikietal. 1993
Saikietal. 1993
Saikietal. 1993
Saikietal. 1993
Saikietal. 1993
Saikietal. 1993
Saikietal. 1993
Saikietal. 1993
Saikietal. 1993
Site
29
20
7
19
30
3
22
23
S46
ET6
ET6
GT5
GT5
GT4
GT4
SJR2
SJR2
SJR3
SJR3
SJR1
SJR1
ET7
c c
8.80
3.00
0.18
16.80
17.30
0.10
4.60
7.80
2.30
1.03
1.03
4.50
4.50
1.39
1.39
1.25
1.25
0.45
0.45
0.22
0.22
0.16
;t *^s(
1.15
1.15
14.95
14.95
8.40
8.40
5.00
5.00
1.25
1.25
0.50
0.50
0.76
p
;d ^part
15.40 12.10
41.00 22.00
2.80 1.49
1.20 9.00
47.30 32.30
0.30 0.20
44.00 24.30
10.80 9.30
2.30
1.09
1.09
9.73
9.73
4.90
4.90
3.13
3.13
0.85
0.85
0.36
0.36
0.46
P
*^ invert
18.40
11.40
2.90
4.30
22.50
2.30
7.60
11.30
3.20
0.44
0.86
4.60
3.30
3.40
3.70
3.80
2.80
1.50
1.10
0.89
1.30
1.10
Ratio
1.52
0.52
1.95
0.48
0.70
11.50
0.31
1.22
1.39
0.40
0.79
0.47
0.34
0.69
0.76
1.22
0.90
1.77
1.30
2.47
3.61
2.42
B-53 Draft Document
-------
Saikietal. 1993
ET7
0.16 0.76
0.46
1.10
2.42
-Inverts
10 20 30 40
partic.
Median ratio: 1.22
R2: 0.69
F: 46.9
df: 21
P: < 0.001
Earthworms and Leeches (Annelida)
Study
Lemly 1985
Lemly 1985
Lemly 1985
Site
Badin Lake
Belews Lake
High Rock Lake
C i C,i ,
\^JI|P *^det
8.20
62.70
8.25
CSed Cpart Cinvert Ratio
0.91
8.27
0.79
4.56
35.49
4.52
8.10
51.15
9.05
1.78
1.44
2.00
-inverts
60 -
50 -
40 -
30 -
20 -
10 -
0
0
10
20
^partic.
30
40
Median ratio: 1.78
R2:
F:
df:
P:
1.00
2426
1
< 0.001
B-54 Draft Document
-------
Midges (Chironomidae)
Study
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Grassoetal. 1995
Lambing etal. 1994
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki etal. 1993
Saiki etal. 1993
Saiki etal. 1993
Saiki etal. 1993
Saiki etal. 1993
Saiki etal. 1993
Saiki etal. 1993
Saiki etal. 1993
Saiki etal. 1993
Saiki etal. 1993
Saiki etal. 1993
Saiki etal. 1993
Saiki etal. 1993
Site (
29
19
30
3
22
27
12
23
17
S46
Kesterson
Pond 1 1
Kesterson
Pond 2
Kesterson
Pond 2
Kesterson
Pond8
San Luis
Drain
San Luis
Drain
Volta Pond
26
Volta Pond
26
Volta Pond
7
Volta Pond
7
ET6
ET6
GT5
GT5
GT4
GT4
SJR2
SJR2
SJR3
SJR3
SJR1
SJR1
ET7
ca,g
8.80
16.80
17.30
0.10
4.60
10.35
2.30
7.80
1.87
2.30
18.15
152.7
152.7
136.5
67.00
67.00
0.42
0.42
1.03
1.03
4.50
4.50
1.39
1.39
1.25
1.25
0.45
0.45
0.22
0.22
0.16
Cdet
47.95
44.65
44.65
92.00
275.0
275.0
1.01
1.01
1.39
1.39
1.15
1.15
14.95
14.95
8.40
8.40
5.00
5.00
1.25
1.25
0.50
0.50
0.76
CSed
15.40
1.20
47.30
0.30
44.00
6.50
0.30
10.80
0.40
8.56
34.82
34.82
6.05
79.90
79.90
0.29
0.29
0.39
0.39
^part
12.10
9.00
32.30
0.20
24.30
8.43
1.30
9.30
1.14
2.30
18.15
44.65
44.65
92.00
79.90
79.90
0.42
0.42
0.89
0.89
1.09
1.09
9.73
9.73
4.90
4.90
3.13
3.13
0.85
0.85
0.36
0.36
0.46
^invert
58.20
15.30
59.30
2.50
18.80
26.70
7.70
34.20
2.07
9.70
71.00
200.0
290.0
220.0
190.0
284.0
1.74
1.30
3.00
1.30
0.58
1.00
8.90
7.20
5.40
6.90
6.00
4.10
1.50
1.60
0.47
1.00
0.53
Ratio
4.81
1.70
1.84
12.50
0.77
3.17
5.92
3.68
1.82
4.22
3.91
4.48
6.49
2.39
2.38
3.55
4.18
3.13
3.37
1.46
0.53
0.92
0.92
0.74
1.10
1.41
1.92
1.31
1.77
1.89
1.31
2.78
1.16
B-55 Draft Document
-------
Saikietal. 1993
ET7
0.16 0.76
0.46
0.84
1.85
350 i
300 -
250 -
200 -
^-inverts ^Q .
100 -
0 o ^
^x^ o
_/^
50 - 0^ 0
LJt o
o
slope.
Cinvert Ratio
77.60 2
74.10 2
110.00 3
54.00 1
89.10 1
28.80 0
43.70 0
1.92
0.20
1.24
5
0.36
0.05 and negative
38
27
37
75
92
62
94
B-56 Draft Document
-------
Water boatmen (Corixidae)
Study
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Lambing etal. 1994
Rinellaetal. 1994
Rinellaetal. 1994
Rinellaetal. 1994
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki and Lowe 1987
Schuleretal. 1990
Schuleretal. 1990
Schuleretal. 1990
Schuleretal. 1990
Schuleretal. 1990
Schuleretal. 1990
Schuleretal. 1990
Schuleretal. 1990
Site
18
29
20
7
3
22
12
23
S46
G
A
Q
Kesterson
Pond 1 1
Kesterson
Pond 1 1
Kesterson
Pond8
Kesterson
PondS
Volta Pond
26
Volta Pond
26
Volta Pond
7
Volta Pond
7
Kesterson
Pond 1 1
Kesterson
Pond 11
Kesterson
Pond 1 1
Kesterson
Pond 2
Kesterson
Pond 2
Kesterson
Pond 7
Kesterson
Pond 7
Kesterson
Pond 7
Calg Cdet t
7.60
8.80
3.00
0.18
0.10
4.60
2.30
7.80
2.30
0.84
2.21
1.42
18.15 47.95
18.15 47.95
136.50 92.00
136.50 92.00
0.42 1.01
0.42 1.01
1.39
1.39
53.70
53.70
53.70
52.50
52.50
87.10
87.10
87.10
>-^
4.30
15.40
41.00
2.80
0.30
44.00
0.30
10.80
0.50
0.40
0.50
8.56
8.56
6.05
6.05
0.29
0.29
0.39
0.39
11.50
11.50
11.50
9.30
9.30
5.90
5.90
5.90
^part
5.95
12.10
22.00
1.49
0.20
24.30
1.30
9.30
2.30
0.67
1.31
0.96
18.15
18.15
92.00
92.00
0.42
0.42
0.89
0.89
32.60
32.60
32.60
30.90
30.90
46.50
46.50
46.50
^invert
8.40
29.40
11.00
4.20
4.20
9.90
7.30
15.50
3.40
1.38
2.98
2.00
24.00
16.00
20.00
24.00
2.15
0.87
1.76
1.53
15.90
64.60
15.10
20.00
10.00
23.00
30.90
6.46
Ratio
1.41
2.43
0.50
2.82
21.00
0.41
5.62
1.67
1.48
2.06
2.28
2.08
1.32
0.88
0.22
0.26
5.17
2.10
1.98
1.72
0.49
1.98
0.46
0.65
0.32
0.49
0.66
0.14
B-57 Draft Document
-------
Rinella and Schuler
1992
18
0.59
0.59
2.70
4.58
70 -1
60 -
50 -
40 -
^-inverts ^Q . o
20 - o I
10 -Ler^^
of
u ™
0
0
Median ratio: 1.48
0
o ^^
~^»"
o
•
50
^^^
-^^ 8
I
100
R2: 0.25
F: 9.17
df: 27
P: < 0.001
Crayfish (Astacidae)
Study
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Butler etal. 1993
Butler etal. 1993
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Site
29
19
30
22
27
SP2
SP2
AK
AK
DD
DD
HD1
HD1
HD2
HD2
ME2
ME2
ME4
ME4
ME3
ME3
NW
NW
SD
c , c,, ,
*^alg *^det
8.80
16.80
17.30
4.60
10.35
1.60
1.60
0.45
0.45
0.88
0.88
0.59
0.59
0.45
0.45
1.11
1.11
1.04
1.04
0.82
0.82
3.45
3.45
0.77
sed
15.40
1.20
47.30
44.00
6.50
0.50
0.50
0.20
0.20
0.70
0.70
0.20
0.20
1.10
1.10
0.50
0.50
0.40
0.40
1.60
1.60
0.50
-------
Crayfish (Astaddae)
Butler et al
Butler et al
Butler et al
Butler et al
Butler et al
Butler et al
Butler et al
Butler et al
Butler et al
Saiki et al.
Saiki et al.
Saiki et al.
Saiki et al.
Saiki et al.
Saiki et al.
Saiki et al.
Saiki et al.
Saiki et al.
Saiki et al.
Saiki et al.
Saiki et al.
Saiki et al.
Saiki et al.
C
^-inverts
. 1995
. 1995
. 1995
. 1997
. 1997
. 1997
. 1997
. 1997
. 1997
1993
1993
1993
1993
1993
1993
1993
1993
1993
1993
1993
1993
1993
1993
40 -
35 -
30 -
25 -
20 -
15 -
10 -
s -
n J
o
p
X
«&•<" g
•BtO c
mo
SD
YJ2
YJ2
CHK
MN2
MUD2
MUD2
TRH
TRH
ET6
ET6
GT5
GT5
GT4
GT4
SJR2
SJR2
SJR3
SJR3
SJR1
SJR1
ET7
ET7
o
/*"
,/"
0
0
0
1
0
1
1
1
1
1
1
4
4
1
1
1
1
77
31
31
19
79
30
30
25
25
03
03
50
50
39
39
25
25
0.45
0.45
0
0
0
0
/
^
'"'
O
22
22
16
16
O
'-
1
1
14
14
15
15
95
95
8.40
8.40
5
5
1
1
0
0
0
0
00
00
25
25
50
50
76
76
0.50 0.64
0.10 0.21
0.10 0.21
1.19
0.79
.30
.30
.25
.25
.09
.09
9.73
9.73
4.90
4.90
3.13
3.13
0.85
0.85
0.36
0.36
0.46
0.46
Median ratio:
R2:
F:
df:
P:
1.40
1.40
1.50
0.90
0.83
3.10
3.80
0.98
1.60
0.67
0.83
5.20
4.40
3.10
3.20
1.70
1.90
0.77
1.30
0.50
0.74
0.87
0.85
1.46
0.74
130.8
45
< 0.001
2
6
7
0
1
2
2
0
1
0
0
0
20
83
32
76
06
38
92
78
28
62
76
53
0.45
0
0
0
0
0
1
1
2
1
1
63
65
54
61
91
53
39
06
91
87
10
20
30
40
c
*~-
B-59 Draft Document
-------
True flies (Diptera)
Study
Site Ca,g C
Schuleretal. 1990 Kesterson 53.70
Pond 1 1
Schuleretal. 1990 Kesterson 53.70
Pond 11
Schuleretal. 1990 Kesterson 52.50
Pond 2
Schuleretal. 1990 Kesterson 52.50
Pond 2
Schuleretal. 1990 Kesterson 52.50
Pond 2
Schuleretal. 1990 Kesterson 87.10
Pond 7
Schuleretal. 1990 Kesterson 87.10
Pond 7
Schuleretal. 1990 Kesterson 87.10
140 -
120 -
100 -
SO -
C
Mom-is 5Q .
40 -
20 -
n ,
Pond 7
o
o
a- — -g
0
- c,,,
11
11
9
9
9
5
5
5
C
50
50
30
30
30
90
90
90
Median
part
32.60
32.60
30.90
30.90
30.90
46.50
46.50
46.50
ratio:
R2:
F:
df:
P:
Not used because P >
si one
^invert
126.00
85.10
117.00
93.30
105.00
95.50
97.70
102.00
2.81
0.07
0.46
6
0.65
0.05 and
Ratio
3.87
2.61
3.79
3.02
3.40
2.05
2.10
2.19
negative
40
60
B-60 Draft Document
-------
Mayflies (Ephemeroptera)
Study
Site
Rinella etal. 1994 A
Casey 2005
Casey 2005
Casey 2005
Casey 2005
Casey 2005
Casey 2005
14 -
12 -
10 -
8 •
c
^-'inverts g .
4 -
""* »
0 -
(
Deerlick Creek
Luscar Creek
Deerlick Creek
Luscar Creek
Deerlick Creek
Luscar Creek
o ^
C C C C
2.21 0.40
1.00 0.20
5.50 3.20 2.40
1.00 0.20
5.50 3.20 2.40
1.00 0.20
5.50 3.20 2.40
o Median ratio:
--""""t> R2: 0.58
0 F: 6.77
df: 5
,art Cjnvert Ratio
1.31 9.65
7.39
0.60 6.40 10.67
3.20 8.20
0.60 5.70
3.20 9.70
0.60 6.80 1
3.20 12.30
7.39
2.56
9.50
3.03
1.33
3.84
P: 0.04
Not used because of anomalously high
ratio. TTF from physiological data used
1 1 2
instead.
3 4
Snails (Gastropoda)
Study
Site (
Butler etal. 1995 WC
Butler etal. 1995 WC
Butler etal. 1995 WC
Butler etal. 1997 DCP1
Butler etal. 1997 MNP2
Butler etal. 1997 CHP
Butler etal. 1997 LCHP1
^alg »^det »^sed ^part ^invert RatlO
3.30 1.50
3.30 1.50
3.30 1.50
1.00 2.10
5.40 6.70
4.00 2.10
0.33 1.10
2.40 3.70
2.40 3.90
2.40 2.00
1.55 3.50
6.05 2.00
3.05 19.00
0.72 0.32
1.54
1.63
0.83
2.26
0.33
6.23
0.45
B-61 Draft Document
-------
zu -
15 -
CIHWJS I0 "
5 -
n -
o
o
024
0
6
Median ratio:
R2:
F:
df:
P:
Not used because P >
S
1.54
0.01
0.07
5
0.93
0.05.
Zooplankton
Study
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Lambing et al.
Saiki and Lowe
Saiki and Lowe
Saiki and Lowe
Saiki and Lowe
Saiki and Lowe
Saiki and Lowe
Site
29
20
7
19
3
27
12
23
1988 12
1987 Kesterson
Pond 11
1987 Kesterson
Pond 2
1987 Kesterson
Pond8
1987 VoltaPond26
1987 VoltaPond7
1987 Volta
Wasteway
Saiki etal. 1993 ET6
Saiki etal. 1993 ET6
Saiki etal. 1993 GT5
Saiki etal. 1993 GT5
Saiki etal. 1993 GT4
Saiki etal. 1993 GT4
Saiki etal. 1993 SJR2
Saiki etal. 1993 SJR2
Saiki etal. 1993 SJR3
Saiki etal. 1993 SJR3
c,,,
8.80
3.00
0.18
16.80
0.10
10.35
2.30
7.80
1.40
18.15
152.70
136.50
0.42
0.87
1.03
1.03
4.50
4.50
1.39
1.39
1.25
1.25
0.45
0.45
Cdet
47
44
95
65
92.00
1
1
2
1
1
14
14
8
8
5
5
1
1
01
39
03
15
15
95
95
40
40
00
00
25
25
CSed
15.40
41.00
2.80
1.20
0.30
6.50
0.30
10.80
0.30
8.56
34.82
6.05
0.29
0.39
0.24
Cpart
12
22
1
9
0
8
1
9
0
18
44
10
00
49
00
20
43
30
30
85
15
65
92.00
0
0
0
1
1
9
9
4
4
3
3
0
0
42
89
87
09
09
73
73
90
90
13
13
85
85
Cinvert
31
11
3
7
3
42
5
15
2
68
30
00
30
70
40
50
80
40
60
30
83.00
100
1
2
2
1
1
2
5
4
4
2
4
1
1
00
46
90
80
20
50
40
40
50
40
60
30
60
80
Ratio
2
0
2
0
17
5
4
1
3
3
1
1
3
3
3
1
1
0
0
0
0
0
1
1
2
59
50
22
86
00
04
46
66
06
76
86
09
51
26
21
10
38
25
56
92
90
83
38
89
12
B-62 Draft Document
-------
Zooplankton
Saikietal. 1993 SJR1 0.22 0.50 0.36
Saikietal. 1993 SJR1 0.22 0.50 0.36
Saikietal. 1993 ET7 0.16 0.76 0.46
Saikietal. 1993 ET7 0.16 0.76 0.46
140 -|
120 -
100 -
80 -
C
»- inverts 6Q .
40 -
20 -
n 1
---"'"'"' O
„ /""" Median ratio:
o ^
o ---^'
R2:
o „„-"-" F:
°/~ df:
»! o P.
1.40 3.89
1.30 3.61
0.63 1.38
1.40 3.08
1.89
0.76
85.7
27
< 0.001
50
"partic.
100
Ratio
Special case of Odonates (Damselflies and Dragonflies) consuming invertebrates
n = Number of invertebrate food species co-occurring with an Odonate species.
Cpart = Average selenium concentration in particulate material (mg/kg):
(Cal3+Cd3et+Csed)
Cfood = Median selenium concentration in all invertebrate tissues that co-occur
with an Odonate species (mg/kg)
C damsel = Selenium concentration in damselfly tissue (mg/kg)
Cdragon = Selenium concentration in dragonfly tissue (mg/kg)
_ Cfood
('part
cdamsel cdragon
('food ('food
B-63 Draft Document
-------
Co-occurring potential food species of damselflies and dragonflies (Odonata)
Study
Saiki and Lowe
Saiki and Lowe
Saiki and Lowe
Saiki and Lowe
Saiki and Lowe
Saiki and Lowe
Saiki and Lowe
Saiki and Lowe
Saiki and Lowe
Saiki and Lowe
1987
1987
1987
1987
1987
1987
1987
1987
1987
1987
Schuleretal. 1990
Schuleretal. 1990
Schuleretal. 1990
Schuleretal. 1990
Schuleretal. 1990
Schuleretal. 1990
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Grassoetal. 1995
Site
Kesterson
11
Kesterson
2
Kesterson
2
Kesterson
0
o
Kesterson
e
Pond
Pond
Pond
Pond
Pond
o
Volta Pond 26
Volta Pond 26
Volta Pond 7
Volta Pond 7
Volta
Wasteway
Kesterson
11
Kesterson
11
Kesterson
2
Kesterson
2
Kesterson
7
Kesterson
7
29
20
7
19
30
3
22
27
23
17
Pond
Pond
Pond
Pond
Pond
Pond
Co-occurs with: n
dragonflies
dragonflies
dragonflies
dragonflies
dragonflies
dragonflies
dragonflies
dragonflies
dragonflies
dragonflies
dragonflies
dragonflies
dragonflies
dragonflies
dragonflies
dragonflies
damselflies
damselflies
damselflies
damselflies
damselflies
damselflies
damselflies
damselflies
damselflies
damselflies
4
4
4
5
5
4
4
5
5
2
10
10
8
8
11
11
3
2
2
2
2
3
3
1
3
1
cpa
18
44
44
92
15
65
65
00
92.00
0
0
0
0
0
32
42
42
89
89
87
60
32.60
30.90
30
46
46
12
22
1
9
32
0
24
8
9
1
90
50
50
10
00
49
00
30
20
30
43
30
14
Cfood
47.5
206.5
206.5
120
120
.52
.52
.53
.53
.83
75.85
75.85
93.3
93.3
69.2
69.2
29.4
11.2
3.55
9.8
40.9
2.5
9.9
26.7
15.5
2.07
Ratio
2
4
4
1
1
3
3
1
1
2
2
2
3
3
1
62
62
62
30
30
65
65
72
72
10
33
33
02
02
49
1.49
2
0
2
1
1
12
0
3
1
1
43
51
39
09
27
50
41
17
67
82
B-64 Draft Document
-------
Co-occurring potential food species of damselflies and
250 -i
200 -
150 -
Cfood 1QO .
50 -
0 <
O
^^
O ^
Q^~^Q
9^-0
f^r CD
0 20 40 60 80 100
r*
*- particulate
dragonflies (Odonata)
Median ratio:
R2:
F:
df:
P:
2.21
0.54
28.7
24
< 0.001
Damselflies (Anisoptera)
Study Site
Birkner 1978 29
Birkner 1978 4
Birkner 1978 25
Birkner 1978 20
Birkner 1978 7
Birkner 1978 19
Birkner 1978 6
Birkner 1978 30
Birkner 1978 3
Birkner 1978 22
Birkner 1978 27
Birkner 1978 23
Birkner 1978 11
Grassoetal. 1995 17
Grassoetal. 1995 9
100
80
60
(^
*- damsel
40
20
n
/""
/"'''
,,-••'"
O ^-''* o
/^
o /""'
£b
dfj
Cp
food ^damsel
29.4
1.95
18.7
11.2
3.55
9.8
4.2
40.9
2.5
9.9
26.7
15.5
5.9
2.07
8.2
Median ratio:
R2:
F:
df:
P:
Ratio
55
1.8
21.9
18.7
4.4
28.4
11.1
53.3
3.1
15.8
45.1
18.4
7.7
1.75
6.98
2.88
0.89
104.4
13
<0.001
1.87
0.92
1.17
1.67
1.24
2.90
2.64
1.30
1.24
1.60
1.69
1.19
1.31
0.85
0.85
20
40
60
-'fond
B-65 Draft Document
-------
Dragonflies (Zygopterd)
Study
Mason et al. 2000
Mason et al. 2000
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki and Lowe 1987
Schuleretal. 1990
Schuleretal. 1990
Schuleretal. 1990
Schuleretal. 1990
Schuleretal. 1990
Schuleretal. 1990
Sorenson & Schwarzbach 1991
200 i
150 -
r 100 -
*- dragon
50 -
r i i
o° 3''
>-_
q,-^P
i
Site (
BK
HCRT
Kesterson Pond 1 1
Kesterson Pond 2
Kesterson Pond 2
Kesterson Pond 8
Kesterson Pond 8
Volta Pond 26
Volta Pond 26
Volta Pond 7
Volta Pond 7
Volta Wasteway
Kesterson Pond 1 1
Kesterson Pond 1 1
Kesterson Pond 2
Kesterson Pond 2
Kesterson Pond 7
Kesterson Pond 7
5
o
_,.-•" o
-1 p
^food ^dragon
1.845
4.305
47.5
206.5
206.5
120
120
1.52
1.52
1.53
1.53
1.83
75.85
75.85
93.3
93.3
69.2
69.2
0.42
Median ratio:
R2:
F:
df:
P:
Ratio
1.665
2.81
53
155
171
95.5
105
1.4
1.42
1.2
1.4
2.5
63.1
95.5
110
65
61.7
56.2
0.49
1.97
0.95
343.5
17
<0.001
0.90
0.65
1.12
0.75
0.83
0.80
0.88
0.92
0.93
0.78
0.92
1.37
0.83
1.26
1.18
0.70
0.89
0.81
1.17
100
200
300
B-66 Draft Document
-------
Vertebrates:
Selenium concentration in invertebrate tissue ((^g/g)
Average selenium concentration in the whole-body offish (|ig/g)
Rati
0_ cflsh
r
^invert
Black bullhead (Ameiurus melas)
Study
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Lemly 1985
Mueller et al.
Mueller et al.
Lemly 1985
Mueller et al.
Site
Sand
Sand
Sand
Sand
Sand
Creek at Colfax
Creek at Colfax
Creek at Colfax
Creek at Colfax
Creek at Colfax
Badin Lake
1991 Lake
CO
1991 Lake
CO
High
Meredith near Ordway,
Meredith near Ordway,
Rock Lake
1991 Pueblo Reservoir near Pueblo,
CO
Butler et al . 1 99 1 Sweitzer Lake
Lemly 1985
50 -
40 -
30 -
CM 2Q _
10 -
0 -
Belews Lake
^^
w
I 1
o
~*^""^ o
^^
1 1 1
c
*^ invert
2
2
2
2
2
5
Cfish
81
81
81
81
81
18
6.40
6.40
6
8
29
45
75
70
80
53
Median ratio:
R2:
F:
df:
P:
Ratio
1.95
2.37
2.73
3.21
3.96
4.19
9.20
9.70
5.26
7.40
39.00
28.11
0.91
0.79
38.1
10
< 0.001
0.70
0.84
0.97
1.14
1.41
0.81
1.44
1.52
0.78
0.85
1.31
0.62
10
20 30 40 50
c,
invert.
B-67 Draft Document
-------
Black crappie (Pomoxis nigromaculatus)
Study
Site
Butler et al. 1995 Totten Reservoir
Butler etal. 1995 Summit Reservoir
Peterson et al
Peterson et al
Mueller et al.
.1991 Ocean Lake, west side
.1991 Ocean Lake, west side
1991 Lake Meredith near
Ordway, CO
Lambing et al. 1994 Priest Butte Lakes near
Choteau
Lambing et al. 1994 Priest Butte Lakes near
Choteau
Lambing et al. 1994 Priest Butte Lakes near
Choteau
Lambing et al. 1994 Priest Butte Lakes near
Choteau
Lambing et al. 1994 Priest Butte Lakes near
Choteau
Lambing et al. 1994 Priest Butte Lakes near
70 1
60 -
50 -
c 40 "
•fish 30 .
20 -
10 -
0 -
Choteau
0
0
^invert
1
1
3
3
Cfish
07
85
83
83
6.40
14
14
14
15
15
15
00
00
00
00
00
00
J£ Median ratio:
,xx^
^^
jg^0
Ofjfe*'*'^
i > « i
R2.
F:
df:
P:
Ratio
2.50
1.70
4.20
6.32
13.00
39.00
41.00
47.00
40.00
57.00
63.00
2.67
0.92
97.9
9
< 0.001
2
0
1
1
2
2
2
3
2
3
4
35
92
10
65
03
79
93
36
67
80
20
s 10 15 20
r*
*~ invert.
B-68 Draft Document
-------
Blacknose dace (Rhinichthys atratulus)
Study
Mason et al. 2000
Mason et al. 2000
Mason et al. 2000
- -
- -
Qish l '
1 -
fi
0 1
Site
BK
BK
BK
0
0
o
1 2
^-invert.
^invert ^fish
1.43
1.43
1.43
Median ratio: 1.01
R2: 0.0
F: 0.0
df: 1
P: 1.0
2 Not used because P >
Ratio
1.13
1.45
1.74
0.05.
0.79
1.01
1.21
Bluegill (Lepomis macrochirus)
Study
Saikietal. 1993
Saikietal. 1993
Saikietal. 1993
Saikietal. 1993
Hermanutz et al. 1996
Saikietal. 1993
Saikietal. 1993
Butler etal. 1995
Hermanutz et al. 1996
Saikietal. 1993
Saikietal. 1993
Hermanutz et al. 1996
Saikietal. 1993
Saikietal. 1993
Hermanutz et al. 1996
Saikietal. 1993
Saikietal. 1993
Saikietal. 1993
Saikietal. 1993
Hermanutz et al. 1996
Site
ET6
ET6
ET7
ET7
MSO I
SJR1
SJR1
TT
MSO III
SJR3
SJR3
MSO II
SJR2
SJR2
MSO III
GT4
GT4
GT5
GT5
MSO II
P P
^invert ^fish
0.85
0.85
0.86
0.86
0.87
0.95
0.95
1.07
1.20
1.50
1.50
1.70
3.30
3.30
3.95
4.05
4.05
4.90
4.90
5.05
Ratio
1.40
2.20
1.20
1.20
1.55
0.87
1.40
2.30
1.83
1.90
2.00
1.55
2.70
3.30
4.21
4.30
4.50
5.00
6.40
3.86
1.66
2.60
1.40
1.40
1.78
0.92
1.48
2.16
1.52
1.27
1.33
0.91
0.82
1.00
1.06
1.06
1.11
1.02
1.31
0.76
B-69 Draft Document
-------
Bluegill (Lepomis macrochirus)
Study
Hermanutz et al. 1996
Hermanutz et al. 1996
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Mueller etal. 1991
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Hermanutz et al. 1996
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Hermanutz et al. 1996
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Hermanutz et al. 1996
Hermanutz et al. 1996
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Site
MSOII
MSO III
transect 3
transect 3
transect 3
transect 3
Rl
transect 3
transect 3
transect 3
transect 3
MSO III
transect 3
transect 3
transect 3
transect 3
transect 3
transect 3
transect 4
transect 4
transect 4
transect 4
transect 4
transect 4
MSOII
transect 3
transect 3
transect 3
transect 3
MSOII
MSOII
transect 3
transect 3
transect 4
transect 4
transect 3
transect 3
transect 3
transect 3
transect 4
transect 4
^invert ^fish
5.05
5.55
8.60
8.60
8.60
8.60
8.70
9.25
9.25
9.25
9.25
10.00
11.40
11.40
11.95
11.95
15.20
15.20
15.70
15.70
15.70
15.70
16.45
16.45
16.63
16.95
16.95
16.95
16.95
17.30
17.30
17.90
17.90
18.25
18.25
20.35
20.35
20.70
20.70
20.90
20.90
Ratio
4.88
13.77
7.64
7.64
11.90
14.30
5.20
7.64
9.05
16.70
19.00
10.32
30.32
34.50
21.28
28.60
38.73
48.80
15.16
16.70
20.20
21.28
13.10
13.63
24.29
16.70
33.38
38.10
48.54
16.76
20.99
33.38
44.00
16.69
20.20
50.07
83.30
27.27
41.70
27.27
39.30
0.97
2.48
0.89
0.89
1.38
1.66
0.60
0.83
0.98
1.81
2.05
1.03
2.66
3.03
1.78
2.39
2.55
3.21
0.97
1.06
1.29
1.36
0.80
0.83
1.46
0.99
1.97
2.25
2.86
0.97
1.21
1.86
2.46
0.91
1.11
2.46
4.09
1.32
2.01
1.30
1.88
B-70 Draft Document
-------
Bluegill (Lepomis macrochirus)
Study
Crutchfield 2000
Crutchfield 2000
Hermanutz et al. 1996
Hermanutz et al. 1996
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
250 -|
200 -
150 -
100 -
50 - jjfr
,, -Btffrw^ *V
Site
transect 4
transect 4
MSOI
MSOI
transect 3
transect 3
transect 3
transect 3
transect 3
transect 3
transect 4
transect 4
transect 4
transect 4
transect 4
transect 4
transect 4
transect 4
transect 4
transect 4
transect 4
transect 4
transect 4
transect 4
transect 4
transect 4
transect 4
transect 4
0
o
o
3D ^L,--0""""""'""']!
|jg-e-r 8
y "
^invert ^fish
20.90
20.90
21.19
21.19
21.80
21.80
21.80
21.80
23.40
23.40
25.40
25.40
30.00
30.00
30.70
30.70
33.20
33.20
33.25
33.25
38.55
38.55
43.90
43.90
48.90
48.90
49.30
49.30
Median ratio:
R2:
F:
df:
P:
Ratio
43.96
52.40
18.13
18.28
26.20
28.03
33.38
65.50
46.25
84.50
30.32
45.20
51.60
60.70
86.51
102.40
53.13
59.50
54.66
61.90
83.46
237.00
37.97
48.80
62.18
156.00
72.75
92.90
1.48
0.57
115.5
87
< 0.001
2.10
2.51
0.86
0.86
1.20
1.29
1.53
3.00
1.98
3.61
1.19
1.78
1.72
2.02
2.82
3.34
1.60
1.79
1.64
1.86
2.16
6.15
0.86
1.11
1.27
3.19
1.48
1.88
0 20 40
^-invert.
60
B-71 Draft Document
-------
Bluehead sucker (Catostomus discobolus)
Study
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Site Cjnvert Cf,sh
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
1995
1995
1995
1995
1995
1995
1995
1995
1993
1993
1993
1995
1995
1993
1995
1995
1993
1993
1993
1994
1994
1995
1995
1994
1997
1997
1997
1997
1997
1993
1995
1995
1995
1995
1995
1997
1997
1993
1993
1997
1997
AK 0.78
HD1 0.83
HD1 0.83
HD1 0.83
HD1 0.83
DD 0.86
DD 0.86
DD 0.86
Dl
Bl
Bl
ME2
ME2
B2
SD
SD
D2
D2
PI
COL1
RB3
YJ2
YJ2
.20
.25
.25
.25
.25
.35
.40
.40
.45
.45
.50
.50
.60
.65
.65
NFK3 2.00
MN2 2.20
MUD 2.30
MUD 2.30
CHK 2.40
CHK 2.40
Ul 2.45
SJ1 2.50
SJ1 2.50
SJ1 2.50
ME3 2.55
ME3 2.55
MN3 2.70
MN1 2.90
SP1 2.95
SP2 3.40
MUD2 3.45
MUD2 3.45
0
0
0
1
1
0
0
1
2
1
2
0
1
1
1
1
1
2
2
1
13
0
2
1
1
1
2
1
1
4
0
1
1
1
1
1
1
5
7
2
5
Ratio
94
83
86
20
40
64
88
30
80
90
20
83
30
80
50
80
60
30
20
60
00
96
80
40
20
80
30
20
60
80
94
20
20
70
80
50
40
10
10
50
20
1
1
1
1
1
0
1
1
2
1
1
0
1
1
1
1
1
1
1
1
8
0
1
0
0
0
1
0
0
1
0
0
0
0
0
0
0
1
2
0
1
21
01
04
45
70
74
02
51
33
52
76
66
04
33
07
29
10
59
47
07
13
58
70
70
55
78
00
50
67
96
38
48
48
67
71
56
48
73
09
72
51
B-72 Draft Document
-------
Bluehead sucker (Catostomus discobolus)
Study
Site
Butler etal. 1997 MUD2
Butler etal. 1993 F2
Butler etal. 1991 4
Butler etal. 1993 F2
Butler etal. 1994 BSW1
Butler etal. 1997 WBR
Butler etal. 1997 WBR
Butler etal. 1995 NW
Butler etal. 1995 NW
Butler etal. 1994 LZA1
Butler etal. 1994 RBI
Butler etal. 1994 GUN2
35 -I
30 -
25 -
r 20 -
*-ttsh |5 .
10 -
5 -
n -
0
o
o o _«— Tr-""""""
«£§-s — -""""""""""
AR*i
0 10 20
C-
^invert ^fish
3.45
3.90
3.90
4.80
5.00
5.05
5.05
5.10
5.10
19.00
21.00
28.00
Median ratio:
R2'
F:
df:
P
30
Ratio
5.60
10.00
1.80
0.94
33.00
1.80
2.80
7.20
9.30
9.00
22.00
3.60
1.04
0.16
9.6
51
< 0.001
1.62
2.56
0.46
0.20
6.60
0.36
0.55
1.41
1.82
0.47
1.05
0.13
Brook stickleback (Culaea inconstans)
Study
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Site
SWA1
SWA1
SWA1
SWA1
SWA1
SWA1
SWA1
SWA1
SWA1
SWA1
SWA1
SWA1
SWA1
Cp
invert ^fish
2.81
2.81
2.81
2.81
2.81
3.64
3.64
3.64
3.64
3.64
3.64
3.64
3.64
Ratio
4.40
4.59
4.66
5.00
5.21
3.69
4.16
4.21
4.62
4.78
4.98
5.06
6.28
1.57
1.64
1.66
1.78
1.86
1.02
1.14
1.16
1.27
1.31
1.37
1.39
1.73
B-73 Draft Document
-------
Brook stickleback (Culaea inconstans)
Lambing etal. 1994 S38
Lambing etal. 1994 S37
Lambing etal. 1994 S36
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
SW2-1
SW2-1
SW2-1
SW2-1
SW2-1
SW2-1
SW2-1
SW2-1
SW2-1
SW2-1
SWB
SWB
SW1
SW1
SW1
SW1
SW11
SW11
SW2-1
Lambing etal. 1994 S34
Lambing etal. 1994 Sll
Lambing etal. 1994 Sll
50 -
40 -
30 -
10 -
0 -
(
o
Q O
| -x^g
O^g""*
el o0 o
) 5 10 15 20
C
^invert.
4
5
6
6
6
6
6
6
6
6
6
6
6
7
7
7
7
7
7
70
30
30
60
60
60
60
60
60
60
60
60
60
06
06
82
82
82
82
8.41
8.41
9
14
14
14
14
00
50
50
Median ratio:
R2:
F:
df:
P:
17.00
6.10
5.30
21.14
23.21
23.64
25.89
27.71
32.97
34.54
37.05
39.26
43.38
15.74
17.15
9.96
10.38
10.58
11.98
6.36
6.45
21.09
35.00
22.00
26.00
1.69
0.27
13.3
36
< 0.001
3.62
1.15
0.84
3.21
3.52
3.58
3.93
4.20
5.00
5.24
5.62
5.95
6.58
2.23
2.43
1.27
1.33
1.35
1.53
0.76
0.77
2.31
2.50
1.52
1.79
B-74 Draft Document
-------
Brook trout (Salvelinusfontinalis)
Study
Hamilton and Buhl 2004
Mason et al. 2000
Mason et al. 2000
Mason et al. 2000
Mason et al. 2000
Mason et al. 2000
Mason et al. 2000
Butler etal. 1997
Hamilton and Buhl 2005
Hamilton and Buhl 2005
Hamilton and Buhl 2004
Hamilton and Buhl 2004
12 -|
10 -
8 -
Cflstl 6-
2 ~°J&%
fi
Site
use
BK
BK
BK
HCRT
HCRT
HCRT
MN1
LGC
UGC
DVC
use
o
9-^^"^
0 5 10
c
*- invert.
Brown bullhead (Ameiurus
Study
Rinella and Schuler 1992
Mason et al. 2000
Mason et al. 2000
Mason et al. 2000
nebulosus)
Site
HCRT
HCRT
HCRT
^invert *^fish
0.50
1.43
1.43
1.43
2.81
2.81
2.81
2.90
7.80
9.30
12.80
0.50
^
° Median ratio:
R2:
F:
df:
P:
15
Cr<
invert *^fish
1.20
2.81
2.81
2.81
Ratio
2.40
1.21
1.57
1.90
0.99
1.59
2.95
2.20
6.90
9.80
8.00
2.40
0.88
0.83
43.6
9
< 0.001
Ratio
1.90
0.22
1.23
1.83
4.80
0.84
1.10
1.33
0.35
0.57
1.05
0.76
0.88
1.05
0.63
4.80
1.58
0.08
0.44
0.65
B-75 Draft Document
-------
~s _
Qlsh 1 -
1 -
0 -
^"""""""-••s^p
o
) 1 2
r.
^invert.
Median ratio:
R2:
F:
df:
p.
Not used because P >
slope
0.55
0.27
0.73
2
0.58
0.05 and negative
Brown trout (Salmo trutta)
Study
Butler et al.
Butler et al.
Butler et al.
Butler et al.
Butler et al.
Butler et al.
Butler et al.
Butler et al.
Butler et al.
Butler et al.
Butler et al.
Butler et al.
1993
1993
1993
1993
1993
1993
1993
1993
1993
1993
1993
1993
Formation 2012
Butler et al.
Butler et al.
1993
1994
Formation 2012
Formation 2012
Butler et al.
Butler et al.
1993
1991
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Butler et al.
1993
Formation 2012
Butler et al.
1993
Site
LP2
LP2
LP2
LP3
LP3
Bl
B2
B2
B2
D2
D2
D2
SFTC-1
PI
NFK3
SFTC-1
SFTC-1
SP2
12
CC-75
CC-75
CC-350
CC-350
LP4
SFTC-1
SP2
Cr<
invert *^fish
1.00
1.00
1.00
1.12
1.12
1.25
1.35
1.35
1.35
1.45
1.45
1.45
1.63
1.95
2.00
2.42
2.49
2.75
2.80
3.11
3.11
3.16
3.16
3.20
3.21
3.40
Ratio
1.60
1.70
2.10
2.10
2.80
4.20
2.40
2.70
2.70
3.20
3.50
3.50
6.70
3.30
5.00
3.68
2.64
1.20
5.40
4.05
5.35
6.28
8.53
1.80
2.25
3.40
1.60
1.70
2.10
1.88
2.51
3.36
1.78
2.00
2.00
2.21
2.41
2.41
4.11
1.69
2.50
1.52
1.06
0.44
1.93
1.30
1.72
1.99
2.70
0.56
0.70
1.00
B-76 Draft Document
-------
Brown trout (Salmo trutta)
Butler etal. 1993
Butler etal. 1993
Butler etal. 1993
Butler etal. 1991
Butler etal. 1991
Formation 2012
Butler etal. 1993
Formation 2012
Formation 2012
Formation 2012
McDonald and Strosher 1998
Formation 2012
Formation 2012
Formation 2012
Butler etal. 1991
Butler etal. 1994
Butler etal. 1994
Butler etal. 1994
Butler etal. 1994
Formation 2012
Formation 2012
Formation 2012
Butler etal. 1991
Hamilton and Buhl 2005
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
R2
R2
R2
4
4
CC-75
ST2
CC-75
CC-75
CC-350
ER747
CC-150
CC-150
CC-150
10
SMF
SMF
SMF
SMF
CC-3A
CC-3A
CC-3A
3
CC
CC-150
DC-600
DC-600
DC-600
DC-600
DC-600
LSV-4
LSV-4
HS-3
HS-3
CC-350
CC-350
CC-1A
CC-1A
CC-1A
CC-1A
HS-3
CC-1A
CC-150
3.70
3.90
3.90
3.90
3.90
3.97
4.10
4.16
4.16
4.20
4.29
4.46
4.46
4.70
4.80
4.80
4.80
4.80
4.80
5.45
5.45
5.48
6.20
6.70
7.03
7.83
7.83
8.53
8.53
8.65
9.54
9.54
11.40
11.40
11.45
11.45
12.24
12.24
12.24
12.57
13.41
13.55
14.32
5.90
5.40
6.70
3.30
3.50
3.18
6.00
6.60
10.32
5.78
4.80
5.83
8.67
5.20
2.00
8.40
8.54
9.40
21.44
9.20
10.44
11.25
3.50
9.70
10.14
10.54
12.83
6.20
8.54
5.85
15.18
16.20
18.83
20.60
7.95
11.50
9.33
10.51
16.85
9.95
17.89
14.03
7.83
1.59
1.38
1.72
0.85
0.90
0.80
1.46
1.59
2.48
1.38
1.12
1.31
1.94
1.11
0.42
1.75
1.78
1.96
4.47
1.69
1.92
2.05
0.56
1.45
1.44
1.35
1.64
0.73
1.00
0.68
1.59
1.70
1.65
1.81
0.69
1.00
0.76
0.86
1.38
0.79
1.33
1.04
0.55
B-77 Draft Document
-------
Brown trout (Salmo trutta)
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Butler etal. 1994
Butler etal. 1994
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Butler etal. 1994
Butler etal. 1994
Formation 2012
100 -|
80 -
60 -
Cfwh 4Q _
20 -
f! -
5LAol|$|ir
^ppfpfflp^
CC-3A
CC-3A
HS
HS
HS
HCC1
HCC1
LSV-2C
LSV-2C
HS-3
LSV-2C
HS-3
LSV-2C
HS
HS
GUN2
GUN2
LSV-2C
0
8
r-iJLjt-— '
,9— -f^QPo
o
14
14
15
15
18
21
21
22
22
24
26
26
26
27
27
28
28
30
50
50
70
70
70
00
00
62
62
70
31
55
95
80
80
00
00
00
Median ratio:
R2:
F:
df:
P:
15.38
19.68
16.52
25.00
24.90
35.68
42.00
12.78
19.45
23.68
22.67
28.97
20.96
22.80
32.63
5.90
80.27
19.53
1.44
0.55
102.7
85
< 0.001
1.06
1.36
1.05
1.59
1.33
1.70
2.00
0.56
0.86
0.96
0.86
1.09
0.78
0.82
1.17
0.21
2.87
0.65
10 20 30
*--invert.
40
B-78 Draft Document
-------
Bullhead (Ameiurus sp.)
Study
Butler
Butler
Butler
Butler
et
et
et
et
al.
al.
al.
al.
1995
1993
1993
1994
Site
ME3
R2
R2
BSW1
^invert
2.
3
3
5
Cfish
.55
.70
.70
.00
3
3
4
4
Ratio
.00
.50
.00
.10
1
0
1
0
.18
.95
.08
.82
c
3 -
fish _,
Median ratio: 1.01
R2:
F:
df:
P:
0.77
6.58
2
0.13
Not used because P > 0.05
-invert.
B-79 Draft Document
-------
Channel catfish (Ictalurus punctatus)
Study
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1997
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
Butler etal. 1993
Butler etal. 1993
Butler etal. 1993
Butler etal. 1993
Butler etal. 1993
Butler etal. 1993
Butler etal. 1997
Mueller et al. 1991
Butler etal. 1991
Butler etal. 1991
Butler etal. 1991
Butler etal. 1991
Butler etal. 1991
Butler etal. 1991
Butler etal. 1991
Site
TT
SJ1
SJ1
MN4
18
18
18
18
18
18
18
18
18
18
18
18
18
18
18
LP4
LP4
R2
R2
R2
R2
MN5
Rl
7
7
7
7
7
7
7
^invert ^fish
1.07
2.50
2.50
2.65
3.10
3.10
3.10
3.10
3.10
3.10
3.10
3.10
3.10
3.10
3.10
3.10
3.10
3.10
3.10
3.20
3.20
3.70
3.70
3.90
3.90
8.60
8.70
29.80
29.80
29.80
29.80
29.80
29.80
29.80
Ratio
1.00
2.80
4.10
4.20
.40
.50
.60
.70
.70
.80
.80
.90
2.00
2.10
2.20
2.20
2.30
2.40
3.10
2.68
3.30
3.00
3.31
2.17
7.87
5.00
2.20
18.08
18.66
28.03
31.85
32.40
36.95
39.50
0.94
1.12
1.64
1.58
0.45
0.48
0.52
0.55
0.55
0.58
0.58
0.61
0.65
0.68
0.71
0.71
0.74
0.77
1.00
0.84
1.03
0.81
0.90
0.56
2.02
0.58
0.25
0.61
0.63
0.94
1.07
1.09
1.24
1.33
B-80 Draft Document
-------
Channel catfish (Ictaluruspunctatus)
fish
50 -
40 -
30 -
20 -
10 -
0
Median ratio: 0.73
0
10 20
r
""invert.
30
40
F:
df:
P:
0.89
250.6
32
< 0.001
B-81 Draft Document
-------
Common carp (Cyprinus carpio)
Study
Rinella and Schuler 1992
Butler etal. 1993
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Low and Mullins 1990
Rinella and Schuler 1992
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Garcia-Hernandez et al. 2000
Butler etal. 1994
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Butler etal. 1993
Peterson et al. 1991
Peterson et al. 1991
Peterson et al. 1991
Peterson et al. 1991
Peterson et al. 1991
Butler etal. 1993
GEI2013
GEI2013
GEI2013
GEI2013
Butler etal. 1991
Site
Malheur Lake
D2
ME4
ME4
ME4
7
Malheur Lake
SJ1
SJ1
ME3
ME3
MN1
MN1
MN1
NFK2
18
18
18
18
18
18
SW4-
SW4-
SW4-
SW4-
SW4-
SW4-
SW4-
R2
7
7
7
7
7
R2
SW88
SW88
SW88
SW88
9
^invert ^fish
1.20
1.45
1.55
1.55
1.55
1.60
2.05
2.50
2.50
2.55
2.55
2.70
2.70
2.70
3.00
3.10
3.10
3.10
3.10
3.10
3.10
3.10
3.33
3.33
3.33
3.33
3.33
3.33
3.33
3.70
3.83
3.83
3.83
3.83
3.83
3.90
3.96
3.96
3.96
3.96
4.10
Ratio
2.00
3.70
3.70
3.80
3.90
0.30
2.20
3.40
5.30
4.40
5.20
5.40
5.80
9.80
3.30
4.90
3.20
3.90
4.60
4.70
4.80
5.30
3.91
4.36
4.48
4.60
4.78
7.29
9.61
3.30
4.24
4.41
4.73
5.16
5.21
4.80
3.88
5.33
5.49
5.66
3.90
1.67
2.55
2.39
2.45
2.52
0.19
1.07
1.36
2.12
1.73
2.04
2.00
2.15
3.63
1.10
1.58
1.03
1.26
1.48
1.52
1.55
1.71
1.18
1.31
1.35
1.38
1.44
2.19
2.89
0.89
1.11
1.15
1.23
1.35
1.36
1.23
0.98
1.35
1.39
1.43
0.95
B-82 Draft Document
-------
Common carp (Cyprinus carpio)
Butler etal. 1993
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Butler etal. 1991
Butler etal. 1994
Lemly 1985
Low and Mullins 1990
Mueller et al. 1991
Mueller et al. 1991
Butler etal. 1991
Mueller etal. 1991
Mueller etal. 1991
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Lemly 1985
GEI2013
GEI2013
Butler etal. 1993
Grassoetal. 1995
Grassoetal. 1995
Grassoetal. 1995
May et al. 2008
May et al. 2008
GEI2013
R2
SW9
SW9
SW9
SW9
SW9
SW9
SW9
SW9
SW9
SW9
SW9
SW9
SW9
10
BSW1
Badin lake
5
A6
A3
3
R2
R2
SW2-
SW2-
SW2-
SW2-
SW2-
SW2-
SW2-
SW2-
SW2-
SW2-
High Rock Lake
SWB
SWB
F2
9
9
9
SSW
SSAU
SW11
4.30
4.45
4.45
4.45
4.45
4.45
4.45
4.45
4.45
4.45
4.45
4.45
4.45
4.45
4.80
5.00
5.18
5.60
5.60
6.00
6.20
6.40
6.40
6.60
6.60
6.60
6.60
6.60
6.60
6.60
6.60
6.60
6.60
6.75
7.06
7.06
7.50
7.59
7.59
7.59
7.60
8.35
8.41
5.00
3.64
3.70
3.77
3.80
3.90
4.14
4.26
4.41
4.50
4.53
4.69
5.61
6.13
10.30
12.00
6.50
1.20
3.40
6.50
2.20
14.00
14.40
22.96
24.27
25.09
26.73
26.74
28.74
29.73
31.74
36.81
41.57
5.03
12.50
15.61
5.80
4.70
4.93
5.51
10.40
7.59
3.56
1.16
0.82
0.83
0.85
0.85
0.88
0.93
0.96
0.99
1.01
1.02
1.05
1.26
1.38
2.15
2.40
1.26
0.21
0.61
1.08
0.35
2.19
2.25
3.48
3.68
3.80
4.05
4.05
4.36
4.51
4.81
5.58
6.30
0.75
1.77
2.21
0.77
0.62
0.65
0.73
1.37
0.91
0.42
B-83 Draft Document
-------
Common carp (Cyprinus carpio)
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Mueller et al. 1991
Butler etal. 1997
Mueller et al. 1991
May et al. 2008
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
May et al. 2008
May et al. 2008
May et al. 2008
Lambing etal. 1994
Lambing etal. 1994
May et al. 2008
Butler etal. 1994
May et al. 2008
Butler etal. 1994
Butler etal. 1991
Butler etal. 1991
Butler etal. 1991
Butler etal. 1991
Lemly 1985
SW11
SW11
SW11
SW11
SW11
SW11
SW11
SW11
SW11
SW11
A2
MN5
Rl
NSK
SW2-
SW2-
SW2-
SW2-
SW2-
SW2-
sso
NSCL
SSAL
S34
S34
KR
RBI
NSP
GUN2
7
7
7
7
Belews Lake
8.41
8.41
8.41
8.41
8.41
8.41
8.41
8.41
8.41
8.41
8.50
8.60
8.70
8.81
9.14
9.14
9.14
9.14
9.14
9.14
10.00
10.70
11.50
14.00
14.00
17.20
21.00
24.00
28.00
29.80
29.80
29.80
29.80
45.53
3.60
3.79
3.95
4.14
4.34
6.43
7.21
7.50
7.90
11.84
7.30
16.00
5.60
9.33
13.29
13.77
20.49
23.65
24.84
27.27
8.48
10.80
10.50
19.00
32.00
7.78
5.10
10.30
63.00
25.80
31.00
40.00
50.00
43.66
0.43
0.45
0.47
0.49
0.52
0.77
0.86
0.89
0.94
1.41
0.86
1.86
0.64
1.06
1.45
1.51
2.24
2.59
2.72
2.99
0.85
1.01
0.91
1.36
2.29
0.45
0.24
0.43
2.25
0.87
1.04
1.34
1.68
0.96
B-84 Draft Document
-------
Common carp (Cyprinus carpio)
70 i
60 -
50 -
40-
*- ash 30 .
20 -
10 -
0 -
o
l^^
O ^i*****^
8Q ^.^"""Q
D f^^"^1^
jHr o 0°
r*sp i— i i i
0 10 20 30 40
C
*- invert.
0 Median ratio:
R2:
'
50
F
df:
P:
1.29
0.44
89.7
16
< 0.001
Creek chub (Semotilus atromaculatus)
Study
Site
Mason et al. 2000 HCRT
Mason et al. 2000 HCRT
Mason et al. 2000 HCRT
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
SW4-1
SW4-1
SW4-1
SW4-1
SW4-1
SW4-1
SW4-1
LG1
LG1
LG1
LG1
LG1
LG1
LG1
LG1
LG1
LG1
LG1
LG1
LG1
LG1
LG1
LG1
p
^invert
2
2
2
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
Cr,sh
81
81
81
33
33
33
33
33
33
33
37
37
37
37
37
39
39
39
39
39
39
39
39
39
39
56
Ratio
0.49
1.18
1.97
4.65
4.96
5.52
6.11
6.31
6.53
6.67
3.41
3.58
3.75
3.78
4.10
3.23
3.72
3.74
3.78
3.89
4.03
4.12
5.11
5.21
5.34
3.28
0.18
0.42
0.70
1.40
1.49
1.66
1.84
1.90
1.96
2.01
1.01
1.06
1.11
1.12
1.22
0.95
1.10
1.10
1.12
1.15
1.19
1.22
1.51
1.54
1.58
0.92
B-85 Draft Document
-------
Creek chub (Semotilus atromaculatus)
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
LG1
LG1
LG1
LG1
LG1
LG1
LG1
LG1
LG1
CC1
CC1
CC1
CC1
CC1
CC1
CC1
CC1
CC1
CC1
CC1
CC1
CC1
CC1
CC1
CC1
CC1
CC1
CC1
CC1
CC1
CC1
CC1
CC1
CC1
3.56
3.56
3.56
3.56
3.56
3.56
3.56
3.56
3.56
3.76
3.76
3.76
3.76
3.76
4.69
4.69
4.69
4.69
4.69
4.69
4.69
4.69
4.69
4.69
5.86
5.86
5.86
5.86
5.86
5.86
5.86
5.86
5.86
5.86
3.37
3.82
3.86
4.02
4.16
4.49
4.53
4.63
4.77
5.43
5.57
6.51
6.71
7.12
3.99
4.06
4.08
4.25
4.44
4.48
4.50
4.72
5.24
5.44
4.98
5.39
5.77
6.39
6.43
6.50
6.57
7.42
7.42
7.47
0.95
1.07
1.09
1.13
1.17
1.26
1.27
1.30
1.34
1.44
1.48
1.73
1.78
1.89
0.85
0.87
0.87
0.91
0.95
0.96
0.96
1.01
1.12
1.16
0.85
0.92
0.99
1.09
1.10
1.11
1.12
1.27
1.27
1.28
B-86 Draft Document
-------
Creek chub
8 -,
6-
CT • 4 ~
"i _
0 -
0
(Semotilus atromaculatus)
Is
*Br^
Hi
SB
o
0
o
2 4
""invert.
O
jj
1
6
Median ratio: 1.12
R2: 0.30
F: 25.11
df: 58
1 P: < 0.001
8
Cutthroat trout (Oncorhynchus clarkii)
Study
Hamilton and Buhl 2004
McDonald and Strosher 1998
Hamilton and Buhl 2005
McDonald and Strosher 1998
Hamilton and Buhl 2005
Hamilton and Buhl 2004
Hamilton and Buhl 2005
McDonald and Strosher 1998
Hamilton and Buhl 2005
Hamilton and Buhl 2004
Hamilton and Buhl 2004
Hamilton and Buhl 2004
60
50
40
Cnsi, 30
20
'I
.x-''
1 I
Site
ShpC
ER745
SC
ER747
UAC
ACM
DC
ER746
BGS
DVC
UEMC
LEMC
^
^
1
Cjnvert Cf,sh Ratio
1.90 1.80
2.74 5.40
4.10 3.50
4.29 6.57
5.00 6.60
6.70 6.30
8.70 11.00
10.70 12.71
10.80 12.20
12.80 10.20
26.90 27.00
75.20 52.30
-O
Median ratio: 1.07
R2: 0.97
F: 325.2
df: 10
1 P: < 0.001
0.95
1.97
0.85
1.53
1.32
0.94
1.26
1.19
1.13
0.80
1.00
0.70
20
40
80
r
""invert.
B-87 Draft Document
-------
Fathead minnow (Pimephales promelas)
Study
Butler etal. 1997
Butler etal. 1997
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1993
Butler etal. 1993
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1997
Butler etal. 1997
Butler etal. 1997
Butler etal. 1997
Butler etal. 1995
Butler etal. 1995
Birkner 1978
Birkner 1978
Butler etal. 1997
Butler etal. 1997
Butler etal. 1997
Grassoetal. 1995
Grassoetal. 1995
Grassoetal. 1995
Grassoetal. 1995
Grassoetal. 1995
Grassoetal. 1995
Butler etal. 1993
Butler etal. 1995
Butler etal. 1995
Site
MNP1
MNP1
AK
AK
AK
HD1
HD1
HD1
DD
DD
DD
HD2
HD2
Dl
Dl
ME2
SD
SD
SD
ME4
ME4
TRH
TRH
TRH
TRH
YJ2
YJ2
1
4
TR25
TR25
TR25
10
10
10
17
17
17
Ul
ME3
ME3
^invert ^fish
0.70
0.70
0.78
0.78
0.78
0.83
0.83
0.83
0.86
0.86
0.86
0.98
0.98
1.20
1.20
1.25
1.40
1.40
1.40
1.55
1.55
1.60
1.60
1.60
1.60
1.65
1.65
1.75
1.80
1.80
1.80
1.80
1.85
1.85
1.85
1.91
1.91
1.91
2.45
2.55
2.55
Ratio
1.70
1.80
2.60
2.80
2.90
2.50
2.60
3.90
3.40
3.60
3.90
1.50
1.60
3.70
3.80
4.80
3.00
4.00
4.90
1.40
5.90
2.20
3.00
4.20
4.30
4.00
11.00
2.10
2.10
4.00
5.20
6.00
2.74
2.79
2.90
6.59
6.60
7.30
6.40
4.30
4.40
2.43
2.57
3.35
3.61
3.74
3.03
3.15
4.73
3.95
4.19
4.53
1.53
1.63
3.08
3.17
3.84
2.14
2.86
3.50
0.90
3.81
1.38
1.88
2.63
2.69
2.42
6.67
1.20
1.17
2.22
2.89
3.33
1.48
1.51
1.57
3.45
3.46
3.82
2.61
1.69
1.73
B-88 Draft Document
-------
Fathead minnow (Pimephales promelas)
Study
Butler etal. 1995
Butler etal. 1994
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Butler etal. 1993
Lambing etal. 1994
Butler etal. 1993
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Butler etal. 1993
Butler etal. 1995
Butler etal. 1997
Butler etal. 1997
Butler etal. 1997
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Site
ME3
AD
SWA1
SWA1
SWA1
SWA1
SWA1
SWA1
SWA1
SWA1
SWA1
SWA1
WSB2
S48
SP2
SW4-1
SW4-1
SW4-1
SW4-1
SW4-1
SW4-1
SW4-1
SW4-1
SW4-1
SW4-1
LG1
LG1
LG1
LG1
LG1
LG1
SP2
ME1
MUD2
MUD2
MUD2
LG1
LG1
LG1
LG1
LG1
^invert ^fish
2.55
2.70
2.81
2.81
2.81
2.81
2.81
2.81
2.81
2.81
2.81
2.81
3.00
3.05
3.15
3.33
3.33
3.33
3.33
3.33
3.33
3.33
3.33
3.33
3.33
3.39
3.39
3.39
3.39
3.39
3.39
3.40
3.40
3.45
3.45
3.45
3.56
3.56
3.56
3.56
3.56
Ratio
5.30
9.60
3.89
3.98
4.04
4.33
4.48
4.53
4.81
5.00
5.24
5.76
8.10
2.50
8.20
4.85
5.25
5.39
5.88
5.89
6.07
6.11
6.61
6.67
6.87
3.60
3.89
4.27
4.45
5.18
5.51
6.00
5.60
6.50
7.70
12.00
3.26
3.35
3.72
4.09
4.20
2.08
3.56
1.39
1.42
1.44
1.54
1.60
1.61
1.71
1.78
1.87
2.05
2.70
0.82
2.60
1.46
1.58
1.62
1.77
1.77
1.83
1.84
1.99
2.01
2.07
1.06
1.15
1.26
1.31
1.53
1.63
1.76
1.65
1.88
2.23
3.48
0.92
0.94
1.05
1.15
1.18
B-89 Draft Document
-------
Fathead minnow (Pimephales promelas)
Study
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Butler et al.
Butler et al.
Butler et al.
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Butler et al.
Butler et al.
Butler et al.
Butler et al.
Butler et al.
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Butler et al.
Butler et al.
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Butler et al.
Butler et al.
Butler et al.
1993
1993
1994
1993
1993
1994
1993
1993
1993
1994
1993
1993
1993
Site
LG1
LG1
LG1
LG1
LG1
WSB2
WSB2
CF1
SWA1
SWA1
SWA1
SWA1
SWA1
SWA1
SWA1
SWA1
SWA1
SWA1
SB2
R2
PSW1
SB2
SB2
CC1
CC1
CC1
CC1
CC1
R2
LSW1
SW88
SW88
SW88
SW88
SW88
SW88
SW88
SW88
Rl
Rl
ST2
^invert
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
4
4
4
Cfish
.56
.56
.56
.56
.56
.60
.60
.60
.64
.64
.64
.64
.64
.64
.64
.64
.64
.64
.65
.70
.70
.75
.75
.76
.76
.76
.76
.76
.90
.90
.96
.96
.96
.96
.96
.96
.96
.96
.00
.00
.10
4
4
5
5
5
4
10
7
3
3
4
4
4
4
4
4
5
5
9
6
22
5
8
3
5
7
8
9
6
73
4
4
5
5
5
5
6
6
11
11
7
Ratio
.81
.86
.05
.47
.56
.20
.00
.90
.62
.72
.07
.43
.52
.66
.68
.76
.45
.71
.90
.60
.00
.70
.60
.79
.23
.36
.69
.07
.60
.00
.73
.96
.13
.55
.56
.86
.07
.32
.00
.00
.60
1.35
1.37
1.42
1.54
1.56
1.17
2.78
2.19
1.00
1.02
1.12
1.22
1.24
1.28
1.29
1.31
1.50
1.57
2.71
1.78
5.95
1.52
2.29
1.01
1.39
1.96
2.31
2.41
1.69
18.72
1.20
1.25
1.30
1.40
1.41
1.48
1.53
1.60
2.75
2.75
1.85
B-90 Draft Document
-------
Fathead minnow (Pimephales promelas)
Study
Butler etal. 1993
Butler etal. 1997
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Butler etal. 1993
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Butler etal. 1993
Butler etal. 1991
Butler etal. 1994
Lemly 1985
Lambing etal. 1994
Lambing etal. 1994
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Butler etal. 1991
Lambing etal. 1994
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Site
ST2
MNP2
SW9
SW9
SW9
SW9
SW9
SW9
ST2
CC1
CC1
CC1
CC1
CC1
WSB2
10
TGC
Badin Lake
S39
S39
CC1
CC1
CC1
CC1
CC1
CC1
CC1
3
S46
SW1
SW1
SW1
SW1
SW1
SW1
SW1
SW1
SW1
SW2-1
SW2-1
SW2-1
^invert ^fish
4.10
4.40
4.45
4.45
4.45
4.45
4.45
4.45
4.50
4.69
4.69
4.69
4.69
4.69
4.75
4.80
4.90
5.18
5.85
5.85
5.86
5.86
5.86
5.86
5.86
5.86
5.86
6.20
6.20
6.54
6.54
6.54
6.54
6.54
6.54
6.54
6.54
6.54
6.60
6.60
6.60
Ratio
16.00
11.00
5.57
5.93
6.14
6.20
6.56
7.57
12.80
5.92
6.49
7.14
7.59
7.68
17.10
8.10
11.00
2.54
7.90
21.00
6.68
7.73
7.88
8.45
9.21
9.70
11.69
9.50
5.10
7.01
7.86
7.98
8.23
8.50
9.48
9.95
10.09
10.19
12.51
12.83
14.80
3.90
2.50
1.25
1.33
1.38
1.39
1.47
1.70
2.84
1.26
1.39
1.52
1.62
1.64
3.60
1.69
2.24
0.49
1.35
3.59
1.14
1.32
1.35
1.44
1.57
1.66
2.00
1.53
0.82
1.07
1.20
1.22
1.26
1.30
1.45
1.52
1.54
1.56
1.90
1.95
2.24
B-91 Draft Document
-------
Fathead minnow (Pimephales promelas)
Study
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Lemly 1985
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Butler etal. 1994
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Site
SW2-1
SW2-1
SW2-1
SW2-1
SW2-1
SW2-1
SW2-1
we
we
we
High Rock Lake
SWB
SWB
SWB
SWB
SWB
SWB
SWB
SWB
SWB
SWB
SWB
SWB
SWB
SWB
SWB
SWB
SWB
SWB
SWB
CRC
SW1
SW1
SW1
SW1
SW1
SW1
SW1
SW1
SW1
SW1
^invert ^fish
6.60
6.60
6.60
6.60
6.60
6.60
6.60
6.75
6.75
6.75
6.75
7.06
7.06
7.06
7.06
7.06
7.06
7.06
7.06
7.06
7.44
7.44
7.44
7.44
7.44
7.44
7.44
7.44
7.44
7.44
7.50
7.82
7.82
7.82
7.82
7.82
7.82
7.82
7.82
7.82
7.82
Ratio
16.70
17.21
18.27
20.13
20.66
26.75
30.48
18.40
22.90
26.40
3.21
7.38
8.49
8.61
8.72
9.02
9.11
9.30
9.53
9.80
9.36
9.46
9.78
9.87
10.66
10.97
11.22
12.25
12.43
12.46
20.40
8.45
8.88
9.11
9.15
9.41
9.82
11.07
11.15
11.23
13.76
2.53
2.61
2.77
3.05
3.13
4.06
4.62
2.73
3.39
3.91
0.48
1.05
1.20
1.22
1.24
1.28
1.29
1.32
1.35
1.39
1.26
1.27
1.32
1.33
1.43
1.48
1.51
1.65
1.67
1.68
2.72
1.08
1.14
1.16
1.17
1.20
1.26
1.42
1.43
1.44
1.76
B-92 Draft Document
-------
Fathead minnow (Pimephales promelas)
Study
Butler etal. 1994
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Butler etal. 1997
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Butler etal. 1997
Butler etal. 1997
Birkner 1978
Lambing etal. 1994
Lambing etal. 1994
Lambing etal. 1994
Birkner 1978
Butler etal. 1994
Butler etal. 1994
Birkner 1978
Lemly 1985
Butler etal. 1994
Site
IW
SW11
SW11
SW11
SW11
SW11
SW11
SW11
MN5
SW2-1
SW2-1
SW2-1
SW2-1
SW2-1
SW2-1
SW2-1
SW2-1
WCP
WCP
22
S34
Sll
Sll
23
GUN2
MKP
27
Belews Lake
OMD
^invert ^fish
8.35
8.41
8.41
8.41
8.41
8.41
8.41
8.41
8.60
9.14
9.14
9.14
9.14
9.14
9.14
9.14
9.14
9.70
9.70
11.30
14.00
14.50
14.50
15.50
28.00
32.00
34.60
45.53
73.00
Ratio
10.00
4.68
5.29
5.34
5.38
5.38
5.70
7.05
7.30
13.31
15.63
15.77
16.79
17.00
18.21
19.39
22.50
10.00
15.00
11.00
25.00
11.00
33.00
34.50
7.50
51.00
79.00
23.03
13.00
1.20
0.56
0.63
0.63
0.64
0.64
0.68
0.84
0.85
1.46
1.71
1.73
1.84
1.86
1.99
2.12
2.46
1.03
1.55
0.97
1.79
0.76
2.28
2.23
0.27
1.59
2.28
0.51
0.18
B-93 Draft Document
-------
Fathead minnow
Study
100 -
80 -
60 -
Co* 4Q _
£2
20 - dj
Ml
o ^*
0
(Pimephales promelas)
Site
o
o ^~*^~
i£^-^^o
p* o °
1 1 1 1
20 40 60 SO
^"invert.
^invert ^fish
Median ratio:
R2:
F:
df:
P:
Ratio
1.57
0.22
64.4
232
< 0.001
Flannelmouth sucker (Catostomus latipinnis)
Study
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1993
Butler etal. 1993
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1993
Butler etal. 1994
Butler etal. 1994
Butler etal. 1994
Butler etal. 1994
Butler etal. 1994
Butler etal. 1994
Butler etal. 1994
Butler etal. 1994
Site
AK
AK
AK
AK
HD1
HD2
HD2
HD2
HD2
LP3
LP3
ME2
ME2
ME2
ME2
PI
COL1
COL1
COL1
COL1
COL1
COL1
COL1
COL1
P P
^invert ^fish
0.78
0.78
0.78
0.78
0.83
0.98
0.98
0.98
0.98
1.12
1.12
1.25
1.25
1.25
1.25
1.50
1.50
1.50
1.50
1.50
1.50
1.50
1.50
1.50
Ratio
0.82
0.90
1.10
1.10
2.90
0.49
0.54
0.62
0.96
0.92
1.40
1.40
1.60
2.00
2.20
2.40
0.50
0.60
0.63
0.92
1.00
1.60
1.70
1.80
1.06
1.16
1.42
1.42
3.52
0.50
0.55
0.63
0.98
0.83
1.26
1.12
1.28
1.60
1.76
1.60
0.33
0.40
0.42
0.61
0.67
1.07
1.13
1.20
B-94 Draft Document
-------
Flannelmouth sucker (Catostomus latipinnis)
Study
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
1994
1994
1995
1995
1995
1995
1995
1994
1995
1995
1995
1995
1997
1997
1997
1993
1997
1997
1995
1995
1995
1995
1995
1995
1995
1995
1995
1995
1995
1997
1997
1995
1995
1995
1997
1997
1993
1993
1994
1994
1991
Site
COL1
COL1
ME4
ME4
ME4
ME4
ME4
RB3
MP
MP
YJ2
YJ2
MNQ
MNQ
MNQ
PI
MUD
MUD
SJ1
SJ1
SJ1
SJ1
SJ1
SJ1
ME3
ME3
ME3
ME3
ME3
MN4
MN4
MN1
MN1
MN1
MN3
MN3
LP4
LP4
PSW1
LSW1
4
^invert
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
Cfish
50
50
55
55
55
55
55
60
60
60
65
65
80
80
80
95
30
30
50
50
50
50
50
50
55
55
55
55
55
65
65
70
70
70
70
70
20
20
70
90
90
2
3
29
1
1
1
2
2
3
3
1
2
4
0
1
1
2
3
4
1
1
2
2
3
5
9
1
4
6
2
2
2
2
9
6
1
Ratio
90
90
30
50
90
40
00
00
20
40
60
40
10
20
50
50
70
10
61
10
50
20
19
20
70
70
10
40
60
10
60
70
80
50
30
60
40
60
40
70
70
1
1
0
0
1
1
1
18
0
0
0
1
1
1
1
0
1
1
0
0
0
0
1
1
0
0
0
0
1
1
3
0
1
2
0
0
0
0
2
1
0
27
27
84
97
23
55
94
13
75
88
97
45
17
78
94
77
17
78
24
44
60
88
27
68
67
67
82
94
41
92
62
63
78
41
85
96
75
81
54
72
44
B-95 Draft Document
-------
Flannelmouth sucker
Study
Butler etal. 1991
Butler etal. 1991
Butler etal. 1991
Butler etal. 1994
Butler etal. 1994
Butler etal. 1997
Butler etal. 1997
Butler etal. 1994
Butler etal. 1994
Butler etal. 1991
35 -|
30 - o
25 -
10 - 000 %_
(Catostomus latipinnis)
Site
9
9
10
BSW1
CRC
MN5
NW2
LZA1
RBI
7
0
°^^
0
0 10 20 30 40
r
'-invert.
^invert ^fish
4.10
4.10
4.80
5.00
7.50
8.60
11.40
19.00
21.00
29.80
Median ratio:
R2:
F:
df:
P:
Ratio
1.50
6.00
2.50
9.60
12.00
8.40
11.00
17.00
4.60
22.00
1.06
0.36
41.6
73
< 0.001
0.37
1.46
0.52
1.92
1.60
0.98
0.96
0.89
0.22
0.74
Gizzard shad (Dorosoma cepedianum)
Study
Mueller etal. 1991
Mueller etal. 1991
Mueller etal. 1991
Site
R2
Rl
Rl
c c
^invert ^fish
6.40
8.70
8.70
Ratio
14.30
7.50
11.00
2.23
0.86
1.26
B-96 Draft Document
-------
20 -
15 -
10 -
o
•v
o
10
Median ratio: 1.26
R2:
F:
df:
P:
0.74
2.78
1
0.39
'invert.
Not used because P > 0.05 and negative
slope.
Goldeye
Study
Roddy
Roddy
Roddy
Roddy
Roddy
Roddy
Roddy
Roddy
Roddy
Roddy
et
et
et
et
et
et
et
et
et
et
(Hiodon alosoides)
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
1991
1991
1991
1991
1991
1991
1991
1991
1991
1991
Site
18
18
18
18
18
18
18
18
18
18
invert
3.
3.
3.
3.
3.
3.
3.
3.
3.
3.
Cfish
10
10
10
10
10
10
10
10
10
10
2
2
2
2
2
2
2
3
3
4
Ratio
.00
.10
.20
.30
.40
.70
.90
.40
.60
.70
0
0
0
0
0
0
0
1
1
1
.65
.68
.71
.74
.77
.87
.94
.10
.16
.52
5 -,
4 -
-fish
1 -
0
o
e
Median ratio: 0.82
R2:
F:
df:
P:
0.0
0.0
8
1.0
-invert.
B-97 Draft Document
-------
Green sunfish (Lepomis cyanellus)
Study
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1997
Butler etal. 1997
Butler etal. 1995
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
GEI2013
GEI2013
GEI2013
Butler etal. 1997
Butler etal. 1997
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Butler etal. 1991
Lemly 1985
Butler etal. 1991
Lemly 1985
GEI2013
Butler etal. 1997
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Butler etal. 1994
Butler etal. 1991
Site
HD1
HD1
MP
TRH
TR25
ME3
SWA1
SWA1
SWA1
SWA1
SWA1
18
18
18
18
18
LG1
LG1
LG1
MUD2
MUD2
SW88
SW88
SW9
SW9
SW9
SW9
SW9
10
Badin Lake
3
High Rock Lake
SWB
CHI
SW11
SW11
SW11
SW11
SW11
LZA1
7
^invert ^fish
0.83
0.83
1.60
1.60
1.80
2.55
2.81
2.81
2.81
2.81
2.81
3.10
3.10
3.10
3.10
3.10
3.39
3.39
3.39
3.45
3.45
3.96
3.96
4.45
4.45
4.45
4.45
4.45
4.80
5.18
6.20
6.75
7.44
7.50
8.41
8.41
8.41
8.41
8.41
19.00
29.80
Ratio
1.30
1.30
1.90
3.30
4.40
5.00
2.96
3.21
3.24
3.69
3.88
2.80
3.80
4.00
5.20
5.70
4.11
4.33
5.71
7.00
7.60
7.14
7.41
4.38
5.06
5.53
5.80
7.29
7.90
3.43
6.40
3.30
11.94
9.50
4.54
4.84
5.34
7.00
7.13
37.00
15.20
1.58
1.58
1.19
2.06
2.44
1.96
1.06
1.14
1.16
1.32
1.38
0.90
1.23
1.29
1.68
1.84
1.21
1.28
1.68
2.03
2.20
1.81
1.87
0.98
1.14
1.24
1.30
1.64
1.65
0.66
1.03
0.49
1.61
1.27
0.54
0.58
0.63
0.83
0.85
1.95
0.51
B-98 Draft Document
-------
Green sunfish (Lepomis cyanellus)
Butler etal. 1991
Lemly 1985
Belews Lake
29.80
45.53
25.10
21.96
0.84
0.48
40 -
30 -
Qish 20 H
10 -
0
0
10
o
c.
Median ratio: 1.27
20 30 40
50
R2:
F:
df:
P:
0.59
57.9
41
< 0.001
invert.
Iowa darter (Etheostoma exile)
Study
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Site
7
20
22
23
^invert ^fish
3.75
11.20
11.30
15.50
Ratio
2.10
36.30
23.00
41.90
0.56
3.24
2.04
2.70
fish
50 -
40 -
30 -
20 -
10 -
0
0
c,
Median ratio: 2.37
R2:
F:
df:
P:
0.90
17.3
2
0.055
10
15
20
Not used because P > 0.05
invert.
B-99 Draft Document
-------
Largemouth bass (Micropterus salmoides)
Saiki et al.
Saiki et al.
Saiki et al.
Saiki et al.
Saiki et al.
Saiki et al.
Rinella and
Saiki et al.
Saiki et al.
Butler et al
GEI2013
Study
1993
1993
1993
1993
1993
1993
Schiller 1992
1993
1993
1995
Garcia-Hernandez et al. 2000
Saiki et al.
Saiki et al.
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Saiki et al.
Saiki et al.
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Saiki et al.
Saiki et al.
GEI2013
GEI2013
GEI2013
GEI2013
1993
1993
1993
1993
1993
1993
Site
ET6
ET6
ET7
ET7
SJR1
SJR1
Malheur Lake
SJR3
SJR3
MP
SWA1
Cienga Wetland
SJR2
SJR2
SW4-
SW4-
SW4-
SW4-
SW4-
SW4-
LG1
SW88
SW88
SW88
SW88
SW88
SW88
GT4
GT4
SW9
SW9
SW9
SW9
SW9
SW9
GTS
GTS
SW11
SW11
SW11
SW11
^invert
0
0
0
0
0
0
1
1
1
1
2
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
8
8
8
8
85
85
86
86
95
95
20
50
50
60
81
00
30
30
33
33
33
33
33
33
39
96
96
96
96
96
96
05
05
45
45
45
45
45
45
90
90
41
41
41
41
Cfish
1
1
0
1
0
1
0
1
1
1
3
5
2
2
5
5
5
5
6
7
4
4
5
5
5
6
6
4
4
5
5
6
6
7
7
6
6
5
5
5
6
00
40
86
00
80
80
92
70
80
40
17
10
20
40
53
65
72
80
34
14
29
87
73
77
93
62
84
00
70
78
79
19
87
27
36
80
90
02
19
77
26
Ratio
1
1
1
1
0
1
0
1
1
0
1
1
0
0
1
1
1
1
1
2
1
1
1
1
1
1
1
0
1
1
1
1
1
1
1
1
1
0
0
0
0
18
66
00
16
85
90
77
13
20
88
13
70
67
73
66
70
72
74
91
15
27
23
45
46
50
67
73
99
16
30
30
39
54
63
65
39
41
60
62
69
74
B-100 Draft Document
-------
Largemouth bass (Micropterus salmoides)
Study
GEI2013
GEI2013
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
Crutchfield 2000
40 -I
30 -
C0sh 20 -
c
10 - -^^--J5
jf^ •
n IP* ^*
Site
SW11
SW11
transect 3
transect 3
transect 3
transect 3
transect 3
transect 3
transect 3
transect 3
transect 3
transect 3
transect 3
transect 3
transect 4
transect 4
transect 4
transect 4
transect 4
transect 4
transect 4
transect 4
transect 4
transect 4
transect 4
transect 4
0
o 8
^^*x"^ ri
U^8 8
r o
i i i i
^invert
8.41
8.41
8.60
8.60
8.60
8.60
9.25
9.25
9.25
9.25
11.40
11.40
11.95
11.95
15.70
15.70
15.70
15.70
16.45
16.45
18.25
18.25
20.90
20.90
20.90
20.90
Median
Cfish
6.48
7.22
8.92
9.50
10.32
11.40
10.70
10.96
14.00
14.20
23.70
26.12
19.62
22.30
13.50
14.78
29.69
34.90
10.70
11.20
14.78
17.20
20.20
24.34
28.60
30.83
ratio: 1.27
R2: 0.72
F: 164.3
df: 65
P: < 0.001
Ratio
0.77
0.86
1.04
1.10
1.20
1.33
1.16
1.18
1.51
1.54
2.08
2.29
1.64
1.87
0.86
0.94
1.89
2.22
0.65
0.68
0.81
0.94
0.97
1.16
1.37
1.48
10 15 20
-- invert.
B-101 Draft Document
-------
Longnose dace (Rhinichthys cataractae)
Study
Site
Lambing etal. 1994 S33
Mueller et al.
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Mueller et al.
Hamilton and
Hamilton and
10 -I
15 -
tfisl, 10 '
5 -
n -
1991 Al
LG1
LG1
LG1
LG1
LG1
LG1
LG1
LG1
LG1
LG1
LG1
LG1
1991 Tl
Buhl 2005 CC
Buhl 2005 BGS
o
c*^---
J^^^ °
08
o
0 5 10
r
*- invert.
^invert *^fish
2.40
2.70
3.37
3.37
3.37
3.37
3.37
3.39
3.39
3.56
3.56
3.56
3.56
3.56
5.40
6.70
10.80
Median ratio:
R2:
F:
df:
, P:
15 Not used because P >
Ratio
5.30
2.10
5.05
5.57
6.57
6.75
10.08
10.69
12.77
8.95
9.63
11.41
11.94
12.04
16.90
13.40
10.90
2.52
0.17
3.16
15
0.07
0.05
2.21
0.78
1.50
1.65
1.95
2.00
2.99
3.15
3.77
2.52
2.71
3.21
3.36
3.39
3.13
2.00
1.01
Longnose sucker (Catostomus catostomus)
Study
Site
Minnow 2007 FL17
Butler etal. 1994 NFK2
Butler etal. 1994 NFK2
Butler etal. 1994 NFK2
Butler etal. 1994 NFK2
Butler etal. 1994 NFK2
Cr<
invert *^fish
3.03
3.10
3.10
3.10
3.10
3.10
Ratio
1.40
2.10
2.50
2.70
2.80
2.90
0.46
0.68
0.81
0.87
0.90
0.94
B-102 Draft Document
-------
Longnose sucker (Catostomus catostomus)
Butler etal. 1994 NFK2 3.10 3.00
Butler etal. 1994 NFK2 3.10 3.20
Butler etal. 1994 NFK2 3.10 3.30
Butler etal. 1994 NFK2 3.10 3.40
Butler etal. 1994 NFK2 3.10 4.00
Mueller etal. 1991 Tl 5.40 3.60
Minnow 2007 FL17 21.22 7.90
10 -I
8 -
6 -
Cfch 4
.,
~
0 -
^-"e
^^-^^^ Median ratio: 0.90
^f^^"^ R2: 0.83
5 F: 54.7
0 df: 11
11111 P: <0.001
0 5 10 15 20 25
f
^invert.
0.97
1.03
1.06
1.10
1.29
0.67
0.37
Mosquitofish (Gambusia sp.)
Study Site Cinvert Cflsh Ratio
Lemly 1985 Badin Lake 5.18 5.44
Lemly 1985 High Rock Lake 6.75 5.75
Lemly 1985 Belews Lake 45.53 44.15
50 -i
40 -
30 -
c:asb _
10 •
n -
^x"'0
.^^ Median ratio: 0.97
.^
^^ R2: LOO
^^ F: 1326
oeT df: 1
P: 0.019
1.05
0.85
0.97
10
20 30
40
50
r
^invert.
B-103 Draft Document
-------
Mottled sculpin (Cottus bairdii)
Study
Hamilton and Buhl 2004
Butler etal. 1993
Butler etal. 1993
Butler etal. 1993
Butler etal. 1993
Butler etal. 1993
Butler etal. 1993
Butler etal. 1993
Butler etal. 1993
Hamilton and Buhl 2004
Butler etal. 1993
Butler etal. 1994
Butler etal. 1997
Butler etal. 1997
Butler etal. 1997
Lambing etal. 1994
Butler etal. 1991
Butler etal. 1997
Butler etal. 1997
Butler etal. 1994
Butler etal. 1991
Butler etal. 1991
Butler etal. 1991
Hamilton and Buhl 2005
Butler etal. 1991
Hamilton and Buhl 2004
Hamilton and Buhl 2005
Butler etal. 1993
Hamilton and Buhl 2004
Hamilton and Buhl 2005
Hamilton and Buhl 2005
Hamilton and Buhl 2004
Butler etal. 1994
Site
use
LP2
LP2
LP3
LP3
LP3
PI
PI
PI
ShpC
PI
NFK3
MN2
CHK
CHK
S33
12
MN1
MN1
NFK2
4
4
10
UAC
3
ACM
CC
F2
LBR
DC
BGS
DVC
HCC1
^invert ^fish
0.50
1.00
1.00
1.12
1.12
1.12
1.50
1.50
1.50
1.90
1.95
2.00
2.20
2.40
2.40
2.40
2.80
2.90
2.90
3.10
3.90
3.90
4.80
5.00
6.20
6.70
6.70
7.50
7.70
8.70
10.80
12.80
21.00
Ratio
5.30
2.20
3.10
3.90
4.20
4.90
5.10
6.40
6.70
4.10
7.30
5.80
2.60
3.10
4.40
3.70
4.20
3.20
3.40
6.40
2.60
4.40
5.00
6.20
6.50
8.30
8.20
9.90
5.20
12.00
12.30
8.80
5.60
10.60
2.20
3.10
3.50
3.77
4.39
3.40
4.27
4.47
2.16
3.74
2.90
1.18
1.29
1.83
1.54
1.50
1.10
1.17
2.06
0.67
1.13
1.04
1.24
1.05
1.24
1.22
1.32
0.68
1.38
1.14
0.69
0.27
B-104 Draft Document
-------
Mottled sculpin (Cottus bairdii)
14 -
12 -
10 -
8 -
6 -
4 -
2 -
0
o o
Median ratio: 1.38
R2:
F:
df:
P:
0.27
11.62
31
< 0.001
0
10 15
20
Mountain whitefish (Prosopium williamsoni)
Study
Low and Mullins 1990
McDonald and Strosher 1998
Minnow 2007
McDonald and Strosher 1998
Minnow 2007
Minnow 2007
Minnow 2007
Minnow 2007
Site
7
ER745
EL12
ER747
MI3
MI2
ELI
FO23
c c
^invert ^fis
1.60
2.74
4.01
4.29
6.21
6.69
7.08
10.00
h Ratio
1.40
4.17
6.60
4.93
9.12
10.16
9.12
10.20
0.88
1.52
1.65
1.15
1.47
1.52
1.29
1.02
-fish
14 -|
12 -
10 -
8 -
6 -
4 -
2 -
0
Median ratio: 1.38
0
10
15
R2:
F:
df:
P:
0.83
30.3
6
<0.001
•- invert.
B-105 Draft Document
-------
Northern pike (Esox
Study
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
3 -
3 -
~* „
p T -
*~ash -
1 -
1 -
A
0
lucius)
Site
PU
PU
TT
TT
TT
TT
TT
TT
o
O^""""'""^
O
1 1 2
f1
*-• invert.
^invert
0.61
0.61
1.07
1.07
1.07
1.07
1.07
1.07
Median
Cfish
0.93
.40
.80
.80
.90
.91
.97
2.68
ratio: 1.79
R2: 0.62
F: 9.63
df: 6
P: 0.013
Ratio
1.52
2.30
1.69
1.69
1.78
1.79
1.85
2.51
Northern plains killfish (Fundulus kansae)
Study
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Birkner 1978
Site
3
11
23
27
30
p
^ invert
3.10
5.65
15.50
34.60
45.05
Wish
7.70
5.00
23.10
31.90
57.40
Ratio
2.48
0.88
1.49
0.92
1.27
B-106 Draft Document
-------
70 -I
60 -
50 -
r 40 -
20 -
10 -
0.
o
^x-**X*^"^°
Median ratio: 1.27
R2: 0.93
F: 37.8
df: 3
P: 0.008
0 10 20 30 40 50
*- invert.
Rainbow trout (Oncorhynchus mykiss)
Study
Site
Butler etal. 1993 LP2
Butler etal. 1993 LP2
Butler etal. 1993 LP3
Butler etal. 1995 MP
Butler etal. 1995 MP
Butler etal. 1995 MP
Butler etal. 1995 MP
Butler etal. 1994 NFK3
Butler etal. 1997 MN2
Butler etal. 1997 MN2
Butler etal. 1997 CHK
Butler etal. 1997 CHK
Butler etal. 1997 CHK
Butler etal. 1997 CHK
Butler etal. 1997 CHK
Butler etal. 1997 MN3
Butler etal. 1997 MN3
Butler etal. 1997 MN3
Butler etal. 1997 MN1
Butler etal. 1997 MN1
Butler etal. 1997 MN1
Butler etal. 1994 NFK2
Butler etal. 1994 NFK2
Butler etal. 1993 F2
Butler etal. 1991 4
Casey 2005
Casey 2005
Deerlick Cr.
Deerlick Cr.
P P
^invert ^fis
1.00
1.00
1.12
1.60
1.60
1.60
1.60
2.00
2.20
2.20
2.40
2.40
2.40
2.40
2.40
2.70
2.70
2.70
2.90
2.90
2.90
3.10
3.10
3.90
3.90
4.45
4.45
Ratio
1.27
1.40
1.90
2.10
2.30
2.50
3.06
4.70
2.10
2.80
2.20
2.29
2.50
2.80
2.90
2.60
3.69
4.90
2.50
2.60
3.20
3.60
35.68
7.60
3.50
2.09
3.34
1.27
1.40
1.70
1.31
1.44
1.56
1.91
2.35
0.95
1.27
0.92
0.96
1.04
1.17
1.21
0.96
1.37
1.81
0.86
0.90
1.10
1.16
11.51
1.95
0.90
0.47
0.75
B-107 Draft Document
-------
Rainbow trout (Oncorhynchus mykiss)
Butler et al. 1
Butler et al. 1
Low and Mul
Casey 2005
Casey 2005
Butler et al. 1
Butler et al. 1
40 -i
30 -
10 -
0 -
(
993
997
lins 1990
994
994
o
)
(
F2
WBR
5
Luscar Cr.
Luscar Cr.
HCC1
GUN2
o
£--— -^^^
o
10 20 30
r-,
L'lnvm,
4.80 7.60
5.05 5.10
5.60 2.60
9.95 11.16
9.95 13.71
21.00 26.76
28.00 5.40
Median ratio: 1.19
R2: 0.16
F: 6.22
df: 32
P: 0.005
1.58
1.01
0.46
1.12
1.38
1.27
0.19
Red shiner (Cyprinella lutrensis)
Study
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1997
Butler etal. 1994
Butler etal. 1994
Butler etal. 1994
Lemly 1985
Mueller et al.
1991
Butler etal. 1991
Lemly 1985
May et al. 2008
Butler etal. 1994
May et al. 2008
Mueller etal. 1991
Butler etal. 1997
Site
ME4
YJ2
SJ1
ME3
ME3
MN4
AD
LW
LSW1
Badin Lake
A3
3
High Rock Lake
SSW
IW
SSAU
A2
MN5
P P
^invert ^fis
1.55
1.65
2.50
2.55
2.55
2.65
2.70
3.00
3.90
5.18
6.00
6.20
6.75
7.60
8.35
8.35
8.50
8.60
h Ratio
5.10
4.50
3.50
4.20
4.60
4.20
7.30
19.00
14.00
3.56
8.10
7.70
3.70
10.00
83.00
11.20
7.90
4.40
3.29
2.73
1.40
1.65
1.80
1.58
2.70
6.33
3.59
0.69
1.35
1.24
0.55
1.32
9.94
1.34
0.93
0.51
B-108 Draft Document
-------
Red shiner (Cyprinella lutrensis)
Study
May et al. 2008
May et al. 2008
May et al. 2008
May et al. 2008
May et al. 2008
May et al. 2008
May et al. 2008
Lemly 1985
Site
NSK
sso
NSCU
NSCL
SSAL
KR
NSP
Belews Lake
^invert ^fish
8.81
10.00
10.50
10.70
11.50
17.20
24.00
45.53
Ratio
5.81
7.16
7.24
7.36
9.00
7.03
8.62
30.92
0.66
0.72
0.69
0.69
0.78
0.41
0.36
0.68
c
fish
100 -|
so -
60 -
40 -
20 -
0
o o
Median ratio: 1.28
R2:
F:
df:
P:
0.06
1.59
24
0.224
0 10 20 30
r
v'invert.
40
50
Not used because P > 0.05
Redside shiner (Richardsonius balteatus)
Study
Site
invert
cfl;
ish
Ratio
Hamilton and Buhl 2004
Hamilton and Buhl 2004
Hamilton and Buhl 2005
ACM
LBR
BGS
6.70
7.70
10.80
6.00
2.70
13.20
0.90
0.35
1.22
B-109 Draft Document
-------
14 -
12 -
10 -
8 -
t fish g . o
4 - /
2 .
U-
•
0 5
»- -invert
o
/
/
/
/
r
0
1
10
Median ratio:
R2:
F:
df:
P
15 Not used because P >
0.90
0.73
2.68
1
0.397
0.05
Roundtail chub (Gila robusta)
Study
Butler etal. 1994
Butler etal. 1994
Butler etal. 1994
Butler etal. 1994
Butler etal. 1994
Butler etal. 1994
Butler etal. 1994
Butler etal. 1994
Butler etal. 1994
Butler etal. 1994
Butler etal. 1995
Butler etal. 1997
Butler etal. 1994
Butler etal. 1994
Butler etal. 1994
Butler etal. 1994
Butler etal. 1994
Butler etal. 1991
Butler etal. 1994
Butler etal. 1994
Butler etal. 1993
Butler etal. 1994
Butler etal. 1994
Butler etal. 1997
Butler etal. 1994
Butler etal. 1994
Site
COL1
COL1
COL1
COL1
COL1
COL1
COL1
COL1
COL1
RB3
MP
MUD
AD
LW
NFK2
PSW1
LSW1
10
TGC
BSW1
F2
CRC
IW
NW2
RBI
GUN2
c c
^invert ^fish
1.50
1.50
1.50
1.50
1.50
1.50
1.50
1.50
1.50
1.60
1.60
2.30
2.70
3.00
3.10
3.70
3.90
4.80
4.90
5.00
7.50
7.50
8.35
11.40
21.00
28.00
Ratio
2.20
2.50
2.70
3.30
3.70
4.10
5.10
5.30
26.00
5.40
4.20
4.60
7.10
4.50
6.10
7.70
5.80
1.90
10.00
8.10
7.30
19.00
8.50
6.90
5.90
6.80
1.47
1.67
1.80
2.20
2.47
2.73
3.40
3.53
17.33
3.38
2.63
2.00
2.63
1.50
1.97
2.08
1.49
0.40
2.04
1.62
0.97
2.53
1.02
0.61
0.28
0.24
B-110 Draft Document
-------
Roundtail chub (Gila robustd)
30 -
25 -
20 -
10 -
5 -
0 -
o
o
^9, -p
^jgu u o
o^ o
i i
0 10 20
r
^'invert.
Median ratio:
R2
F:
o
Nnt uspd '
30
df:
P:
secause P
1.98
0.01
0.18
24
0.834
>0.05
Sand shiner (Notropis stramineus)
Study
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Site
SW1
SW1
SW1
SW1
SW1
SW1
SW1
SW1
SW1
SW2-1
SW2-1
SW2-1
SW2-1
SW2-1
SWB
SWB
SWB
SWB
SW1
SW1
SW1
SW1
SW1
SW1
SW1
^invert
6
6
6
6
6
6
6
6
6
6
6
6
6
6
7
7
7
7
7
7
7
7
7
7
7
Cfish
54
54
54
54
54
54
54
54
54
60
60
60
60
60
06
06
06
06
82
82
82
82
82
82
82
Ratio
8.43
9.02
9.66
11.21
11.85
11.94
13.50
14.05
14.14
18.70
19.33
19.77
20.39
23.70
8.27
9.01
9.81
10.22
11.33
12.05
12.22
12.55
12.65
12.68
14.13
1.29
1.38
1.48
1.71
1.81
1.83
2.06
2.15
2.16
2.84
2.93
3.00
3.09
3.59
1.17
1.28
1.39
1.45
1.45
1.54
1.56
1.60
1.62
1.62
1.81
B-111 Draft Document
-------
Sand shiner (Notropis stramineus)
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
25 -i
20 -
15 -
cfish io _
5 -
A
0 2
SW1
SW1
SW1
SW2-1
SW2-1
SW2-1
SW2-1
SW2-1
o
i
HT
ii
i i i
468
r
^invert.
7.82
7.82
7.82
9.14
9.14
9.14
9.14
9.14
i
3
Median ratio:
R2:
F:
df:
P:
10
14.43
15.87
16.63
17.84
18.21
18.98
20.12
20.73
1.83
0.12
4.12
31
0.026
1.85
2.03
2.13
1.95
1.99
2.08
2.20
2.27
Sculpin (Cottoidea)
Study
Mason et al. 2000
Mason et al. 2000
Mason et al. 2000
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Site
BK
BK
BK
SFTC-1
SFTC-1
SFTC-1
CC-75
CC-75
CC-350
CC-350
SFTC-1
CC-75
CC-75
CC-75
CC-350
CC-150
CC-150
P P
^invert ^fish
1.43
1.43
1.43
1.63
2.42
2.49
3.11
3.11
3.16
3.16
3.21
3.97
4.16
4.16
4.20
4.46
4.46
Ratio
1.16
2.35
2.64
9.31
5.68
5.87
5.03
5.58
6.47
7.12
3.75
3.77
7.08
7.19
5.28
5.04
6.01
0.81
1.64
1.84
5.71
2.35
2.36
1.62
1.79
2.05
2.26
1.17
0.95
1.70
1.73
1.26
1.13
1.35
B-112 Draft Document
-------
Sculpin (Cottoidea)
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
Formation 2012
CC-150
CC-3A
CC-3A
CC-3A
CC-150
DC-600
DC-600
DC-600
DC-600
DC-600
LSV-4
LSV-4
HS-3
HS-3
CC-350
CC-350
CC-1A
CC-1A
CC-1A
CC-1A
HS-3
CC-1A
CC-150
CC-3A
HS
HS
HS
LSV-2C
LSV-2C
HS-3
LSV-2C
HS-3
LSV-2C
HS
HS
LSV-2C
4.70
5.45
5.45
5.48
7.03
7.83
7.83
8.53
8.53
8.65
9.54
9.54
11.40
11.40
11.45
11.45
12.24
12.24
12.24
12.57
13.41
13.55
14.32
14.50
15.70
15.70
18.70
22.62
22.62
24.70
26.31
26.55
26.95
27.80
27.80
30.00
5.14
11.65
14.45
11.47
10.73
7.96
8.62
7.87
8.50
7.63
18.28
20.01
18.57
21.85
9.53
10.03
8.34
9.94
17.47
7.78
26.63
12.63
7.35
20.20
23.23
23.25
10.95
11.38
17.47
23.93
18.85
23.68
20.32
35.93
41.30
25.95
1.09
2.14
2.65
2.09
1.53
1.02
1.10
0.92
1.00
0.88
1.92
2.10
1.63
1.92
0.83
0.88
0.68
0.81
1.43
0.62
1.99
0.93
0.51
1.39
1.48
1.48
0.59
0.50
0.77
0.97
0.72
0.89
0.75
1.29
1.49
0.87
B-113 Draft Document
-------
Sculpin (Cottoidea)
50
40
CBS. 20
10
0
• Q™&
H
0
Shorthead redhorse
Study
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
Roddy etal. 1991
4 -
3 -
CT _
Bsb -
1 -
n -
o
o
Q° 0 ^^-©tf"6
tftir ° °
o
1 1 R 1
10 20 30 40
*- invert.
(Moxostoma macrolepidotum)
Site
18
18
18
18
18
18
18
18
18
18
18
1
I
Median ratio: 1.29
R2: 0.63
F: 87.0
df: 51
P: <0.001
Cjnvert Cf,sh Ratio
3.10 2.80
3.10 2.90
3.10 2.90
3.10 3.10
3.10 3.30
3.10 3.40
3.10 3.50
3.10 3.60
3.10 3.70
3.10 3.80
3.10 3.80
Median ratio: 1.10
R2: 0.00
F: 0.00
df: 9
P: 1.0
0.90
0.94
0.94
1.00
1.06
1.10
1.13
1.16
1.19
1.23
1.23
Not used because P > 0.05
C..
B-114 Draft Document
-------
Smallmouth bass (Micropterus dolomieu)
Study
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1997
Mueller etal. 1991
Mueller etal. 1991
14 -1
12 -
10 -
r 8 -
*-fisli g .
4 - _^
2 - q§ —
IT F
024
C-
Site
MP
SU
SU
su
su
su
su
MNP3
Rl
Rl
O
-— —
_— -— ~"~ o
o
1 1 1
6 8 10
^invert ^fish
1.60
1.85
1.85
1.85
1.85
1.85
1.85
6.15
8.70
8.70
Median ratio:
R2:
F:
df:
P:
Not used because P >
Ratio
.90
.50
.50
.53
.55
.78
.91
12.00
2.90
4.10
0.83
0.26
2.81
8
0.119
0.05
1.19
0.81
0.81
0.83
0.84
0.96
1.03
1.95
0.33
0.47
Speckled dace (Rhinichthys osculus)
Study
Hamilton and Buhl 2004
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1993
Butler etal. 1993
Butler etal. 1993
Site
use
AK
AK
AK
HD1
HD1
HD1
DD
DD
DD
LP3
Dl
Dl
P P
^invert ^fish
0.50
0.78
0.78
0.78
0.83
0.83
0.83
0.86
0.86
0.86
1.12
1.20
1.20
Ratio
6.90
3.10
4.00
4.30
2.80
3.20
5.30
4.40
5.60
6.00
6.00
3.40
3.50
13.80
4.00
5.16
5.55
3.39
3.88
6.42
5.12
6.51
6.98
5.38
2.83
2.92
B-115 Draft Document
-------
Speckled dace (Rhinichthys osculus)
Study
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
1993
1993
1993
1995
1993
1993
1993
1993
1994
1994
1994
1994
1994
1994
1994
1994
1994
1994
1995
1995
1995
1997
1993
1994
1993
1993
1993
1997
1997
1993
1997
1997
1997
1997
1993
1993
1993
1993
1993
1993
1995
Site
Dl
Bl
Bl
ME2
B2
D2
D2
D2
COL1
COL1
COL1
COL1
COL1
COL1
COL1
COL1
COL1
RB3
YJ2
YJ2
YJ2
MNQ
PI
NFK3
SB1
SB1
SB1
MN2
MN2
ST1
MUD
MUD
CHK
CHK
Ul
Ul
Ul
Ul
Ul
Ul
SJ1
^invert
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
Cfish
.20
.25
.25
.25
.35
.45
.45
.45
.50
.50
.50
.50
.50
.50
.50
.50
.50
.60
.65
.65
.65
.80
.95
.00
.15
.15
.15
.20
.20
.25
.30
.30
.40
.40
.45
.45
.45
.45
.45
.45
.50
3
4
4
6
5
4
6
6
2
5
7
7
8
8
9
9
11
93
6
6
7
5
5
7
7
9
10
2
3
6
6
7
3
5
3
6
7
9
9
9
2
Ratio
.70
.40
.40
.10
.80
.90
.50
.80
.30
.00
.30
.40
.40
.60
.30
.60
.00
.00
.30
.50
.10
.90
.50
.10
.80
.50
.00
.70
.60
.80
.10
.20
.80
.20
.60
.90
.30
.20
.40
.80
.90
3
3
3
4
4
3
4
4
1
3
4
4
5
5
6
6
7
58
3
3
4
3
2
3
3
4
4
1
1
3
2
3
1
2
1
2
2
3
3
4
1
.08
.52
.52
.88
.30
.38
.48
.69
.53
.33
.87
.93
.60
.73
.20
.40
.33
.13
.82
.94
.30
.28
.82
.55
.63
.42
.65
.23
.64
.02
.65
.13
.58
.17
.47
.82
.98
.76
.84
.00
.16
B-116 Draft Document
-------
Speckled dace (Rhinichthys osculus)
Study
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
et
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
al.
1995
1995
1995
1995
1995
1997
1995
1997
1997
1993
1997
1993
1993
1993
1993
1993
1994
1994
1994
1994
1994
1994
1994
1994
1994
1994
1994
1994
1993
1993
1995
1993
1993
1994
1994
1993
1993
1993
1993
1994
1993
Site
SJ1
SJ1
ME3
ME3
ME3
MN4
MN1
MN3
MN3
SP2
MN1
SP1
SP1
SP1
WSB2
WSB2
LW
NFK2
NFK2
NFK2
NFK2
NFK2
NFK2
NFK2
NFK2
NFK2
NFK2
NFK2
LP4
ST2
ME1
WSB2
SB2
CF1
PSW1
SB2
SB2
R2
F2
LSW1
Rl
^invert
2
2
2
2
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
4
Cfish
.50
.50
.55
.55
.55
.65
.70
.70
.70
.75
.90
.95
.95
.95
.00
.00
.00
.10
.10
.10
.10
.10
.10
.10
.10
.10
.10
.10
.20
.35
.40
.60
.60
.60
.70
.75
.75
.90
.90
.90
.00
4
5
2
5
7
7
5
4
6
12
3
7
7
8
6
7
62
4
5
5
6
6
6
6
6
6
7
8
8
9
6
11
12
6
13
7
10
6
8
83
8
Ratio
.30
.10
.80
.50
.00
.90
.50
.30
.00
.00
.70
.00
.30
.90
.20
.60
.00
.80
.40
.70
.10
.20
.30
.40
.70
.90
.40
.70
.70
.30
.40
.70
.10
.10
.00
.80
.80
.00
.90
.00
.50
1
2
1
2
2
2
2
1
2
4
1
2
2
3
2
2
20
1
1
1
1
2
2
2
2
2
2
2
2
2
1
3
3
1
3
2
2
1
2
21
2
.72
.04
.10
.16
.75
.98
.04
.59
.22
.36
.28
.37
.47
.02
.07
.53
.67
.55
.74
.84
.97
.00
.03
.06
.16
.23
.39
.81
.72
.78
.88
.25
.36
.69
.51
.08
.88
.54
.28
.28
.13
B-117 Draft Document
-------
Speckled dace (Rhinichthys
Study
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
et
et
et
et
et
et
et
et
et
et
et
et
et
al. 1991
al. 1993
al. 1993
al. 1993
al. 1993
al. 1993
al. 1991
al. 1994
al. 1994
al. 1994
al. 1997
al. 1997
al. 1995
Hamilton and Buhl 2005
Butler
et
al. 1991
Hamilton and Buhl 2004
Butler
et
al. 1994
Hamilton and Buhl 2004
Butler
Butler
et
et
al. 1994
al. 1997
Hamilton and Buhl 2005
Butler
et
al. 1997
Hamilton and Buhl 2004
Butler
Butler
CB
et
et
sh
al. 1994
al. 1994
100 -|
80 -
60 -
40 -
20 -
n -
O
0
osculus)
Site
9
ST2
ST2
R2
ST2
WSB2
10
SMF
TGC
BSW1
WBR
WBR
NW
LiB
3
ACM
CRC
LBR
IW
MN5
SLC
NW2
DVC
LZA1
GUN2
^invert
4
4
4
4
4
4
4
4
4
5
5
5
5
5
6
6
7
7
8
8
9
11
12
19
28
Cfish
10
10
10
30
50
75
80
80
90
00
05
05
10
40
20
70
50
70
35
60
70
40
80
00
00
Median ratio:
o
^f^ito -c&Q-f
mKsff^
0
.____ *
*o o
R2:
F:
df:
P:
5
8
10
17
15
15
4
7
12
15
5
9
8
5
6
8
13
5
10
14
15
11
7
28
8
Ratio
.70
.50
.70
.10
.70
.60
.80
.80
.00
.00
.50
.70
.70
.80
.50
.50
.00
.60
.00
.00
.20
.00
.50
.00
.90
1
2
2
3
3
3
1
1
2
3
1
1
1
1
1
1
1
0
1
1
1
0
0
1
0
39
07
61
98
49
28
00
63
45
00
09
92
71
07
05
27
73
73
20
63
57
96
59
47
32
2.79
0.01
1.71
118
0.
185
10
r
*--invert.
20
30
Not used because P > 0.05
B-118 Draft Document
-------
Sucker (Catostomidae)
Study
Butler etal. 1995
Butler etal. 1995
Butler etal. 1993
Rinella and Schuler 1992
Butler etal. 1993
Butler etal. 1995
Butler etal. 1993
Butler etal. 1993
Butler etal. 1993
Butler etal. 1991
Butler etal. 1994
Butler etal. 1993
Butler etal. 1993
Butler etal. 1993
Butler etal. 1993
Butler etal. 1993
Butler etal. 1993
40 -
0
30 -
„
fch -
10 -
______ CT
n ^S5b8o
0 2
r
'"invert.
Site
HD2
HD2
Dl
Malheur Lake
B2
YJ2
PI
Ul
Ul
12
NFK2
SB2
R2
R2
ST2
WSB2
F2
o°___—- — —
5 " o
o
468
^invert ^fish
0.98
0.98
1.20
1.20
1.35
1.65
1.95
2.45
2.45
2.80
3.10
3.60
3.90
4.30
4.50
4.75
7.50
Median ratio:
R2:
F:
df:
P:
Not used because P >
Ratio
0.68
0.76
2.30
1.60
1.80
2.20
1.50
2.30
3.60
2.10
35.00
5.10
5.00
2.20
10.00
11.80
4.20
1.33
0.07
1.10
15
0.360
0.05
0.69
0.78
1.92
1.33
1.33
1.33
0.77
0.94
1.47
0.75
11.29
1.42
1.28
0.51
2.22
2.48
0.56
Sunfish (Centrarchidae)
Study
Welsh and Maughan 1994
Welsh and Maughan 1994
Welsh and Maughan 1994
Welsh and Maughan 1994
Welsh and Maughan 1994
Welsh and Maughan 1994
Welsh and Maughan 1994
Site
outfall drain
Pretty Water
Hart Mine Marsh
outfall drain
outfall drain
Pretty Water
Old Channel
P P
^invert ^fish
1.15
1.16
1.20
1.30
1.30
1.50
1.50
Ratio
2.30
1.80
2.40
2.10
2.80
1.60
2.00
2.00
1.56
2.00
1.62
2.15
1.07
1.33
B-119 Draft Document
-------
Sunfish (Centrarchidae)
Welsh and Maughan 1994
Welsh and Maughan 1994
Welsh and Maughan 1994
Welsh and Maughan 1994
Welsh and Maughan 1994
GEI2013
Pretty Water
Cibola Lake
Cibola Lake
Cibola Lake
Oxbow Lake
SW2-1
1.50
1.85
1.90
1.90
3.60
9.14
2.30
5.90
5.30
7.60
11.00
8.10
1.53
3.19
2.79
4.00
3.06
0.89
flsh
12 -
10 -
8 -
6 -
4 -
2 -
0
0246
^invert.
10
Median ratio: 2.00
R2:
F:
df:
P:
0.38
6.66
11
0.013
Tui chub (Gila bicolor)
Study
Sorenson & Schwarzbach 1991
Sorenson & Schwarzbach 1991
Rinella and Schuler 1992
Site
5
4
Harney Lake
Cp
invert ^fish
0.49
0.76
2.05
Ratio
1.20
1.00
3.10
2.45
1.32
1.51
4 -
3 -
3 -
i -
i -
o
0
Median ratio : 1.51
R2:
F:
df:
P-
0.94
15.9
1
0175
Not used because P > 0.05
C-
v'
B-120 Draft Document
-------
Walleye
Study
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
Butler
et
et
et
et
et
et
et
et
et
et
et
Peterson
Peterson
Peterson
Peterson
(Sander vitreus)
al
al
al
al
al
al
al
al
al
al
al
et
et
et
et
1995
1995
1995
1995
1995
1995
1995
1995
1995
1995
1995
al
al
al
al
Mueller et al.
fj
sh
10
8
6
4
7
n
.
-
.
-
1991
1991
1991
1991
1991
0
Site
PU
PU
PU
PU
PU
TT
TT
TT
TT
TT
TT
1
1
1
1
Rl
CL.
0
0
0
0
0
1
1
1
1
1
1
3
3
3
3
8
Cfish
61
61
61
61
61
07
07
07
07
07
07
83
83
83
83
70
Q Median ratio:
JL--
&--— -""^
-4f
|1
B
_ —•
.-—-•^
o
R2:
F:
df:
P:
0
2
2
2
2
4
4
6
8
2
Ratio
89
00
27
66
72
60
86
00
55
68
68
27
79
76
35
40
1
1
2
2
2
1
1
1
2
2
2
1
1
1
2
0
46
64
09
72
82
50
75
88
39
51
51
11
25
77
18
28
1.82
0.24
4.46
14
0.032
4
•>
'invert.
10
B-121 Draft Document
-------
Western mosquitofish (Gambusia a/finis)
Study
Saikietal. 1993
Saikietal. 1993
Saikietal. 1993
Saikietal. 1993
Saikietal. 1993
Saikietal. 1993
Saiki and Lowe 1987
Saiki and Lowe 1987
Saikietal. 1993
Saikietal. 1993
Saiki and Lowe 1987
Saiki and Lowe 1987
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Saikietal. 1993
Saikietal. 1993
GEI2013
Saikietal. 1993
Saikietal. 1993
Saikietal. 1993
Saikietal. 1993
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki and Lowe 1987
Saiki and Lowe 1987
Site
ET6
ET6
ET7
ET7
SJR1
SJR1
Volta Pond 26
Volta Pond 26
SJR3
SJR3
Volta Wasteway
Volta Wasteway
SWA1
SWA1
SWA1
SWA1
SWA1
SJR2
SJR2
SWA1
GT4
GT4
GT5
GT5
Kesterson Pond 1 1
Kesterson Pond 1 1
Kesterson Pond 8
Kesterson Pond 8
Kesterson Pond 2
Kesterson Pond 2
San Luis Drain
San Luis Drain
^invert ^fish
0.85
0.85
0.86
0.86
0.95
0.95
1.42
1.42
1.50
1.50
2.23
2.23
2.81
2.81
2.81
2.81
2.81
3.30
3.30
3.64
4.05
4.05
4.90
4.90
60.65
60.65
102.50
102.50
177.00
177.00
190.00
190.00
Ratio
1.00
1.30
0.90
1.00
0.95
1.30
1.24
1.28
1.70
2.00
1.35
1.36
3.01
3.49
3.66
3.89
4.27
2.20
4.50
2.91
4.50
4.90
11.00
16.00
104.00
130.00
164.00
223.00
224.00
247.00
149.00
332.00
1.18
1.54
1.05
1.16
1.01
1.38
0.87
0.90
1.13
1.33
0.61
0.61
1.07
1.24
1.30
1.39
1.52
0.67
1.36
0.80
1.11
1.21
2.24
3.27
1.71
2.14
1.60
2.18
1.27
1.40
0.78
1.75
B-122 Draft Document
-------
Western mosquitofish (Gambusia a/finis)
50 100 150 200
r1'
*-invert.
Median ratio: 1.25
R2:
F:
df:
P:
0.90
256.4
30
<0.001
Western cutthroat trout (Oncorhynchus clarkii lewisi)
Study
Site
Minnow 2007 BA6
Minnow 2007 AL4
Minnow 2007 MI5
Minnow 2007 EL 12
Minnow 2007 EL 14
Minnow 2007 FO9
Minnow 2007 MI3
Minnow 2007 MI2
Minnow 2007 ELI
Minnow 2007 LI8
Minnow 2007 FO10
Minnow 2007 HA7
Minnow 2007 CL11
70 -i
60 -
50 -
c • 40 "
30
20 -
10 -
n -
0
Q ^^^"^
^^
^X*^^
^^ 0
<%m
^invert
3
3
4
4
4
4
6
6
7
7
17
22
30
27
92
00
01
41
44
21
69
08
81
51
41
87
Cfish
6
4
5
7
4
7
5
5
4
9
45
21
57
Median
Ratio
98
44
12
42
52
80
65
16
82
36
94
10
27
ratio: 1.20
R2: 0.81
F: 47.6
df: 11
P: <0.001
2.13
1.13
1.28
1.85
1.02
1.76
0.91
0.77
0.68
1.20
2.62
0.94
1.86
10
20
30
40
^•invert.
B-123 Draft Document
-------
White sucker (Catostomus commersonii)
Study
Butler etal. 1993
Butler etal. 1993
Butler etal. 1993
Butler etal. 1993
Butler etal. 1993
Butler etal. 1993
Butler etal. 1995
Butler etal. 1995
Grassoetal. 1995
Grassoetal. 1995
Grassoetal. 1995
Grassoetal. 1995
Grassoetal. 1995
Grassoetal. 1995
Butler etal. 1994
Butler etal. 1993
Lambing etal. 1994
Mueller etal. 1991
GEI2013
GEI2013
GEI2013
Mason et al. 2000
Mason et al. 2000
Mason et al. 2000
Butler etal. 1993
Butler etal. 1993
Butler etal. 1993
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Butler etal. 1993
GEI2013
GEI2013
GEI2013
Site
LP3
Bl
D2
D2
PI
PI
MP
SU
17
17
17
17
17
17
NFK3
ST1
S33
Al
SWA1
SWA1
SWA1
HCRT
HCRT
HCRT
WSB2
SP2
LP4
SW4-
SW4-
SW4-
SW4-
SW4-
SW4-
SW4-
SW4-
SW4-
SW4-
ST2
LG1
LG1
LG1
^invert ^fish
1.12
1.25
1.45
1.45
1.50
1.50
1.60
1.85
1.91
1.91
1.91
1.91
1.91
1.91
2.00
2.25
2.40
2.70
2.81
2.81
2.81
2.81
2.81
2.81
3.00
3.15
3.20
3.33
3.33
3.33
3.33
3.33
3.33
3.33
3.33
3.33
3.33
3.35
3.37
3.37
3.37
Ratio
2.50
2.60
1.90
2.50
1.70
1.80
1.40
1.20
2.84
3.19
3.44
3.64
4.00
4.01
3.90
4.90
3.50
4.20
2.83
3.89
4.18
0.81
1.43
1.43
3.90
3.50
2.80
3.01
3.45
3.50
3.62
4.04
4.08
4.13
4.17
4.34
4.78
7.00
3.54
3.55
3.90
2.24
2.08
1.31
1.72
1.13
1.20
0.88
0.65
1.49
1.67
1.80
1.91
2.09
2.10
1.95
2.18
1.46
1.56
1.01
1.39
1.49
0.29
0.51
0.51
1.30
1.11
0.88
0.91
1.04
1.05
1.09
1.22
1.23
1.24
1.25
1.31
1.44
2.09
1.05
1.05
1.16
B-124 Draft Document
-------
White sucker (Catostomus commersonii)
Study
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Butler etal. 1993
Butler etal. 1993
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Butler etal. 1993
Butler etal. 1993
Butler etal. 1993
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Peterson et al. 1991
Peterson et al. 1991
Butler etal. 1993
Butler etal. 1991
GEI2013
GEI2013
Butler etal. 1993
Butler etal. 1993
GEI2013
GEI2013
GEI2013
GEI2013
Site
LG1
LG1
LG1
LG1
LG1
LG1
LG1
LG1
LG1
LG1
LG1
LG1
LG1
WSB2
WSB2
SWA1
SWA1
SWA1
SWA1
SWA1
SWA1
SB2
R2
SB2
CC1
CC1
CC1
CC1
CC1
7
7
R2
4
SW88
SW88
Rl
ST2
SW9
SW9
SW9
SW9
^invert ^fish
3.37
3.37
3.39
3.56
3.56
3.56
3.56
3.56
3.56
3.56
3.56
3.56
3.56
3.60
3.60
3.64
3.64
3.64
3.64
3.64
3.64
3.65
3.70
3.75
3.76
3.76
3.76
3.76
3.76
3.83
3.83
3.90
3.90
3.96
3.96
4.00
4.10
4.45
4.45
4.45
4.45
Ratio
3.95
4.48
3.00
2.72
2.80
2.89
2.99
3.04
3.08
3.13
3.18
3.25
3.27
4.30
6.30
2.83
3.39
3.47
3.55
3.63
3.75
4.30
4.20
4.80
5.99
6.56
7.21
7.42
7.62
3.30
4.64
5.40
5.30
4.63
4.75
9.50
8.30
4.07
4.18
4.19
4.20
1.17
1.33
0.88
0.77
0.79
0.81
0.84
0.86
0.87
0.88
0.89
0.91
0.92
1.19
1.75
0.78
0.93
0.95
0.98
1.00
1.03
1.18
1.14
1.28
1.59
1.74
1.92
1.97
2.03
0.86
1.21
1.38
1.36
1.17
1.20
2.38
2.02
0.91
0.94
0.94
0.94
B-125 Draft Document
-------
White sucker (Catostomus commersonii)
Study
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Butler etal. 1993
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Butler etal. 1991
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Butler etal. 1994
Site
SW9
SW9
CC1
CC1
CC1
CC1
CC1
CC1
CC1
CC1
CC1
CC1
F2
CC1
CC1
CC1
CC1
CC1
CC1
CC1
CC1
CC1
CC1
3
SWB
SWB
SWB
SWB
SWB
SWB
SWB
SWB
SWB
SWB
SWB
SWB
SWB
SWB
SWB
SWB
IW
^invert ^fish
4.45
4.45
4.69
4.69
4.69
4.69
4.69
4.69
4.69
4.69
4.69
4.69
4.80
5.86
5.86
5.86
5.86
5.86
5.86
5.86
5.86
5.86
5.86
6.20
7.06
7.06
7.06
7.06
7.06
7.06
7.06
7.06
7.44
7.44
7.44
7.44
7.44
7.44
7.44
7.44
8.35
Ratio
4.40
5.18
4.51
4.57
4.94
5.02
5.81
6.01
6.43
7.25
8.00
8.52
5.20
5.00
5.37
5.59
5.71
5.90
6.61
6.79
6.82
7.29
7.48
1.80
7.18
7.36
7.98
8.03
9.65
12.76
12.85
13.16
8.21
8.77
8.85
9.87
10.97
13.59
15.75
16.40
9.70
0.99
1.16
0.96
0.98
1.05
1.07
1.24
1.28
1.37
1.55
1.71
1.82
1.08
0.85
0.92
0.95
0.98
1.01
1.13
1.16
1.16
1.25
1.28
0.29
1.02
1.04
1.13
1.14
1.37
1.81
1.82
1.86
1.10
1.18
1.19
1.33
1.48
1.83
2.12
2.21
1.16
B-126 Draft Document
-------
White sucker (Catostomus commersonii)
Study
Mueller etal. 1991
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
Lambing etal. 1994
Lambing etal. 1994
Lambing etal. 1994
Butler etal. 1994
Butler etal. 1994
Butler etal. 1991
50 -|
40 -
30 -
10 - ^A
ft ^^^^
0
Site
Rl
SW2-
SW2-
SW2-
SW2-
SW2-
S34
S34
S34
HCC1
GUN2
7
0
^^*~^"
0 0
10 20 30
r
*" invert.
^invert ^fish
8.70
9.14
9.14
9.14
9.14
9.14
14.00
14.00
14.00
21.00
28.00
29.80
Median ratio:
R2:
F:
df:
P
40
Ratio
3.40
16.54
18.14
18.54
19.16
21.29
25.30
28.00
29.00
3.00
20.00
7.90
1.18
0.49
129.9
134
<0.001
0.39
1.81
1.99
2.03
2.10
2.33
1.81
2.00
2.07
0.14
0.71
0.27
Yellow perch (Perca flavescens)
Study
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Butler etal. 1995
Belize et al. 2006
Belize et al. 2006
Belize et al. 2006
Belize et al. 2006
Peterson et al. 1991
Belize et al. 2006
Belize et al. 2006
Site
PU
TT
TT
MP
MP
MP
Halfway
Geneva
Bethel
McFarlane
7
Long
Ramsey
^invert ^fish
0.61
1.07
1.07
1.60
1.60
1.60
1.74
2.29
2.61
3.79
3.83
4.42
4.97
Ratio
1.10
1.60
1.70
2.00
2.20
2.70
2.72
3.30
3.09
5.40
7.33
6.28
7.64
1.80
1.50
1.60
1.25
1.38
1.69
1.56
1.44
1.19
1.42
1.91
1.42
1.54
B-127 Draft Document
-------
Yellow perch (Perca flavescens)
Study
Site
Belize et al. 2006 Windy
Belize et al. 2006 Nelson
GEI2013
GEI2013
GEI2013
GEI2013
GEI2013
SW11
SW11
SW11
SW11
SW11
Lambing etal. 1994 S34
80 -
60 -
Cash 40 -
20 •
ft
U T
_-****"
^"^^
^oo^-®®®"^ ©
0 5 10
r
'"invert.
C- C
6.32
6.79
8.41
8.41
8.41
8.41
8.41
14.00
o
Median ratio:
.-— R2:
F:
df:
p.
15
Ratio
6.06
10.68
4.54
5.49
5.50
5.58
5.68
67.00
1.42
0.46
16.24
19
<0.001
0
1
0
0
0
0
0
4
96
57
54
65
65
66
68
79
B-128 Draft Document
-------
Table B-6. Final EPA-derived Trophic Transfer Function (TTF)Values
Common name
Species
Bluegill
Bluehead sucker
Brook trout
Brown trout
Common carp
Cutthroat tTrout
Dolly varden
Flannelmouth sucker
Green sunfish
Mountain whitefish
Northern pike
Rainbow trout
Razorback sucker
Roundtail chub
Smallmouth bass
White sucker
Genus
Catostomus
Esox
Lepomis
Micropterus
Oncorhynchus
Family
Catostomidae
Centrarchidae
Cyprinidae
Salmonidae
Order
Perciformes
Median ratio
\\^ egg-ovary ^whole-body)
2.18
1.82
1.45
1.92
1.41
1.45
2.07
1.42
1.41
Median ratio
\\^ egg-ovary ^muscle)
1.09
1.81
1.26
5.80
1.88
1.92
1.02
Muscle to whole-
body correction
factor
1.27
1.27
1.27
1.27
1.27
1.27
1.27
Final CF
values
2.13
1.82
1.38
1.45
1.92
2.30
1.61
1.41
1.45
7.39
2.39
2.44
1.30
2.07
1.42
1.41
1.41
2.39
1.79
1.42
2.37
1.41
1.45
2.00
1.96
1.45
B-129 Draft Document
-------
Common name
Class
Actinopterygii
Median ratio
\\- egg-ovary ^whole-body)
Median ratio
V ^egg-ovary ^muscle)
Muscle to whole-
body correction
factor
Final CF
values
1.71
B-130 Draft Document
-------
APPENDIX C: Summaries of Chronic Studies
Considered For Criteria Derivation
White sturgeon C-2
Sacramento splittail C-6
Fathead minnow C-9
Flannelmouth & razorback suckers C-15
Northern pike C-17
Chinook salmon C-20
Rainbow trout & brook trout C-25
Cutthroat trout C-44
Dolly varden C-57
Brown trout C-60
Desert pupfish C-70
Eastern and western mosquitofish C-87
Striped bass C-89
BluegillsunfishC-91
Largemouth bass C-133
See Appendix D for descriptions of other, less conclusive studies with:
Rainbow trout
Fathead minnow
Sacramento splittail
White sucker
See Appendix D for descriptions of invertebrate studies.
C-l Draft Document
-------
Tashjian, D.H., S.J. The, A. Sogomoyan and S.S.O. Hung. 2006. Bioaccumulation and chronic toxicity
of dietary L-selenomethionine in juvenile white sturgeon (Acipenser transmontanus}. Aquatic
Toxicol.79:401-409.
Test Organism:
Exposure Route:
Test Duration:
Study Design:
Effects Data:
White sturgeon (Acipenser transmontanus)
Dietary only
Seleno-L-methionine was added to an artificial diet consisting of vitamin-free
casein, wheat gluten, egg albumin, dextrin, vitamin mix, BTM-mineral mix,
cellulose, corn oil, cod liver oil, choline chloride and santoquin; the measured
dietary concentrations were 0.4, 9.6, 20.5, 41.7, 89.8, 191.1 mg Se/kg dw.
8 weeks
25 juvenile white sturgeon were placed in each of 24 90-L tanks. Treatments
were randomly assigned to the 24 tanks resulting in 4 replicates per dietary
treatment. Four fish from each tank were sampled after 0, 4 and 8 weeks for
weight, length, liver weight, condition factors, hepatosomatic indices, hemocrit,
histopathology, and selenium measurement in liver, kidney, muscle and gill
tissues. 8 fish after 0 and 8 weeks were sampled for whole body selenium
measurement.
Sturgeon survival did not differ significantly among treatment groups after the 8-
week exposure with a mean survival rate of 99 across all groups. Fish fed 41.7 to
191.1 mg Se/kg dw exhibited significant declines in body weight (see table). All
other endpoints measured were as sensitive or less sensitive to selenium in the
diet as body weight.
Mean (SE) white sturgeon moisture, lipid and whole body Se after 8-week exposure
Treatment
group
0.4
9.6
20.5
41.7
89.8
191.1
Moisture, % ww
76.8 (0.5) b
77.0 (0.7) b
76.8 (0.3) b
77.3 (0.5) b
78.5 (0.3) ab
80.0 (0.4) a
Lipid, % ww
9.5 (4) abc
9.5 (0.9) abc
10.1 (0.4) ab
9.6 (0.7) abc
7.6 (0.4) bed
6.1 (0.4) cd
muscle Se, mg/kg dw
8.2 (0.6) e
17.2 (0.7) d
22.9 (1.5) c
36.8 (1.8) b
52.9 (3 .2) a
54.8 (2.8) a
whole body Se, mg/kg dw
5. 2 (0.4) c
11.8 (0.9) b
14.7 (0.8) b
22.5 (1.4) a
34.4 (2.3) a
27.5 (4.4) a
C-2 Draft Document
-------
Mean (SE) white sturgeon body weight increase after 8-week exposure
Treatment
group
0.4
9.6
20.5
41.7
89.8
191.1
Body weight
increase (%)
282.9 (4.6) a
285.5 (9.9) a
277.7 (6.1) a
191.0 (12.6) b
106.5 (5.8) c
28.6 (3.6) d
muscle Se, mg/kg dw
8.2 (0.6) e
17.2 (0.7) d
22.9 (1.5) c
36.8 (1.8) b
52.9 (3 .2) a
54.8 (2.8) a
whole body Se, mg/kg dw
5.2 (0.4) c
11. 8 (0.9) b
14.7 (0.8) b
22.5 (1.4) a
34.4 (2.3) a
27.5 (4.4) a
Letters denote statistical groupings among treatments within each exposure period (p<0.05).
Chronic Value:
Using the logistic equation with a log transformation of the exposure
concentrations (TRAP program), the ECi0 and EC2o values for reduction
in body weight are 15.08 and 17.82 mg Se/kg dw whole body and 27.76
and 32.53 mg Se/kg dw muscle tissue.
C-3 Draft Document
-------
White sturgeon (Tashjian et al 2006)
350
300
^ 250
tn
CO
£ 200
o
c
IB 150
T3
-° 100
50
0
.
-
\
\
\
V
V
3 .9 1.0 1.1 1.2 1.3 1.4 1.5 1.6 1.7 1.8
Log([Se] muscle, mg/kg dw)
Parameter Summary (Logistic Equation Regression Analysis)
Parameter Guess FinalEst StdError 95%LCL 95%UCL
LogXSO 1.6006 1.6303 0.0314 1.5304 1.7301
S 1.6574 2.938 0.925 -0.005 5.882
YO 284.2 286.3 18.9 226.1 346.5
Effect Concentration Summary
% Effect
50.0
20.0
10.0
5.0
Xp Est
42.69
32.53
27.76
23.98
95%LCL
33.92
21.17
15.63
11.75
95%UCL
53.72
49.99
49.30
48.93
16/19/2009 10:32
MED Toxic Response Analysis Model, Version 1 03
C-4 Draft Document
-------
White sturgeon (Tashjian et al 2006)
350
300
j£ 250
of
CD
2 200
o
IB 150
-o
-° 100
50
0
.(
-
5 .7 .8 .9 1.0 1.1 1.2 1.3 1.4 1.5 1.6
Log([Se] whole body, mg/kg dw)
Parameter Summary (Logistic Equation Regression Analysis)
Parameter Guess FinalEst StdError 95%LCL 95%UCL
LogX50 1.3403 1.3750 0.0643 1.1702 1.5797
S 2.283 2.794 1.908 -3.277 8.865
YO 284.2 294.2 45.0 151.0 437.3
Effect Concentration Summary
% Effect
50.0
20.0
10.0
5.0
Xp Est
23.71
17.820
15.078
12.926
95%LCL
14.80
6.890
4.160
2.587
95%UCL
37.99
46.090
54.655
64.584
06/19/2009 10:42
MED Toxic Response Analysis Mode), Version 1.03
C-5 Draft Document
-------
Teh, S.J., X. Deng, D-F Deng, F-C Teh, S.S.O. Hung, T.W. Fan, J. Liu, R.M. Higasi. 2004 Chronic
effects of dietary selenium on juvenile Sacramento splittail (Pogonichthys macrolepidotus). Environ. Sci.
Technol. 38: 6085-6593.
Test Organism:
Exposure Route:
Dietary Treatments:
Test Duration:
Study Design:
Effects Data:
Sacramento splittail (Pogonichthys macrolepidotus); juveniles 7-mos.old
Dietary only
8 graded levels of dietary Se; dietary levels obtained by combining selenized
yeast with Torula (non-active) yeast. Selenized yeast contained approximately
21% of Se as selenomethionine and proteinaceous Se forms. Diet was formulated
as pellets by mixing dry ingredients with water and oil, fan-dried, crumbled and
sieved. Analyzed levels: 0.4 (no selenized yeast), 0.7, 1.4, 2.7, 6.6, 12.6, and 57.6
mg/kg.
Fish were fed twice daily with a daily feeding rate of 3% BW in first 5 months
and then adjusted to 2% BW thereafter.
9 months
A flow-through system with 40 fish/tank (24 total tanks) was used; each tank
held 90 L. Flow rate was 4 L/min. Water temperature was maintained at 23°C for
6 months and then 18°C for last 3 months due to failure of water heating system.
5 fish were sampled from each tank at 5 and 9 months and measured for gross
deformities, length, weight, Se in liver and muscle. Sections of the liver were
kept for histopathology. Condition factor (100 x BW/length), heptatosomatic
index (100 x liver weight/BW), BCF (total organ Se/dietary Se) were determined.
Mortality was observed in the two highest dietary treatments: 10 and 34.3%,
respectively. No mortalities were observed in fish fed diets # 12.6 mg/kg. No
significant difference in growth offish fed 12.6 mg/kg Se in diet, but there was in
the fish fed 26.6 mg/kg Se. See table below for levels of Se in fish at 9 months
and associated effects.
Authors determined prevalence of deformities was higher in fish fed 6.6 and 12.6
mg/kg Se in their diet, however a dose-response relationship did not occur (e.g.,
no deformities in high concentration). Gross pathology was a more sensitive
endpoint than growth.
C-6 Draft Document
-------
Summary of effects and assoc. dietary and tissue concentrations in Sacramento splittail after 9
month exp.
Dietary conc'n mg/kg
Se in liver, mg/kg dw
Se in muscle, mg/kg dw
0.4
20.1
6.6
0.7
18.6
6.9
1.4
20.0
9.2
2.7
23.0
10.1
6.6
26.8
15.1
12.6
31.3
18.9
26.0
40.4
29.4
57.6
73.7
38.7
Liver histopathology (mean lesions scores, N=15)
Macrophage aggregate
Glycogen depletion
Single cell necrosis
Fatty vacuolar degerneration
Eosinophilic protein droplets
Sum of mean lesion scores
0.13
0
0
0
0
0.13
0.07
0
0
0
0
0.07
0.2
0.2
0
0
0
0.4
0.27
0
0.07
0.2
0
0.54
0.40
0.4
0.13
0.53
0
1.46
0.20
0.2
0
0.07
0
0.47
0.20
0
0.07
0.2
0.07
0.54
0.85
1.38
0.46
0.08
0.85
3.62
Gross Pathology (No. of deformities, N=15)
Facial deformities (eye, jaw, and mouth)
Body deformities (kyphosis, lordosis,
scoliosis)
Prevalence of deformity (%)
0
0
0
1
0
6.7
0
4
26.7
1
2
20
5
3
53.3
3
1
26.7
0
1
6.7
0
0
0
Chronic Value:
Comments:
Using gross pathology as the endpoint (prevalence of deformities, %), the
NOAEC is 10.1 mg Se/kg dw and the LOAEC is 15.1 mg/kg Se dw in muscle
tissue; MATC or CV = 12.34 mg/kg Se in muscle dw.
The above concentrations in juvenile muscle tissue cannot be exactly translated
into an equivalent egg-ovary or whole-body concentration in adult splittail. But
using the median egg-ovary to muscle ratio of 1.59 for the family Cyprinidae, the
NOEC and MATC would represent 16.1 and 19.6 mg Se/kg egg-ovary. Using the
median muscle to whole-body ratio of 1.26 for the family Cyprinidae, the NOEC
and MATC would represent 8.04 and 9.83 mg Se/kg whole body. However,
appropriateness of these conversion estimates rests upon uncertain assumptions
that the muscle concentrations in juvenile splittails equal those of adult splittails
under the same exposure conditions, and that splittail tissue ratios are those
typical of the family Cyprinidae.
The authors observed deformities including spinal deformities using fish that
were 7-months-old at test initiation. This is the only study in which deformities
were observed in fish that were not exposed maternally.
Deng et al. (2008) exposed Sacramento splittail juveniles (21-day post hatch) to
dietary selenium and dietary methylmercury in a two factorial design for four
weeks. No adverse effects (growth, condition factor, lethargy or abnormalities)
C-7 Draft Document
-------
were observed in the selenium only exposures. The splittail accumulated
approximately 3.5 mg Se/kg ww muscle in the highest dietary exposure (35 mg
Se/kg. Using the average percent moisture in fish muscle of 78.4% (May et al.
2000), the dw Se concentration is 16.2 mg Se/kg muscle indicating the
recommended CV does not over-estimate an effect concentration.
Rigby et al. (2010) re-analyzed the juvenile Sacramento splittail data generated in
the Teh et al. (2004) study. The authors used logistic regression to estimate EC
values for deformities on a culled data set which eliminated the three highest
dietary treatments due to their departure from a standard concentration-response
relationship. The ECi0 value for the culled data set was 7.9 mg Se/kg dw muscle
which is lower than the recommended CV of 12.3 mg Se/kg dw muscle. Due to
the lack of a concentration-response relationship across the entire dietary range
and the lack of effects in the Deng et al. (2008) study, an ECi0 of 7.9 mg Se/kg
dw muscle is too uncertain for a recommended CV. Although the recommended
CV of 12.3 mg Se/kg dw muscle is based on deformities (an uncertain response),
it is considered representative of an effect level for this species because of the
significant reductions in growth at the two highest test concentrations.
C-8 Draft Document
-------
Bennett, William N., Arthur S. Brooks, and Martin E. Boraas. 1986. Selenium uptake and transfer in
an aquatic food chain and its effects on fathead minnow larvae. Arch. Environ. Contain. Toxicol. 15:513-
517.
Test Organism:
Exposure Route:
Test Duration:
Study Design:
Fathead minnow (Pimephalespromelas; 2 to 8 day-old larvae).
Dietary only
Green alga, Chlorellapyrenoidosa were exposed to Se (H275SeO4) in culture
water for 3 days. Rotifers, Brachionus calyciflorus, were cultured in chambers
with selenium containing green algae at the ratio of 25 |o,g algae/ml to 50 jog
rotifer/ml for 5 hr. The rotifers were filtered to separate them from the algae and
immediately heat-killed. The Se concentration in the rotifers was measured for
75Se activity.
9 to 30 days
Selenium uptake by larval fathead minnows was measured in three experiments.
Se-contaminated and control rotifers for feeding to larval fish were prepared in
advance using the low algae:rotifer ratio. Daily equal volumes of rotifers were
divided among five 800 mL polypropylene larval chambers. Three chambers
received Se-contaminated rotifers and two received control rotifers. The rotifers
were dead at the time of feeding (heat killed).
Larval fish were hatched from eggs spawned in the laboratory. After hatching,
active larvae were divided equally among the larval test chambers (daily renewal
exposures using dechlorinated Lake Michigan water). Larvae were initially fed
rotifers raised on control algae (no selenium). The age of the larvae when first
fed Se-contaminated rotifers was 4, 9, and 3 days post-hatch for experiments 1, 2,
and 3, respectively. Larval fish were fed Se-contaminated rotifers for 7, 9, and 7
days in the 3 experiments. A post-exposure observation period of 19 and 2 days
was used for experiments 1 and 2, respectively. During this time the larvae were
fed control rotifers. Daily, larvae from a replicate were removed from the test
chamber, washed, placed in a 20 ml vial, and counted for 75Se activity for 20 min.
All larvae were then placed in test chambers with fresh food rations. At the end
of the study all fish were individually dried and weighed.
C-9 Draft Document
-------
Initial feeding of control diet
(days)
Day Se diet first fed
Day Se diet last fed
Observation days on control diet
Age at study termination (days)
Experiment 1
3
4
11
19
30
Experiment 2
8
9
17
2
19
Experiment 3
2
3
9
0
9
Effects Data:
Mean food Se concentration
(mg/kg)
Food intake (|o,g rotifers/larva)
Initial larvae mean dry wt. at start
of Se-laden food (|o,g)
Final larvae mean dry wt. (|o,g) at
end of test
Final mean larval Se content (|o,g
Se/larva)b
Final mean larval Se
concentrations (mg Se/kg dw)
Experiment 1
>70
50
90
1470 (Control)
800 (Treatment)3
0.0062
43.0
Experiment 2
68
1330
400
1888 (Control)
1354 (Treatment)3
0.0700
51.7
Experiment 3
55
1190
100
475 (Control)
4 16 (Treatment)
0.0248
61.1
a Significantly different from the control.
b Values when Se-laden feeding was ended.
Chronic Value:
Selenium was measured in the test water during the feeding exposures, but the
concentrations were insignificant (0.84 |o,g/L). Survival was not affected by the
selenium exposures. Preliminary tests showed that fathead minnow larvae would
reach plateau concentrations of selenium within the 7- to 9-day exposure periods.
The food supply was sufficient to sustain growth of the larvae during the study,
according to the authors. The authors state that selenium uptake and higher
selenium content in experiment 2 larvae was due to their larger size and ability to
consume more rotifers/unit time. Se-exposed larvae were significantly smaller
(p<0.05) in mass than controls for experiments 1 and 2.
GM of mean larval Se concentrations measured in the three experiments, i.e.,
43.0, 51.7, and 61.1 mg/kg dw WB, respectively, is 51.40 mg Se/kg dw.
C-10 Draft Document
-------
Dobbs, M.G., D.S. Cherry, and J. Cairns, Jr. 1996. Toxicity and bioaccumulation of selenium to a
three-trophic level food chain. Environ. Toxicol. Chem. 15:340-347.
Test Organism:
Exposure Route:
Rotifer (Brachionus calyciflorus}, and fathead minnow (Pimephales promelas)
12 to 24 hr-old at start.
Dietary and waterborne
Water
Filtered and sterilized natural creek water supplemented with nutrients (Modified
Guillard's Woods Hole Marine Biological Laboratory algal culture medium) for
algal growth. Sodium selenate (Na2SeO4) was added to test water to obtain
nominal concentrations of 100, 200, or 400 (ig Se/L. Concentrations remained
stable and equal in each trophic level.
Control Diet
No selenium was added to the water medium for the alga; green alga was free of
selenium for the rotifer; and rotifers were free of selenium for the fathead
minnow.
Selenium Diet
Sodium selenate was added to the culture medium for the alga; green alga
thereby contained a body burden for the rotifer; and rotifers thereby contained a
body burden for the fathead minnow.
Dietary Treatments: Each trophic level had a different treatment. The green alga was exposed directly
from the water (1, 108.1, 204.9, 397.6 (ig total Se/L); rotifers were exposed from
the water (1, 108.1, 204.9, 393.0 (ig total Se/L) and the green alga as food (2.5,
33, 40, 50 mg Se/kg dry wt); and the fathead minnow were exposed from water
(1, 108.1, 204.9, 393.0 ng total Se/L) and the rotifer as food (2.5, 47, 53, 60 mg
Se/kg dry wt.).
Test Duration:
Study Design:
25 days
A flow-through system utilizing a stock solution of filtered and sterilized creek
water controlled at 25°C was used to expose three trophic levels of organisms.
Approximately one liter of media was pumped from the algal chamber into the
rotifer chamber each day. A cell density between 3 and 6 x 106 cells/ml was
delivered to the rotifer chambers. Rotifers were started at a density of 151.4 ± 7.7
females/ml and one liter/day of rotifers containing culture water was
intermittently pumped into the minnow chamber. (B. calyciflorus has a life span
of about 7 days at 25°C.) The pump was necessary to overcome the swimming
ability of rotifers to avoid an overflow tube. Larval fathead minnows
(35/chamber) were prevented from escaping by a screened overflow. Chambers
were cleaned daily and aeration was provided. All chambers were duplicated for
test replication and water was measured for selenium on days 0, 2,6,7, 11, 14,
17, 20, and 24. All algal and rotifer biomass and selenium samples were made on
these days. Fathead minnow chambers were measured for biomass, dissolved
C-11 Draft Document
-------
Effects Data:
selenium, and tissue selenium concentrations of days 0, 7, 11, 14, 20, and 24.
Additional measurements were made in the 200 (ig Se/L test chambers on the
fathead minnow on day 16. Selenium concentrations were maintained near the
nominal concentrations and the standard deviation of mean concentrations was
less than 4 percent.
Rotifers. Rotifers did not grow well and demonstrated reduced survival at all
selenium exposure concentrations during the 25 day test. By test day 7 only the
lowest test concentration (108.1 ug/L) had surviving rotifers which showed a
decrease in selenium content from test days 18 through 25. A reduction in rotifer
biomass was discernable by test day 4 in the selenium treatments and since all
test concentrations had viable rotifer populations present, the effect level was
calculated using these data.
Effect of Dietary and Waterborne Selenium on Rotifers after 4 Days Exposure
Se in water, jig/L
1
108.1
202.4
393
Se in diet, mg/kg dw
2.5
33
40
50
Se in rotifer tissue,
mg/kg dw
2.5
40
54
75
rotifer biomass, mg/ml
dw
0.028
0.025
0.011
0.003
Fathead minnows. Due to the reduction of rotifer biomass in the higher test
concentrations, fish mortality and reduction in fish growth observed in the latter
days of the test was difficult to discern between effects from starvation and
selenium toxicity. The data from test day 8 was selected for determining the
effect of selenium on fathead minnows because starvation could be excluded as a
variable.
C-12 Draft Document
-------
Effect of Dietary and Waterborne Selenium on Larval Fathead Minnows after 8 Days Exposure
Se in water, jig/L
1
108.1
202.4
393
Se in diet, nig/kg dw
2.5
47
53
60
Se in fathead minnow
tissue, mg/kg dw
2.5
45
75
73
Average fish weight,
mg dw
0.8
0.7
0.4
0.2
Chronic Value:
Rotifers
Fish
42.36 mg Se/kg dw (EC20)
< 73 mg Se/kg dw (LOAEC) - not amenable to statistical treatment; the LOAEC
was based on the observation that a >50 percent reduction in mean fish weight
occurred at this tissue concentration.
C-13 Draft Document
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Schultz, R. and R. Hermanutz. 1990. Transfer of toxic concentrations of selenium from parent to
progeny in the fathead minnow (Pimephalespromelas). Bull. Environ. Contam. Toxicol. 45:568-573.
Test Organism:
Exposure Route:
Study Design:
Effects Data :
Chronic Value:
Fathead minnow (Pimephales promelas; Adults)
Dietary and waterborne
Selenite was added to artificial streams which entered the food web; thus, fish
were also exposed to selenium in the diet.
Four Monticello artificial streams were used for the study which lasted from
September 1987 to September 1988. For each study, two streams (treated) were
dosed continuously to achieve 10 |o,g/L and two streams served as controls. Mean
selenium concentrations at the head of the treated streams were 9.8 V 1.2 and
10.3 V 1.7 |o,g/L, respectively. The concentrations of selenium measured in the
water from controls streams were all less than the detection limit, i.e., 2 :g/L.
Spawning platforms were submerged into each stream. One subset of six embryo
samples (n = 2000 embryos per sample) were collected from the streams for
selenium analysis. Another subset often embryo samples were reared in
incubation cups receiving the same stream water dosed with sodium selenite via a
proportional diluter. The treated embryos in egg cups received an average 9.7 V
2.6 :g Se/L. Samples of hatched larvae were analyzed for selenium content while
others were inspected for occurrence of edema and lordosis. Prior to test
termination, female parents were seined. The mean selenium content in the
ovaries of seven to eight females from the treated and control streams was
reported.
Edema and lordosis occurred in approximately 25 percent of the fish spawned
and reared in 10 :g Se/L. Corresponding occurrence in control fish incubated in
the egg cups was only 1 and 6 percent, respectively. Selenium residues in the
ovaries of females from the control and treated streams were 0.77 and 5.89 mg
Se/kg ww. Using 75.3 percent moisture content in the eggs/ovaries (average
value for fathead minnow ovaries and eggs from GEI Consultants 2008 and
Rickwood et al. 2008), these concentrations equate to 3.12 and 23.85 mg Se/kg
dw.
The NOAEC for egg/ovary is <23.85 mg Se/kg dw.
C-14 Draft Document
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Beyers, D.W. and Sodergren, C. 200 la. Evaluation of interspecific sensitivity to selenium exposure:
Larval razorback sucker versus flannelmouth sucker. Larval Fish Laboratory. Department of Fishery and
Wildlife Biology, Colorado State University, Fort Collins, Colorado.
Test Organism:
Exposure Route:
Study Design:
Effects Data :
Chronic Value:
Larval flannelmouth sucker (Catostomus latipinnis) and larval razorback sucker
(Xyrauchen texanus)
Dietary and waterborne - laboratory exposure (28-d early life stage)
Continuous flow diluter supplied a range of aqueous test concentrations <1, 25.4,
50.6, 98.9, and 190.6 :g/L selenate. Well water was used as the dilution water.
Across the range of aqueous exposure concentrations, each test chamber was fed
the same daily ration of living rotifers containing selenium at <0.702, 1.35, 2.02,
4.63, and 8.24 mg/kg dw, respectively. Rotifers accumulated selenium from
algae (Chlorella vulgaris) exposed to 0, 25, 50, 100, and 200 :g/L selenate.
Replicated (n=4) exposure beakers using a randomized, balanced 5x2 factorial
design (1st factor - selenium; 2nd factor - species). Survival was monitored daily
and growth measured at the end of the 28-day exposure. Selenium was measured
in the larvae at the end of the 28-day exposure.
No survival effects were observed and there were no decreases in fish weight or
length. Fish mass was found to increase as a function of selenium concentration.
The chronic values for the flannelmouth sucker and razorback sucker were >10.2
and >12.9 mg Se/kg dw, respectively, based on the concentrations of selenium
measured in whole-body tissue of larval fish at the highest water and dietary
selenium concentrations.
C-15 Draft Document
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Beyers, D.W. and Sodergren, C. 2001b. Assessment of exposure of larval razorback sucker to selenium
in natural waters and evaluation of laboratory-based predictions. Larval Fish Laboratory. Department of
Fishery and Wildlife Biology, Colorado State University, Fort Collins, Colorado.
Test Organism:
Exposure Route:
Study Design:
Effects Data:
Chronic Value:
Larval razorback sucker (Xyrauchen texanus)
Dietary and waterborne - laboratory exposure (28-d early life stage)
Larvae were exposed in a daily static-renewal system to control water
(reconstituted very hard) and site waters: De Beque, Orchard Mesa, North Pond
diluted 50%, and North Pond. Each water type received either a control diet
(rotifers) or a diet previously exposed to the site water (site food: rotifers fed
algae exposed to respective site water).
Replicated (n=4) exposure beakers using a randomized, balanced 5x2 factorial
design (1st factor - test water type; 2nd factor - rotifers cultured in control water or
in site water). Survival was monitored daily and growth measured at the end of
the 28-day exposure. Selenium was measured in the larvae at the end of the 28-
day exposure.
No survival effects were observed. There were no significant decreases in growth
offish exposed to both site water and site food compared to fish exposed to
control water and control food. There was a significant increase in growth offish
exposed to site water and control food relative to fish exposed to control water
and control food (p<0.0001). There were reductions in the growth offish (14%)
exposed to site water and site food compared to site water and control food
(pO.OOOl). Due to the lack of a dose-response relationship in both the
concentration of selenium in the food (rotifers) and growth, and the concentration
of selenium in the fish larvae and growth, the authors did not attribute the effect
of site food on the growth offish to selenium.
The NOAEC for the razorback sucker larvae in the four site water types based on
selenium in whole-body tissue were: De Beque >5.45 mg Se/kg dw; Orchard
Mesa >11 mg Se/kg dw; North Pond 50% dilution >41.1 mg Se/kg dw; North
Pond >42 mg Se/kg dw. Because no significant effects were observed in larvae
exposed to North Pond water at >42 mg Se/kg dw whole-body tissue, this value
was selected as the chronic value for the study.
C-16 Draft Document
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Muscatello, J.R., P.M. Bennett, K.T. Himbeault, A.M. Belknap and D.M. Janz. 2006. Larval
deformities associated with selenium accumulation in northern pike (Esox lucius) exposed to metal
mining effluent. Environ. Sci. Technol. 40:6506-6512.
Test Organism:
Exposure Route:
Test Duration:
Study Design:
Effects Data:
Northern pike (Esox lucius)
Dietary and waterborne - field exposure
Eggs were collected in the field and incubated in the laboratory. The test was
terminated when the majority of the fry exhibited swim-up and had absorbed the
yolk.
The study area was Key Lake uranium milling operation in north-central
Saskatoon. Spawning northern pike were collected from four sites, one reference
(Davies Creek) and three exposure sites, David Creek near-field (high exposure),
Delta Lake (medium exposure), and David Creek far-field (low exposure). The
exposure sites were located approximately 2, 10 and 15 km downstream of the
effluent discharge. Milt and ova were stripped from ripe fish and eggs were
fertilized in the field. Females were saved for metal analysis and age
determination. Subsamples of ova (prior to fertilization) were collected for metal
analysis.
Although the study sites represent open systems where fish can potentially
migrate among sites, radiotelemetry data from tagged adult pike (Muscatello and
Janz, unpublished data) indicate high site fidelity at the "high" and "medium"
exposure sites (lakes). In contrast, the "low" exposure site likely represents pike
that migrated from further downstream sites that were likely of similar Se
exposures as the reference site.
Eggs were incubated using a two-way ANOVA experimental design using water
collected from reference or exposure sites. So, embryos originating from
reference or exposure site females were incubated in either reference or
appropriate exposure water. In addition, embryos from reference site females
were incubated in water from all four study sites. 50 viable embryos from each
individual female were transferred to each of four replicate incubation chambers.
Cumulative time to 50% eyed, 50% hatch and 50% swim-up were determined.
When the majority of the fry exhibited swim-up and had absorbed the yolk, the
remaining fry were preserved and examined for deformities.
Mean egg diameter and fertilization success did not differ among sites.
Cumulative embryo mortality throughout incubations was not significantly
different among the sites ranging from 45 to 60%. There were no significant
differences in the cumulative time to reach 50% eyed embryos, 50% hatch or
50% swim-up among treatments. Difference in the percent total deformities
between test waters used during embryo incubation exposures were not
significant, so the data were combined for each site (see Table below).
C-17 Draft Document
-------
Selenium concentrations in eggs and muscle from female northern pike collected from reference
and exposed sites and associated total deformities in embryos
Site
Davies Creek
Davies Creek
Davies Creek
Davies Creek
Davies Creek
David Creek (far field)
David Creek (far field)
David Creek (far field)
David Creek (far field)
David Creek (far field)
Delta Lake
Delta Lake
Delta Lake
David Creek (near field)
David Creek (near field)
Site ID
Reference
Reference
Reference
Reference
Reference
Low
Low
Low
Low
Low
Medium
Medium
Medium
High
High
Female
1
2
3
4
5
1
2
3
4
5
1
2
3
1
2
[Se] mg/kg dw
Egg
3.45
2.72
3.39
3.72
2.69
3.39
4.07
4.07
4.07
3.4
43.19
24.53
26.14
48.23
N/A*
Muscle
0.86
1.89
2.56
1.34
1.04
1.95
2.04
1.26
2.48
1.26
17
16.52
16.52
47.82
28.72
Total
deformities %
17
2.5
15.51
7.13
10.41
20.32
13.19
15.33
18.83
11.8
37.8
31.71
26.29
39.5
N/A*
* female had no eggs
Significant increases in total deformities (edema, skeletal deformities,
craniofacial deformities and fin deformities) were observed in fry originating
from pike collected at the medium exposure site. Determination of an effect level
for the percent total deformities relative to the concentration of selenium in eggs
or in female muscle tissue was not amenable to analysis by TRAP. One
requirement of TRAP is to have a response greater than 50%, which was not
satisfied with the available data.
When data are not amenable to determining an effect level using a software
program, such as TRAP, one way to estimate the effect level is to make a direct
measurement of effect at an exposure or tissue concentration. For example, if
only a control and one exposure concentration, 10 |o,g/L, were tested in an acute
toxicity test and there was 100% survival in the control and 35% in the 10 |o,g/L,
the effect level would be an EC35 of lOjo, g/L. Such an approach was used to
estimate effect in the Muscatello et al. data. Because no significant differences
were observed in either selenium concentrations in eggs or percent total
deformities between the reference and low exposure site, the data from these 10
sites were combined. Similarly, the egg selenium and total deformity data were
combined for the 4 medium and high exposure sites. These means, geometric for
the selenium concentrations and arithmetic for the percent total deformities, are
given in the following table.
C-18 Draft Document
-------
Mean selenium in northern pike egg and effect values for reference and exposure sites
Sites
Reference sites
(includes low exposure)
exposure sites
[Se] in eggs,
mg/kg dw
(geometric mean)
3.462
34.00
Total deformities,
%
(arithmetic mean)
13.20
33.82
Total deformities, %
(accounting for reference
deformities and transformed
to new scale)8
0
23.76
The % total deformities in the reference and exposed sites were normalized to the reference effect
(13.2%) and then transformed to a new scale (100%). i.e, Abbott's formula.
The percent affected becomes 24% or an EC24 and the effect level is 34.00 mg
Se/kg dw in eggs
Chronic Value:
EC24 = 34.00 mg Se/kg dw in eggs. Note: an ECio cannot be estimated with the
data.
C-19 Draft Document
-------
Hamilton, S.J., K.J. Buhl, N.L. Faerber, R.H. Wiedermeyer and F.A. Bullard. 1990. Toxicity of
organic selenium in the diet of chinook salmon. Environ. Toxicol. Chem. 9:347-358.
Test Organism:
Exposure Route:
Chinook salmon (Oncorhynchus tshawytscha Walbaum; swim-up larvae)
Dietary only
Control Diet
Oregon moist pellet diet where over half of the salmon meal was replaced with
meal from low-selenium mosquitofish (1.0 mg Se/kg dw) collected from a
reference site.
Selenium Diet # 1
Oregon moist pellet diet where over half of the salmon meal was replaced with
meal from high-selenium mosquitofish (35.4 mg Se/kg dw) collected from the
San Luis Drain, CA, termed SLD diet.
Selenium Diet #2
Oregon moist pellet diet where over half of the salmon meal was replaced with
meal from low-selenium mosquitofish same as in the control diet, but fortified
with seleno-DL-methionine (35.5 mg Se/kg dw), termed SeMet diet.
Dietary Treatments: Each selenium diet was formulated to contain about 36 mg Se/kg dw as the high
exposure treatment. The remaining treatments were achieved by thoroughly
mixing appropriate amounts of high-exposure treatment diet with control diet to
yield the following nominal concentrations (3,5, 10, and 18 mg Se/kg dw).
Test Duration:
Study Design:
90 days
Each dietary treatment was fed twice each day to swim-up larvae (n=100) in each
of two replicate aquaria that received 1 L of replacement water (a reconstituted
experimental water that simulated in quality a 1:37 dilution of water from the San
Luis Drain, CA minus the trace elements) every 15 minutes (flow-through
design). Mortality was recorded daily. Growth was evaluated at 30-day intervals
by measuring the total lengths and wet weights of two subsets of individual fish
(n=10x2) held in separate 11.5 L growth chambers within each replicate
aquarium. Tissue samples were collected for whole-body selenium
determinations (dw basis) at 30-day intervals throughout the study; 10, 5, and 2
fish were sampled from each duplicate treatment after 30, 60, and 90 days of
exposure, respectively. Concentrations of selenium measured in water were
below the limit of detection (1.5-3.1 |o,g/L) in all dietary selenium exposure
concentrations.
C-20 Draft Document
-------
Effects Data:
The magnitude of reduced growth was most evident in the weight of the fish,
although total length was significantly reduced in fish fed high Se-laden diets as
well. The effect of increasing dietary selenium on mean larval weight was similar
in both the SLD and seleno-methionine diets.
Effect of San Luis Drain Diet on Growth and Survival of Chinook Salmon Larvae after 60 Days
Se in diet, nig/kg dw
1
3.2
5.3
9.6
18.2
35.4
Se in chinook salmon,
mg/kg dw
0.9
3.3
4.5
8.4
13.3
29.4
Mean larval weight, g
3.35
2.68
2.76
2.8
2.62
1.4
Survival, %
99
97.3
93
95
92.4
89
Effect of Seleno-methionine Diet on Growth and Survival of Chinook Salmon Larvae after 60
Days
Se in diet, mg/kg dw
1
3.2
5.3
9.6
18.2
35.4
Se in chinook salmon,
mg/kg dw
0.9
2
3.1
5.3
10.4
23.4
Mean larval weight, g
3.35
3.08
3.22
3.07
2.61
1.25
Survival, %
99
100
95
94.1
92.4
62.5
C-21 Draft Document
-------
Chronic Value:
Due to unacceptable control mortality of swim-up larvae in control treatments
after 90 days (33.3 percent - SLD diet; 27.5 percent - SeMet diet), chronic values
had to be determined from respective values reported after 60 days (tables
above).
Analysis of the elemental composition of the SLD diet indicated that B, Cr, Fe,
Mg, Ni and Sr were slightly elevated compared to the control and SeMet diets.
No additional analyses were performed to determine the presence of other
possible contaminants, i.e., pesticides.
Diet
type
SLD
SeMet
EC2o values
Survival
(after 60 d of
exposure)
Tissue Se
(mg/kg dw)
NAa
NAa
Growth
(after 60 d of exposure)
Whole body Tissue Se
(mg/kg dw)
15.73
10.47
ECio values
Growth
(after 60 d of exposure)
Whole body Tissue Se
(mg/kg dw)
11.14
7.355
The EC2o and ECio values for survival of swim-up larvae versus levels of selenium for the SLD and
SeMet dietary exposure could not be estimated using non-linear regression.
C-22 Draft Document
-------
Hamilton et al (1990) Chinook Salmon fed SLD Diet
Logistic Equation, Three Parameter Model, Se concentrations logio transformed
4.0 r
3.5
4
_ 3.0
i> 2.5
CO
>
CO
CO
CD
1.5
1.0
.5
0
0 .2 .4 .6 .8 1.0 1.2
Log(Se in Chinook Salmon mg/kg dw)
1.4
1.6
Guess
FinalEst SE
95%LCL 95%UCL
LogXSO
StDev
YO
%Effect
50
20
10
5
Total
Model
Error
1.453
1.353
2.968
XpEst
28.379
15.734
11.143
8.1085
DF
5
2
3
1.453
1.353
2.968
95% LCL
16.62
5.7003
2.4771
1.1213
SS
2.0749
1.8202
0.2547
7.30E-02
6.67E-01
1.89E-01
95% UCL
48.458
43.431
50.127
58.637
MS
0.41498
0.91009
8.49E-02
1.2206 1.6854
-7.71E-01 3.4769
2.3651 3.5709
F P
10.719 0.95699
C-23 Draft Document
-------
Chinook salmon SeMet diet (Hamilton et al. 1990)
.2 .4 .6 .8 1.0 1.2
Log([Se]whole body, mg/kg dw)
1.4
Parameter Summary (Logistic Equation Regression Analysis)
Parameter Guess FinalEst StdError 95%LCL 95%UCL
LogXSO 1.3148 1.2823 0.0242 1.2053 1.3593
S 0.6971 1.3214 0.1826 0.7404 1.9025
YO 3.217 3.239 0.067 3.027 3.452
Effect Concentration Summary
% Effect
50.0
20.0
10.0
5.0
Xp Est
19.156
10.472
7.355
5.312
95%LCL
16.045
7.516
4.595
2.899
95%UCL
22.870
14.591
11.775
9.733
06/19/2009 13 52
MED Toxic Response Analysis Model, Version 1 03
C-24 Draft Document
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Hilton, J.W. and P.V. Hodson. 1983. Effect of increased dietary carbohydrate on selenium metabolism
and toxicity in rainbow trout (Salmo gairdneri). J. Nutr. 113:1241-1248.
Test Organism:
Exposure Route:
Test Treatments:
Test Duration:
Study Design:
Effects Data:
Rainbow trout (Oncorhynchus mykiss; juvenile; approx. 0.6 g each)
Dietary only
Low carbohydrate diet (LCD)
This diet contained capelin oil at 11 percent of the diet with cellulose as the
filler.
High carbohydrate diet (HCD)
This diet contained cerelose at 25 percent of the diet with cellulose as the filler.
For both diets, the selenium was supplemented as sodium selenite which was
mixed with cellulose and then added to the diet as a selenium premix.
The two diets were supplemented with selenium (as sodium selenite) at the rate
of 0, 5, or 10 mg/kg dw to make up the six different dietary selenium treatments
(n = 3 low carbohydrate diet; n= 3 high carbohydrate diet). The six diets were fed
to duplicate groups of 100 fish. The trout were fed to satiation 3-6 times per day.
Measured concentrations of selenium in the low carbohydrate diet were: 0.6
(control), 6.6, and 11.4 mg/kg dw, and the measured concentrations of selenium
in the high carbohydrate diet were: 0.7 (control), 6.6, and 11.8 mg/kg dw. The
tanks received a continuous flow of water with a flow rate of 3-4 liters per
minute.
16 weeks
Body weights, feed: gain ratios, and total mortalities were determined after each
28-day interval. After 16 weeks, approximately 20 fish were randomly removed
from each tank, weighed, and blood was collected for hemoglobin, hematocrit,
and plasma glucose, protein, and calcium determination. The livers and kidneys
were then dissected. The livers were assayed for glycogen content, and samples
of both liver and kidney were assayed for selenium content. Additional
subsamples offish were sacrificed and assayed for selenium content and for ash,
crude protein, and moisture content (n=6 per treatment). Finally, 30 fish were
killed, their livers and kidneys dissected, and analyzed for Ca, Cu, Fe, Mg, P, and
Zn content.
The only overt sign of selenium toxicity was food avoidance observed in trout
fed the highest selenium content in both low and high carbohydrate diets, which
led to significantly reduced body weight after 16 weeks. There were no
significant differences detected between treatment groups in hematological
parameters. Kidney, liver, and carcass selenium levels increased with increasing
selenium content of the diet, however, only the liver selenium concentrations
were significantly affected by dietary selenium level, dietary carbohydrate level,
and the interaction between the two treatments. Mineral analysis of the kidney
showed significantly higher levels of calcium and phosphorous in trout reared on
the two highest levels of dietary selenium. Concentrations of copper in the liver
C-25 Draft Document
-------
increased significantly with increasing dietary selenium levels and decreasing
dietary carbohydrate levels.
Effect of Selenium in Low carbohydrate Diet to Rainbow Trout
Se in diet, nig/kg dw
0.6
6.6
11.4
Se in trout liver, mg/kg dw
0.8
38.3
49.3
Trout weight, kg/100 fish
3.3
3.3
1.8
Effect of Selenium in High carbohydrate Diet to Rainbow Trout
Se in diet, mg/kg dw
0.7
6.6
11.8
Se in trout liver, mg/kg dw
0.6
21.0
71.7
Trout weight, kg/100 fish
2.7
2.3
1.4
Chronic Value:
The following table lists the NOAEC, LOAEC and MATC for both diets in liver
tissue. EC values could not be determined for this study. Data did not meet
minimum requirements for analysis.
Diet
Low carb
high carb
NOAEC, mg Se/kg dw
liver
38.3
21.0
LOAEC, mg Se/kg dw
liver
49.3
71.7
MATC, mg Se/kg dw
liver
43.5
38.8
C-26 Draft Document
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Hicks, B.D., J.W. Hilton, and H.W. Ferguson. 1984. Influence of dietary selenium on the occurrence of
nephrocalcinosis in the rainbow trout, Salmo gairdneri Richardson. J. Fish Diseases. 7:379-389.
(Note: These data are the exact same as reported for the low carbohydrate diet in Hilton and Hodson
1983, with the addition of prevalence of nephrocalcinosis occurring in trout after 16 to 20 weeks of
consuming the contaminated test diets).
Test Organism:
Exposure Route:
Test Treatments:
Test Duration:
Study Design:
Effects Data:
Chronic Value:
Rainbow trout (Oncorhynchus mykiss; juvenile; approx. 0.6 g each)
Dietary only
This diet contained capelin oil at 11 percent of the diet with cellulose as the filler.
The selenium was supplemented as sodium selenite which was mixed with
cellulose and then added to the diet as a selenium premix.
The test diet was supplemented with selenium (as sodium selenite) at the rate of
0, 5, or 10 mg/kg dw to make up the three different dietary selenium treatments.
The three diets were fed to duplicate groups of 100 fish. The trout were fed to
satiation 3-6 times per day. Measured concentrations of selenium in the low
carbohydrate diet were: 0.6 (control), 6.6, and 11.4 mg/kg dw. The tanks received
a continuous flow of water with a flow rate of 3-4 liters per minute.
16 to 20 weeks
See Hilton and Hodson (1983). After 20 weeks on the test diets, ten fish were
randomly removed from each treatment. Tissues for histopathological
examination included the stomach, intestine and pyloric ceca (including
pancreas), spleen, liver, heart, kidney, skin, muscle, and gills.
Only effects of selenium on kidney tissue are included in the article. The kidneys
of the 10 trout fed the highest selenium content in the diet exhibited normal
appearance. Five of these trout exhibited precipitation of calcium in the tubules
with some epithelial necrosis, but no loss of epithelial continuity. Extensive
mineralized deposition of Ca within the tubules, tubular dilation and necrosis of
tubular epithelium, ulceration of tubules, and intestinal Ca mineralization was
observed in four of the ten fish.
Same as for growth of rainbow trout reported by Hilton and Hodson (1983). The
MATC estimated for growth of rainbow trout relative to final concentration of
selenium in liver tissue of trout reared on the low carbohydrate diet is the GM of
38.3 (NOAEC) and 49.3 (LOAEC) mg/kg dw, or 43.45 mg/kg dw.
EC values could not be determined for this study. Data did not meet minimum
requirements for analysis.
C-27 Draft Document
-------
Hilton, J.W., P.V. Hodson, and S.J. Slinger. 1980. The requirements and toxicity of selenium in
rainbow trout (Salmo gairdneri). J. Nutr. 110:2527-2535.
Test Organism:
Exposure Route:
Test Duration:
Study Design:
Effects Data:
Rainbow trout (Oncorhynchus mykiss; juvenile; approx. 1.28 g each)
Dietary only
A casien-torula yeast diet was formulated to contain geometrically increasing
levels of selenium from 0 to 15 mg/kg dw. The selenium was supplemented as
sodium selenite which was mixed with cellulose and then added to the diet as a
selenium premix.
20 weeks
Six test diets were fed to triplicate groups of 75 fish. The trout were fed to
satiation 3-4 times per day, 6 days per week, with one feeding on the seventh
day. Measured concentrations of selenium in the diet were: 0.07 (control), 0.15,
0.38, 1.25, 3.67, and 13.06 mg/kg dw. The tanks received a continuous flow of
dechlorinated tap water from the City of Burlington, Ontario municipal water
supply. The waterborne selenium content of this water was 0.4)0, g/L. During the
experiment, the fish were weighed every 2 weeks with the feeding level adjusted
accordingly. Mortalities were noted daily and the feed consumption for each
treatment was recorded weekly. After 4 and 16 weeks, three to six fish were
randomly removed from each tank, sacrificed, and their livers and kidneys
removed and weighed. An additional three to six fish were then obtained from
each treatment, killed, and prepared for tissue analysis. Organs and carcasses
were freeze-dried for determination of selenium concentration. After 16 weeks,
three more fish were removed. Kidney, liver, spleen and dorsal muscle tissue was
dissected for examination of histopathology. At the end of 8 and 16 weeks, four
to five fish were removed, sacrificed, and a blood sample was taken for
hematological measurements (hematocrit, red blood cell count, and blood iron
concentration). After 20 weeks, three to four more fish were removed, sacrificed,
and a blood sample was taken for measurement of glutathione peroxidase
activity.
There were no significant differences detected between treatment groups in
histopathology, hematology, or plasma glutathione peroxidase activity. Trout
raised on the highest dietary level of selenium (13.06 mg/kg dw) had a
significantly lower body weight and a higher number of mortalities (10.7;
expressed as number per 10,000 fish days) than trout from the other treatments
levels after 20 weeks of exposure.
C-28 Draft Document
-------
Effects on Juvenile Rainbow Trout
Se in diet, mg/kg dw
0.07
0.15
0.38
1.25
3.67
13.06
Se in Liver, mg/kg dw
0.6
0.95
2.4
11
40a
100b
Weight, g/fish
3.2
3.5
3.7
4.1
4.1
1.4
Mortality*
0
0
0.6
0.6
0
10.7
* expressed as number per 10,000 fish-days
a NOAEC
b LOAEC
Chronic Value:
NOAEC = 40 mg Se/kg dw
LOAEC = 100 mg Se/kg dw
MATC = 63.25 mg Se/kg dw
C-29 Draft Document
-------
Holm, J. 2002. Sublethal effects of selenium on rainbow trout (Oncorhynchus mykiss) and brook trout
(Salvelinusfontinalis). Masters Thesis. Department of Zoology, University of Manitoba, Winnipeg, MB.
Holm, J., V.P. Palace, K. Wautier, R.E. Evans, C.L. Baron, C. Podemski, P. Siwik and G. Sterling.
2003. An assessment of the development and survival of rainbow trout (Oncorhynchus mykiss) and brook
trout (Salvelinus fontinalis) exposed to elevated selenium in an area of active coal mining. Proceedings of
the 26th Annual Larval Fish Conference 2003, Bergen, Norway. ISBN 82-7461-059-B.
Holm, J., V.P. Palace, P. Siwik, G. Sterling, R. Evans, C. Baron, J. Werner, and K. Wautier. 2005.
Developmental effects of bioaccumulated selenium in eggs and larvae of two salmonid species. Environ.
Toxicol. Chem. 24: 2373-2381.
Test Organism:
Exposure Route:
Study Design:
Effects Data :
Rainbow trout (Oncorhynchus mykiss; spawning adults) and brook trout
(Salvelinus fontinalis; spawning adults)
Dietary and waterborne - field exposure
Total selenium concentrations measured at the high selenium site ranged from 6
to 32|o,g/L. Selenium was not measured at the reference streams; selenium
concentrations at reference locations in the area ranged from <0.5 to 2.2|o,g/L.
Spawning fish were collected at low selenium or reference streams (Deerlick
Creek, Wampus Creek and Cold Creek), a slightly elevated selenium stream
(Gregg Creek), and an elevated selenium stream (Luscar Creek) in the
Northeastern slopes region of Alberta, Canada. An active coal mine is the source
of selenium in the elevated streams. Eggs and milt from the spawning trout were
expressed by light pressure from abdomen. Individual clutches of eggs were
fertilized from a composite volume of milt derived from 3-5 males. Fertilized
eggs from individual females were reared to swim-up stage and examined for a
number of parameters including percent fertilization, mortality, edema, and
deformities (craniofacial, finfold, and spinal malformations). Similar studies
were conducted in 2000, 2001 and 2002. One notable difference is that the
embryos were incubated at SEC in 2000 and at SEC in 2001. The authors noted
that SEC is a better representation of the actual stream temperature during
embryo development.
Other than selenium, there were no significant differences in the concentrations
of other elements (Al, As, Sb, Ba, Be, Ni, B, Cd, Ca, Cr, Co, Cu, Fe, Pb, Li, Mg,
Mn, Hg, Mo, Ag, Sr, Tl, Th, Sn, Ti, U, V, Zn) in trout eggs between the low level
and elevated selenium streams. There are two ways to approach determination of
effects due to selenium in this study and both are presented here. The first
approach determines effects based on a comparison of average conditions
between streams (between streams approach). For example, if there is a
significant difference between the average frequency of deformities in a
contaminated stream and reference stream, the effect level for the between
streams approach would be the average concentration of selenium in the tissue
from the contaminated stream. The second approach evaluates individual
response variables (e.g., edema, deformities) against the individual selenium
C-30 Draft Document
-------
tissue concentrations for the combined contaminated and reference stream data
set with each year (within streams approach). This approach, which results in EC
estimates (e.g., ECio and EC2o) if the data meet the model assumptions, is
explained in the Calculations of Chronic Values section of the text.
Between streams approach: For each sampling location (stream), data for the
three years (Tables 1 and 2) were combined in the between streams analysis of
variance (ANOVA). For rainbow trout embryos, there were no significant
differences in fertilization, time to hatch and mortality between the streams with
elevated selenium and the reference streams. ANOVA indicated significant
differences in the frequency of embryonic effects between streams (Table 3). The
analysis did not prove useful; however, due to a higher occurrence of effects in
some of the reference streams relative to the exposed streams (Tables 3 and 4).
The between streams analysis, therefore, was not used to determine effect
concentrations for rainbow trout.
ANOVA of brook trout data indicated the only significant difference in
embryonic abnormalities among sites was craniofacial deformities (Tables 5 and
6). Significant differences were also found for fertilization and larval weight. The
highest average percent fertilization was observed at the site with the greatest
concentration of selenium in eggs, which indicates that the differences in
OO '
fertilization among sites were not caused by variation in selenium concentrations.
Because the percent of embryos with craniofacial deformities in Luscar Creek
was 7.9% (2.1% in Cold Creek), it was not considered biologically meaningful.
Likewise the significantly lower larval weights at the exposed sites was not large
(16% lower than Cold Creek larvae) and again coupled with the low occurrence
of abnormalities by the brook trout, a signature of selenium effects, the lower
larval weights were not considered biologically meaningful.
Within streams approach: As with the between streams analysis, data were
combined for the three years of study in the within streams analysis (Tables 1 and
2). Craniofacial deformities, skeletal deformities and edema in rainbow trout
embryo, as a function of selenium in egg ww, were fitted to a logistic curve from
which ECio and EC2o values were calculated (see table below and Figures 1 and
2). EC estimates for finfold deformities, length and weight of rainbow trout
embryos could not be made because of inadequate dose-response. The brook
trout data were not suitable for fitting logistic curves (Figure 3).
C-31 Draft Document
-------
Rainbow Trout EC Estimates using TRAP Logistic Equation, log([Se]egg)
Response
100% -
%craniofacial
100% -
%skeletal
100% -
%edema
EC20
Se, nig/kg ww
11.4
11.0
9.9
Se, nig/kg
dwa
29.4
28.4
25.5
ECio
Se, nig/kg ww
10.3
8.2
9.5
Se, nig/kg
dwa
26.5
21.1
24.5
Comment
Large SE for
steepness
ww to dw was converted using 61.2% moisture for rainbow trout eggs (Seilor and Skorupa, 2001)
C-32 Draft Document
-------
Table 1. Rainbow trout embryo-larval parameters collected from a high Se site (Luscar Creek), an
intermediate Se site (Gregg River), and reference sites (Deerlick Creek and Wampus Creek) in
northeastern Alberta over three consecutive years.
Year
2000
2000
2000
2000
2000
2000
2000
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2002
Site Female #
Luscar
Luscar
Luscar
Deerlick
Deerlick
Deerlick
Deerlick
Luscar
Luscar
Luscar
Luscar
Luscar
Luscar
Luscar
Luscar
Luscar
Luscar
Deerlick
Deerlick
Deerlick
Deerlick
Deerlick
Deerlick
Deerlick
Deerlick
Deerlick
Deerlick
Gregg
Gregg
Gregg
Gregg
Gregg
Wampus
Wampus
Luscar
11
12
14
16
17
18
15
1
3
4
8
14
32
33
39
40
41
8
9
10
16
17
21
22
23
25
39
2
3
5
9
18
9
13
3
Se in eggs,
mg/kg ww
6.84
6.66
11.6
1.78
1.39
1.00
5.01
5.39
8.39
6.48
4.47
10.4
5.64
3.88
5.14
3.36
11.7
3.68
3.08
1.62
2.62
2.79
1.96
3.13
3.03
3.32
2.43
4.57
4.49
4.05
5.09
5.97
2.66
2.04
5.4
%craniofacial %skeletal %finfold
deformities deformities deformitie
s
7.18
1.48
14.43
0.63
0
0
0
7.35
6.29
22.22
12
34.55
8.24
5.26
1.91
11.62
37.67
9.55
5.39
7.89
24.24
14.13
13.27
1.09
9.65
9.25
11.89
11.97
5.58
4.95
20
16.13
16.07
7.84
60.47
13.26
4.43
23.71
1.9
0
0.86
0
6.76
4.97
22.22
9.33
44.85
5.97
6.58
3.18
7.05
83.41
5.45
4.98
7.89
48.48
15.22
35.71
2.17
14.04
13.29
9.09
7.75
9.3
5.45
13.85
19.35
0
9.8
27.9
1.66
0.74
7.22
0.63
0
0
0
3.53
2.98
33.33
2.67
4.24
3.13
9.21
0
5.39
3.59
1.36
0.41
5.26
3.03
4.35
7.14
0
3.51
7.51
7.69
15.49
2.33
2.48
15.38
41.94
1.79
1.31
93
%edema
4.97
1.85
85.57
0.63
0
0
0
2.94
6.95
26.67
10.67
43.64
9.09
3.95
1.27
6.64
87
5.45
2.07
10.53
12.12
20.65
25.51
1.09
7.89
8.09
14.69
7.04
4.65
5.94
16.15
35.48
7.14
7.84
14
C-33 Draft Document
-------
2002
2002
2002
2002
2002
2002
2002
2002
2002
2002
2002
2002
2002
2002
2002
2002
2002
2002
2002
Luscar
Luscar
Luscar
Luscar
Luscar
Luscar
Luscar
Deerlick
Deerlick
Deerlick
Deerlick
Deerlick
Deerlick
Gregg
Wampus
Wampus
Wampus
Wampus
Luscar
8
10
12
22
23
24
26
10
18
21
24
25
26
1
1
2
3
4
28
18.3
22
15.7
20.5
6.3
26.8
6.5
5.9
7.8
5
4.3
4.4
6.6
5.8
3
4
4.6
4.7
7
94.12
100
82.35
100
5.59
100
1.72
5.65
10.77
6.9
2.88
5.3
2.95
4.76
18.84
0
4.1
25
19.23
23.5
64.3
47.1
42.1
6.6
100
1.7
7.26
1.54
6.9
2.88
5.3
1.85
3.81
14.49
0
3.28
20
0
4.4
3.6
66.7
2.1
1.6
0
4.3
7.26
9.23
20.69
21.58
6.82
1.11
3.81
72.46
100
7.58
70
76.9
97.1
100
52.9
100
2.7
100
0.9
3.23
3.08
1.72
0.72
3.03
1.85
3.81
11.59
100
0.61
12.5
0
C-34 Draft Document
-------
Table 2. Brook trout embryo-larval parameters collected from a high Se site (Luscar Creek), an
intermediate Se site (Gregg River), and reference site (Cold Creek) in northeastern Alberta over
three consecutive years.
Year Location Female # Se in egg, mg/kg %craniofaci %skeletal %finfold %edema
2000
2000
2000
2000
2000
2000
2000
2000
2000
2000
2000
2000
2000
2000
2000
2000
2000
2000
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
2001
Luscar
Luscar
Luscar
Luscar
Luscar
Luscar
Luscar
Luscar
Luscar
Luscar
Luscar
Cold
Cold
Cold
Cold
Cold
Cold
Cold
Cold
Cold
Cold
Cold
Cold
Luscar
Luscar
Luscar
Luscar
Luscar
Luscar
Luscar
Luscar
Luscar
Luscar
Luscar
Gregg
Gregg
Gregg
Gregg
Gregg
1
2
3
5
12
13
14
15
16
17
18
21
22
24
25
26
33
34
6
7
8
21
51
3
7
17
19
59
60
61
64
76
82
83
3
22
23
25
31
4.78
4.83
5.98
3.86
6.06
5.8
5.17
9.92
5.03
6.01
12.7
1.15
1.83
0.97
No data
0.59
1.35
2.18
1.79
1.36
0.94
1.07
1.09
8.4
7.26
14.6
9.79
5.8
9.03
7.29
7.08
7.1
6.06
5.82
7.08
7.95
9.23
6.46
7.35
15.38
38.06
7.39
25
16.77
4.06
4.13
16.22
5.61
9.44
14.34
3.26
4.83
1.67
3.31
3.45
6.15
6.45
0
1.61
1.36
0.43
0
0
1.35
2.22
7.55
2.28
3.16
0
1.54
36.71
1.11
6
6.32
0
0.5
0.56
0.51
0
1.49
3.03
5.7
1.83
1.42
0.49
0.54
0
5.83
0.72
1.48
1.38
0
1.1
4.83
0
0
0
0.69
0
0
2.13
0.93
1.62
0.63
2.11
0.46
0
0
2.19
13.29
0.22
2
1.58
0
0.5
0
1.7
0
3.73
0.34
8.77
0.7
0.2
0.36
0.54
0.27
0.83
0
0.89
1.38
0.72
1.66
6.9
1.54
0.81
0
0.46
0.27
0
0
0
0.81
0.32
2.42
0.91
1.05
9.09
0
19.65
0.88
5.6
20.53
1.08
2.51
0.56
0.17
15.38
1.49
0.5
4.82
0
0
0.12
0
0.27
1.11
0.36
0
0.69
0
1.1
0.69
0
0
0
1.38
0.54
0
6.38
0.46
0.27
0
0.3
0.46
1.05
0
0
1.16
0.44
0.8
1.58
0
0
0
0
-------
Table 2. Brook trout embryo-larval parameters collected from a high Se site (Luscar Creek), an
intermediate Se site (Gregg River), and reference site (Cold Creek) in northeastern Alberta over
three consecutive years (continued)
Year Location Female # Se in egg, mg/kg %craniofaci %skeletal %finfold %edema
2001
2001
2001
2002
2002
2002
2002
2002
2002
2002
2002
2002
2002
2002
2002
2002
2002
2002
2002
2002
2002
2002
2002
2002
Gregg
Gregg
Gregg
Luscar
Luscar
Luscar
Luscar
Luscar
Luscar
Luscar
Luscar
Gregg
Gregg
Gregg
Cold
Cold
Cold
Cold
Cold
Cold
Cold
Cold
Cold
Cold
32
33
34
17
23
26
38
42
44
54
56
25
37
39
32
26
2
5
29
23
48
42
22
51
4.91
7.02
5.01
6.28
5.27
6.36
18.9
4.95
6.47
7.96
18.8
6.27
4.58
6.67
0.42
0.89
0.94
1
1.02
1.2
1.25
1.6
1.74
2.11
7.21
1.88
0
1.7
7.34
1.81
0.9
2.79
0
0.33
3.99
1.23
2.99
3.57
0
0
0.96
0.25
0.72
0.35
9.52
0
0
2.17
0.48
1.88
0.37
12.74
0.46
0.52
0.54
0.44
0.25
0.33
0.75
1.23
0
1.19
0.6
0
0.32
0.5
1.09
0.35
4.76
0
0
2.17
3.37
4.38
0
0.85
0
0.26
0
0.15
0
0
0.5
0
0
1.19
0
0
0
0.25
0.36
0.35
2.38
0
1.09
0
0.48
0
0
0.21
0.46
0.26
0.18
0.15
0
0
0.75
0
0
1.19
0
0.29
0
0
0.72
0.35
0
0
1.09
2.17
C-36 Draft Document
-------
Table 3. Results of ANOVA comparing rainbow trout endpoints among sites
% fertilization
Site
Residuals
Df
3
51
Sum of Sq
77.60
20817.33
Mean Sq
25.8653
408.1829
F Value
0.06336703
Pr(F)
0.978935
% mortality
Site
Residuals
Df
3
51
Sum of Sq
3751.51
34510.50
Mean Sq
1250.504
676.676
F Value
1.848008
Pr(F)
0.1502207
% craniofacial deformities
Site
Residuals
Df
3
50
Sum of Sq
8093.97
30449.48
Mean Sq
2697.989
608.990
F Value
4.430272
Pr(F)
0.007732133
% skeletal deformities
Site
Residuals
Df
3
50
Sum of Sq
3279.30
19703.16
Mean Sq
1093.101
394.063
F Value
2.773923
Pr(F)
0.05094422
% finfold deformities
Site
Residuals
Df
3
50
Sum of Sq
6273.17
26886.93
Mean Sq
2091.056
537.739
F Value
3.888612
Pr(F)
0.01417887
% edema
Site
Residuals
Df
3
50
Sum of Sq
8902.51
43012.30
Mean Sq
2967.502
860.246
F Value
3.449597
Pr(F)
0.0233558
C-37 Draft Document
-------
Table 3. Results of ANOVA comparing rainbow trout endpoints among sites (continued)
Fry length
Site
Residuals
Df
3
50
Sum of Sq
5.0847
148.8246
Mean Sq
1.694896
2.976493
F Value
0.5694271
Pr(F)
0.6377436
Fry weight
Site
Residuals
Df
3
48
Sum of Sq
1721.104
7726.859
Mean Sq
573.7012
160.9762
F Value
3.563888
Pr(F)
0.02080915
Table 4. Rainbow trout means (standard deviation) for measurements made in eggs, embryos and
larvae spawned from fish collected at exposed sites (Luscar and Gregg Creeks) and reference sites
(Deerlick and Wampus Creeks).
Parameter
egg Se, mg/kg ww
fertilization, %
mortality, %
craniofacial, %
skeletal, %
finfold, %
edema, %
larval length, mm
larval weight, mg
Site
Luscar Cr.
9.93 (6.77)
77.8 (20.3)
35.0(29.5)
33.3 (37.2)
25.0(27.9)
15.0(27.1)
34.5 (40.3)
18.5 (2.0)
53.3(16.3)
Gregg Cr.
6.52(4.11)
81.2(12.7)
34.2 (32.5)
10.6 (6.5)
9.9 (5.8)
13.6(15.2)
12.2(12.3)
19.4(1.6)
44.6 (10.4)
Deerlick Cr.
3.49(1.90)
77.5 (20.9)
18.1 (14.6)
7.1(6.1)
9.2(12.3)
5.4 (6.2)
6.1 (7.3)
19.0(1.5)
41.2(9.3)
Wampus Cr.
3.5 (1.09)
77.5 (24.1)
37.3 (34.5)
12.0 (9.6)
7.9 (8.2)
42.2 (43.7)
23.3 (37.8)
19.2 (0.9)
40.6 (8.4)
C-38 Draft Document
-------
Table 5. Brook trout means (standard deviation) for measurements made in eggs, embryos and
larva spawned from fish collected at exposed sites (Luscar and Gregg Creeks) and
reference site (Cold Creek).
Parameter
egg Se, mg/kg ww
fertilization, %
mortality, %
craniofacial, %
skeletal, %
finfold, %
edema, %
larval length, mm
larval weight, mg
Site
Luscar Cr.
7.78 (3.80)
92.8 (7.2)
6.5 (8.9)
7.9(10.1)
2.0 (3.3)
1.9(4.1)
1.0(2.9)
17.4(1.1)
31.7(8.6)
Gregg Cr.
6.59(1.39)
78.4(18.2)
2.9(2.3)
2.3 (2.5)
0.8 (0.7)
3.1(6.0)
0.3 (0.6)
17.9(0.9)
31.3(5.4)
Cold Cr.
1.26(0.47)
89.1(19.6)
6.9(12.1)
2.1(2.6)
1.0(1.4)
0.9(1.5)
0.7(1.4)
18.5(1.2)
37.8 (7.2)
Table 6. Results of ANOVA comparing brook trout endpoints among sites
% fertilization
site
Residuals
% mortality
site
Residuals
df
2
60
df
2
60
Sum of Sq
1683.3
12906.4
Sum of Sq
131.4
5433.6
% craniofacial deformities
site
Residuals
df
2
60
Sum of Sq
519.1
3150.6
Mean Sq
841.67
215.11
Mean Sq
65.72
90.56
Mean Sq
259.54
52.51
F Value
3.9128
F Value
0.7257
F Value
4.9427
Pr(F)
0.0253
Pr(F)
0.4882
Pr(F)
0.0103
-------
Table 6. Results of ANOVA comparing brook trout endpoints among sites (continued)
% skeletal deformities
site
Residuals
df
2
60
% finfold deformities
site
Residuals
% edema
site
Residuals
Fry length
site
Residuals
Fry weight
site
Residuals
df
2
60
df
2
60
df
2
60
df
2
60
Sum of Sq
19.2
367.6
Sum of Sq
37.5
895.1
Sum of Sq
4.6
280.6
Sum of Sq
16.1
73.9
Sum of Sq
546.2
3512.9
Mean Sq
9.58
6.13
Mean Sq
18.74
14.92
Mean Sq
2.32
4.68
Mean Sq
8.04
1.23
Mean Sq
273.10
58.55
F Value
1.5631
F Value
1.2562
F Value
0.4966
F Value
6.5265
F Value
4.6644
Pr(F)
0.2179
Pr(F)
0.2921
Pr(F)
0.6110
Pr(F)
0.0027
Pr(F)
0.0131
C-40 Draft Document
-------
Figure 1. Rainbow trout percent normal (100 - % craniofacial deformities) as a
function of the logarithm of selenium concentration in eggs (Exposure Variable).
Untransformed values reported in mg Se/kg tissue wet weight. The curve represents
projections from the fitted logistic equation.
120 r
Log(Se in eggs ww)
Figure 2. Rainbow trout percent normal (100 - % skeletal deformities) as a
function of the logarithm of selenium concentration in eggs (Exposure Variable).
Untransformed values were reported in mg Se/kg tissue wet weight. The curve
represents projections from the fitted logistic equation.
100(,
40 •
.4 .6 .8 1.0
Log(Se in eggs ww)
C-41 Draft Document
-------
Figure 3. Plot of percent abnormal for craniofacial, skeletal and finfold deformities and edema
against selenium concentration in brook trout eggs ww, 2000 and 2001 data.
g. 40 -
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Egg Se Concentration
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C-42 Draft Document
-------
The effect levels determined using the within streams approach resulted in values based on ww in eggs.
The primary tissue for which the reproductive effect levels were based, eggs, was converted from ww to
dw using the average percent moisture of 61.2% for rainbow trout eggs reported by Seilor and Skorupa
(2001).
Chronic Values: Brook trout: between streams approach
No effects at ECio level at 7.78 mg Se/kg eggs ww or 20.5 mg Se/kg eggs dw;
egg. Chronic value is >20.5 mg Se/kg eggs dw.
Rainbow trout: Within streams approach
ECio value (skeletal deformities) at 8.2 mg Se/kg egg ww or 21.1 mg Se/kg egg
dw. Chronic value is 21.1 mg Se/kg eggs dw.
C-43 Draft Document
-------
Kennedy, C.J., L.E. McDonald, R. Loveridge, M.M. Strosher. 2000. The effect of bioaccumulated
selenium on mortalities and deformities in the eggs, larvae, and fry of a wild population of cutthroat trout
(Oncorhynchus clarki lewisi). Arch. Environ. Contam. Toxicol. 39:46-52.
Test Organism:
Exposure Route:
Study Design:
Effects Data :
Cutthroat trout (Oncorhynchus clarki lewisi; spawning adults, 3-6 years)
Dietary and waterborne - field exposure
Total selenium concentrations measured at the time the eggs were taken were
<0.1 ng/L from the reference site and 13.3 to 14.5 |o,g/L at the exposed site.
At reference and exposed site (Fording River, BC, Canada which receives
drainage from open-pit coal mining), eggs were stripped from females (n=20
from reference site; n=17 from exposed site) and fertilized from milt from one
male collected at each site. Fertilized eggs were reared in well water and
examined for time to hatch, deformities (craniofacial, finfold, skeletal and yolk
sac malformations), and mortalities. Inspection of deformities in eggs was
performed using 40X magnification.
No significant correlations between the selenium concentrations in the eggs from
either site and: hatching time (reference, 25.5-26.5 days; exposed, 22-25.5 days);
percent deformities preponding (reference, 0-2.4%; exposed, 0-0.34%); percent
deformities after ponding (reference, 0-0.26%; exposed, 0-0.09%); percent
mortalities preponding (reference, 1.5-70.3%; exposed, 1-100%); percent
mortalities after ponding (reference, 0.3-4.3%; exposed, 1.5-43.7%); total percent
mortalities (reference, 2.8-55.8%; exposed, 3.7-100%). The average selenium
residues in tissues were as follows:
Site
Reference
Exposed
Adult fish liver, mg Se/kg
dw
8.2; Range: 3.4-14.6
36.6; Range: 18.3-1 14
Adult fish muscle, mg Se/kg
dw
2.4; 1.4-3.8
12.5; Range: 6.7-41
eggs, mg Se/kg dw
4.6
21.2
Chronic Value:
>21.2 mg Se/kg dw in eggs
>12.5 mg Se/kg dw in muscle
C-44 Draft Document
-------
Hardy, R.W. 2005. Effects of dietary selenium on cutthroat trout (Oncorhynchus clarki) growth and
reproductive performance. Report for Montgomery Watson Harza. December 14, 2005.
Test Organism:
Exposure Route:
Test Duration:
Study Design:
Effects Data:
Chronic Value:
Cutthroat trout (Oncorhynchus clarki, 0.9 g)
Dietary only
Six experimental dietary treatments were produced by cold extrusion. The
formulation of the diet was designed to be similar to commercial trout diets and
had a proximate composition of 45% protein and 16% lipid. Seleno-methionine
diluted in distilled water (100 ug/L) was added in appropriate volumes to each
batch of feed to facilitate pelleting. Measured dietary selenium concentrations
were 1.2 (control), 3.8, 6.4, 9.0, 11.5, and 12 mg Se/kg dw. Fry were fed initially
at a rate of 10 times per day 6 days each week to apparent satiation. Feeding
frequency decreased as fish grew.
124 weeks (865 days, 2.5 yrs)
Groups of 50 fish were placed into triplicate tanks (145 L) receiving 4-15 L/min
of hatchery water at 14.5EC and fed one of the six experimental diets. The fish in
each tank were bulk-weighed and counted every 14 days for the first 12 weeks of
the experiment, and then every 4 weeks until 48 weeks. Samples offish for
whole-body selenium analysis were taken at each sampling date for the first 12
weeks followed by every 3 months thereafter. After six months of feeding, the
fish were transferred to 575 L tanks and the number of replicate tanks per dietary
treatment was reduced to two. After 80 weeks of feeding, the fish were
transferred to 1050 L outdoor tanks each supplied with 70 L/min of constant
temperature (14.5EC) spring (hatchery) water. After 2.5 years of the feeding trial,
fish were spawned and whole body selenium level, egg selenium level, % eyed
eggs, % hatched eggs, and % deformed larvae were examined.
No signs of toxicity (reduced growth or survival relative to controls) were
observed in fish fed the highest dietary selenium treatment (12 mg Se/kg dw)
after the first 80 weeks of exposure just prior to transfer outdoors. No signs of
clinical disease were evident, and no relationship was found between feed
conversion ratios and the level of selenium added to the feed. Average whole
body selenium levels of female Henry's Lake cutthroat trout at spawning at 2.5 to
3 years of age were 5.87, 9.10, 11.37 and 5.61 mg Se/kg dwinthe four highest
dietary treatments. Average egg selenium levels in the same four dietary
treatments were 6.61, 5.05, 5.18, and 16.04 mg Se/kg dw. Percent survival from
the eyed stage to hatching varied among treatment groups, with the control and
the highest Se dietary treatment having the second highest survival (85%) and the
fifth dietary treatment group the highest (93%). Percent deformed larvae ranged
from a low of 5.6% in controls to a high of 20.2% in the 6.4 mg Se/kg dw dietary
treatment group; larvae in the two highest dietary treatment groups only
exhibited 7 and 6.8 %, respectively.
The chronic value for embryo/larval deformity is a NOAEC of >11.37 mg Se/kg
dw whole-body parent tissue and >16.04 mg Se/kg dw egg.
C-45 Draft Document
-------
Rudolph, B-L, I. Andreller, CJ. Kennedy. 2008. Reproductive success, early life stage development,
and survival of Westslope cutthroat trout (Oncorhynchus clarki lewisi) exposed to elevated selenium in an
area of active coalmining. Environ. Sci. Technol. 42: 3109-3114.
Test Organism:
Exposure Route:
Test duration:
Study Design:
Effects Data:
Westslope cutthroat trout (Oncorhynchus clarki lewisi)
Field collected.
In June, 2005, eggs were collected from 12 females from Clode Pond (exposed
site) and 16 females from O'Rourke Lake (reference site). Milt was obtained
from 3-5 males at each site. Clode Pond is on the property of Fording River Coal
Operations in Southeast British Columbia with reported selenium concentrations
of 93 (ig/L. O'Rourke Lake is an isolated water body into which Westslope
cutthroat trout were stocked in 1985, 1989 and 1992 and has selenium levels
reported <1 (ig/L.
Through the end of yolk sac absorption (at swim-up) by the alevins.
Individual batches of eggs were fertilized in the field with 2 ml composites of
milt. Water-hardened eggs were transported to the rearing laboratory. Eggs and
alevins were monitored daily for fertilization, hatching and mortality. After the
yolk sacs were absorbed, alevins were sacrificed and preserved in Davidson's
solution.
All viable fry (n = 4922) after yolk absorption were observed for the frequency
and severity of skeletal (lordosis, kyphosis, and scoliosis), craniofacial (head,
eyes or jaw), and fin malformations as well as edema. The authors used a
graduated severity index (GSI) for deformities in which fry were scored 0
(normal) to 3 (severe) based on the level of defect.
Eggs with the four highest Se concentrations (86.3 to 140 mg/kg dw) collected
from Clode Pond fish died before reaching the laboratory (Table 1). Excluding
the eggs that died from females CP1, CP3, CP4 and CP5, fertilization (total eggs
reaching the eyed stage/total eggs x 100) was not related to Se concentrations in
the eggs (Table 1). The percent of alevins (post hatch to swim-up stage) that died
was related to the selenium concentration in the eggs; the ECi0 estimated by
TRAP is 24.1 mg Se/kg dw Figure 1). Note: The data used in the TRAP analysis
excluded the variable from OL1. These are data from the reference lake in which
only 57% of the larvae survived. Alevin survival was meaningfully higher in the
other 15 clutches of eggs from the reference site (87.3 to 99.8%). Analysis of
combined egg and alevin survival resulted in a similar ECio estimate. The
selenium in muscle data was not amenable for analysis with alevin survival using
TRAP. ECio and EC2o estimates for muscle were derived using a least squares
regression of the egg and muscle data reported by Rudolph et al.
[Semuscle]= 4.0853 +0.391 [Seegg]
(R2 = 0.9094)
Deformity analysis was not performed on the alevins that died prior to the swim-
up stage. Therefore, due either to dead eggs or dead alevins, the occurrence and
C-46 Draft Document
-------
severity of deformities were assessed on four clutches of eggs from Clode Pond
(CP2, CP6, CPU and CP12) with a range of 11.8 to 20.6 :g Se/g dw and 15 of
the 16 clutches (all eggs died in OL8) from O'Rourke Lake (Table 1). There was
no correlation between egg Se concentration and frequency of deformity or
edema. Statistical differences between sites were observed (p < 0.05) for skeletal
deformities and edema for both the frequency of the occurrence and the severity
score (Table 2). Note: the percent and severity score of skeletal deformities were
greater in the reference site than in the exposed site.
The effect level for this study was based on the alevin mortality data and not the
deformity measurements. Although edema occurred statistically more often at the
exposed site (87.7% at Clode Pond, 61.2% at O'Rourke Lake), it was not
correlated with selenium levels in the eggs. Also the greater occurrence of
skeletal malformations in the reference site confounded the use of statistical
differences between sites to determine effect levels for this study.
Effect Concentration: 24.11 mg Se/kg dw in eggs; 13.51 mg Se/kg dw in muscle
Table 1. Fertilization, egg mortality and alevin mortality for offspring from individual fish
collected in Clode Pond and O'Rourke Lake.
Fish ID
Clode Pond
(exposed site)
CP1
CP2
CP3
CP4
CP5
CP6
CP7
CP8
CP9
CP10
CPU
CP12
avg
SD
Muscle [Se]
mg/kg dw
38.8
11.8
40.4
46.1
50.4
34.7
39
7
35.4
35.5
11.3
13.4
30.3
15.1
Egg [Se] Dead eggs, Dead alevins,
mg/kg dw Fertilization, % % %
88.3
16.1
86.3
121
140
51
65.3
11.8
46.8
75.4
16.9
20.6
61.6
42.4
0
99.7
0
0
0
99
97.2
73.7
91.3
88.2
79.2
98.6
61
45
100
1.8
100
100
100
7.4
8.9
36.1
36.6
17.6
22.1
3
44
42
NA
0.9
NA
NA
NA
92.6
91.1
0.8
63.2
82.4
1.3
5.1
42
44
O'Rourke Lake
(reference site)
OL1
OL2
OL3
OL4
OL5
OL6
OL7
OL8
8.28
7.7
8.16
8.03
8.12
6.61
8.52
7.22
12.9
13.9
12.5
15
14.9
15.2
12.9
12.3
100
93.1
99.4
98.2
89.3
76
99.4
30.5
28.6
53.1
3.9
14.5
19.3
32
2.1
100
42.9
6.9
2.4
12.7
5.3
4
0.2
NA
C-47 Draft Document
-------
Fish ID
OL9
OLIO
OL11
OL12
OL13
OL14
OL15
OL16
avg
SD
Muscle [Se]
mg/kg dw
7.25
7.64
8.74
8.2
7.86
8.5
7.62
8.13
7.9
0.6
Egg [Se]
mg/kg dw Fertilization, %
Dead eggs, Dead alevins,
16.7
13.1
15.6
13.9
15.1
13.1
12.3
12.7
13.9
1.4
96.4
99.1
96.2
99.1
92.6
79.5
92.4
71
88
18
12.8
2.5
10.8
16.4
25.9
22.2
11.8
45.2
25
25
4.5
5.5
2.4
3
2.8
0.5
2.6
4.8
7
10
Table 2. Deformity results (frequency and severity) for offspring from O'Rourke Lake and Clode
Pond. Values are presented as mean ± SE. * indicates a significant difference (p < 0.05)
between means from the two sites.
Frequency of deformity, %
Skeletal*
Craniofacial
Pinfold
Edema*
Severity of deformity, score
Skeletal*
Craniofacial
Pinfold
Edema*
O'Rourke Lake
37.4 ±3.6
10.2 ±2.0
10.6 ±3.1
61.2±4.9
0.47 ±0.07
0.12 ±0.03
0.15 ±0.05
0.61 ±0.05
Clode Pond
16.5 ±2.2
5.7 ± 1.0
7.5 ±3. 84
87.7 ±2.0
0.18 ±0.02
0.06 ±0.01
0.09 ±0.05
0.88 ±0.02
C-48 Draft Document
-------
Figure 1. Survival of Westslope cutthroat trout alevin as a logistic function of the logarithm of the
selenium concentration in eggs.
110
100
90
80
£
_. 70
CO
> 60
1 -
fc 40
<
30
20
10
0
1.1 1.2 1.3 1.4 1.5 1.6
Log(Egg [Se] pg/g dw)
1.7
1.8 1.9
2.0
TRAP Output
Parameter summary
Parameter
Log X50
S
YO
Initial Est.
1.35
3.0
96.35
Final Est.
1.5885
2.663
97.59
Std Error
0.0345
0.694
2.17
95% LCL
1.516
1.209
93.04
95% UCL
1.661
4.116
102.13
Effect concentration summary
p
50
20
10
5
1
Xp estimate
38.77
28.73
24.11
20.51
14.36
95% LCL
32.83
20.95
16.03
12.51
7.22
95% UCL
45.78
39.39
36.26
33.63
28.53
C-49 Draft Document
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Nautilus Environmental. 2011. Evaluation of the Effects of Selenium on Early Life Stage Development
of Westslope Cutthroat Trout from the Elk Valley, BC. Report to Elk Valley Selenium Task Force,
November 24, 2011.
Test Organism:
Exposure Route:
Test Duration:
Study Design:
Effects Data:
Westslope Cutthroat Trout (Oncorhynchus clarki lewisi)
Field collected. Adult fish were collected and spawned from lentic and lotic
environments in areas proximate to Teck Coal's Fording River Operations. Eggs
were also obtained from fish collected from Connor Lake, a lake located within
the Elk valley watershed not exposed to mine discharges and considered a
reference site and a methodological control.
Fertilized eggs were reared in the laboratory until they reached swim-up fry
stage. A subset of fry surviving at swim-up were reared for an additional 28 days.
Gametes were stripped from the ripe adults in the field during June and July 2008
and transported immediately to the laboratory in coolers containing wet ice. Eggs
were fertilized in the laboratory. After stripping the eggs, female fish were
sacrificed and the whole body stored on ice for later Se analysis. For a given
female, approximately 240 fertilized eggs were divided into four replicates of 60
eggs. In cases when fewer eggs were available three replicates of 60 eggs were
used. If less than 180 eggs were available, either 3 or 4 replicates of 30 were
used. Females with less than 90 eggs were not used. The fertilized eggs were
maintained in the laboratory until the fry reached swim-up at which point
deformities were assessed. Survival was also assessed up to swim-up. In test
chambers in which there were at least 40 surviving fish at swim-up, one-half of
the surviving fish were maintained for an additional 28 days. Survival, length,
weight and deformities were assessed in the 28-day post swim-up test.
The number, type and severity of deformities were measured at swim-up and at
the end of the 28-day post swim-up test. Deformity assessments were conducted
on recently killed fresh fish to avoid artifacts caused by preservation. A
graduated severity index (GSI) was assigned to each of four types of
deformity/abnormality: skeletal, craniofacial, finfold and edema. Graduated
Severity Index (GSI) methods followed those described in Holm et al. (2003) and
Rudolph etal (2006; 2008).
Survival of the larvae from hatch through swim-up spawned from the four fish
collected from the reference site, Connor Lake, ranged from 73 to 92% (egg Se
4.32 to 7.31 mg/kg dw) (Table 1). Larval survival at swim-up was also generally
high for fish collected in the Se exposed sites up to egg Se concentration 29.6
mg/kg dw (Table 1, Figure 1). Larvae exposed above this egg Se concentration
had poor to no survival. Larvae from one fish (POOS 11) below this threshold did
have poor survival (11.7%). The authors noted that the many of the eggs from
this fish displayed an unusual distribution of lipid vesicles which resulted in
greater than 50% mortality in the first 24 hours due to egg breakage. The
remaining eggs may have been compromised due to the organic material released
during the egg breakage.
The rate of deformities in larvae at swim-up showed no relationship with Se in
egg through 29.6 mg/kg dw (Table 2).
C-50 Draft Document
-------
The results of the 28-day post swim-up test showed no relationships between
larval survival or deformities and egg Se (Table 3). The authors also measured
the length and weight of larvae at the end of the 28 day test; neither of which
showed a relationship with egg Se concentration.
Se Tissue Concentrations. Two analytical laboratories (A and B) measured Se in
the eggs. The mean difference in egg Se concentrations between the two
laboratories was 34.2%. To better understand the difference between the two
laboratories, five egg samples (i.e., from five different fish) from this study were
sent to both laboratories in 2010. Both laboratories digested the eggs using the
methods they used in their own 2008 original analysis. The respective digestates
were split and then shared between laboratories. Both labs then measured
selenium in their own digestates and the digestate received from the other lab.
The results of this follow-up study showed that when each lab used their own
digestion procedures Laboratory A had on average 43% higher measurements in
the 2008 analysis and 23% higher in the follow-up 2010 analysis. When each lab
measured selenium using the same digestate the difference in the Se
measurements between labs was on average only 1 to 8%. The authors concluded
that although both laboratories employed acceptable and approved practices,
Laboratory A used a more efficient digestion process resulting in higher Se
measurements. To compensate for the reduced Se measurements in Laboratory B,
its values were increased by 34.2%. The measurements made by Laboratory A
are marked in Table 1; unmarked values are Laboratory B measurements
increased by 34.2%.
Effect Concentration: The most sensitive endpoint determined by TRAP was larval survival at swim-
up. TRAP was used to model larval survival with the entire egg Se dataset that
r^ oo
included egg Se measurements from Laboratory A and adjusted measurements
from Laboratory B (EC 10 = 26.6 mg/kg egg dw; Figure 1) and using only the egg
Se measurements from Laboratory A (Figure 2). Because the Laboratory A
dataset estimated slightly lower EC values, the EC 10 of 24.02 mg/kg egg dw is
the selected effect concentration for this study.
C-51 Draft Document
-------
Table 1. Summary of westslope cutthroat trout larvae surviving to swim-up per parent female (fish
ID) including location of collection of parent female and concentration of selenium in the eggs.
Proportion surviving
Fish ID
YO93
CL1
R082
CL4
CL2
CL3
P00815
R026
P00823
R039
R086
R077
R042
R055
R043
R074
POOS 11
P00809
P00803
R078
GO99
O087
O085
WO52
R069
R071
WO94
UT101
Se egg, Replicate
Location mg/kg dw s
Lentic
Referenc
e
Lotic
Referenc
e
Referenc
e
Referenc
e
Lotic
Lotic
Lotic
Lotic
Lotic
Lotic
Lotic
Lotic
Lotic
Lotic
Lotic
Lotic
Lotic
Lotic
Lotic
Lentic
Lentic
Lentic
Lotic
Lotic
Lentic
Lentic
3.88*
4.32
5.21
5.96*
6.82
7.31
7.6
12.53
12.71
12.9
13.4*
14.29
16.44
16.5
16.85
17.8*
19.25
19.72
24.8*
29.61
34.2*
54.7*
56.8*
61.1*
65.61
72.9
73.1
74.67
4
4
3
4
4
4
3
4
4
4
4
3
3
4
4
4
1
4
4
4
4
4
4
4
4
4
4
4
Replicat
e mean
0.8125
0.9167
0.9056
0.7333
0.8333
0.8542
0.8222
0.5792
0.8875
0.6042
0.9417
0.6444
0.8
0.8792
0.8667
0.9375
0.1167
0.7667
0.9375
0.8825
0.2083
0.07083
0
0
0
0
0
0
Replicat
e min
0.6667
0.8833
0.8333
0.6
0.7
0.8167
0.7167
0.5
0.85
0.55
0.85
0.6167
0.7
0.7833
0.7667
0.8833
0.1167
0.65
0.9333
0.8333
0.1667
0.01667
0
0
0
0
0
0
Number Total
Replicat survivor numbe
e max s r
0.9167
1
0.95
0.8
0.9167
0.8833
0.95
0.65
0.95
0.65
0.9833
0.6667
0.9
0.9667
0.9667
0.9833
0.1167
0.8833
0.95
0.9333
0.2667
0.2
0
0
0
0
0
0
195
220
163
176
200
205
148
139
213
145
226
116
72
211
104
225
7
184
225
105
50
17
0
0
0
0
0
0
240
240
180
240
240
240
180
240
240
240
240
180
90
240
120
240
60
240
240
119
240
240
240
240
240
240
240
240
*Laboratory A dataset
C-52 Draft Document
-------
Table 2. Summary of westslope cutthroat trout larval deformities to swim-up per parent female
(fish ID) including location of collection of parent female and concentration of selenium in the eggs.
Fish ID
YO93
CL1
R082
CL4
CL2
CL3
P00815
R026
P00823
R039
R086
R077
R042
R055
R043
R074
P00809
P00803
GO92
R078
GO99
Location
Lentic
Lentic
Lotic
Lentic
Lentic
Lentic
Lotic
Lotic
Lotic
Lotic
Lotic
Lotic
Lotic
Lotic
Lotic
Lotic
Lotic
Lotic
Lotic
Lotic
Lotic
Se egg, Skeletal Craniofacial Finfold Edema Deformities
mg/kg dw combined combined combined combined combined
3.88*
4.32
5.21
5.96*
6.82
7.31
7.6
12.53
12.71
12.9
13.4*
14.29
16.44
16.5
16.85
17.8*
19.72
24.8*
26.1
29.61
34.2*
4.5%
7.6%
1.2%
4.3%
11.1%
5.0%
0.0%
2.1%
1.9%
2.1%
2.7%
1.7%
1.2%
0.0%
0.9%
2.7%
3.9%
2.7%
0.0%
1.8%
14.5%
0.9%
1.9%
1.3%
7.3%
3.7%
2.0%
2.7%
2.1%
2.9%
1.9%
1.0%
10.4%
0.0%
2.8%
2.6%
1.8%
2.8%
0.9%
1.9%
0.0%
53.9%
4.4%
1.0%
2.5%
1.7%
0.8%
1.0%
0.0%
0.7%
1.8%
2.9%
0.0%
0.9%
0.0%
1.0%
1.8%
0.9%
3.3%
0.0%
1.9%
1.0%
6.8%
1.9%
1.0%
0.0%
0.7%
3.0%
0.0%
2.9%
1.4%
5.6%
4.9%
0.0%
12.2%
2.6%
2.9%
1.7%
0.9%
4.7%
0.9%
4.4%
2.9%
28.2%
7.7%
9.5%
3.7%
12.6%
15.9%
7.0%
5.6%
2.1%
7.4%
9.9%
2.7%
15.5%
2.6%
4.7%
4.4%
3.6%
9.0%
4.5%
4.4%
5.7%
64.7%
*Laboratory A dataset
C-53 Draft Document
-------
Table 3. Summary of larval survival and rates deformities after the 28-day post swim-up test per
parent female (fish ID) including location of collection of parent female and concentration of
selenium in the eggs.
Fish ID
CL1
CL2
CL3
CL4
Y093
R082
P00815
P00823
R086
R077
R055
R074
P00809
P00803
Location
Reference
Reference
Reference
Reference
Lentic
Lotic
Lotic
Lotic
Lotic
Lotic
Lotic
Lotic
Lotic
Lotic
Sample
size (n)
112
93
96
68
93
71
69
105
112
36
101
106
65
108
EggSe
(mg/kg dw)
4.3
6.8
7.3
6
3.9
5.2
7.6
12.7
13.4
14.3
16.5
17.8
19.7
24.8
Survival
(%)
99.1
99
91.7
98.6
95.6
87.4
91.1
96.3
97.2
92.4
95.9
93.1
91.7
95.7
Skeletal
(%)
0
0
0
0
0
0
0
0
0
2.8
0
0
0
0
Craniofacial
(%)
0
0
1
0
0
2.9
1.2
0
0.9
2.8
4.6
0
0
0
Pinfold
(%)
0
0
1
4.3
2
0
1.4
0
0
2.8
0
0
0
1
Total
(%)
0
0
2
4.3
2
2.9
2
0
0.9
4.2
4.6
0
0
1
C-54 Draft Document
-------
1.1
1.0
.9
T3
CD
-g
CO
o
o
Q.
0
ol
.7
.6
.5
.4
.3
.2
.1
n
\
*" \
\
\
• \
* \
i i i i i i
.4
.6
1.0 1.2 1.4
Log(Se egg), mg/kg dw
1.6
Figure 1. TRAP 1.20 Analysis type - Tolerance distribution; Model option - Triangular distribution
(3 parameters). Includes Laboratories A and B datasets.
Xp Estimates
p
50
20
10
5
0
Xp Estimate
35.992
29.449
26.616
24.779
20.850
95% LCL
34.831
28.128
25.229
23.356
19.376
95% UCL
37.192
30.832
28.080
26.289
22.437
C-55 Draft Document
-------
1.1
1.0
1
tn
0
0
Q.
o
ol
.7
.6
.5
.4
.3
.2
.1
n
\
\
• \
.4
.6
1.0 1.2 1.4
Log(Se egg), mg/kg dw
1.6
1.5
Figure 2. TRAP 1.20 Analysis type - Tolerance distribution; Model option - Triangular distribution
(3 parameter). Includes Laboratory "A" dataset only. *
Xp Estimates
p
50
20
10
5
0
Xp Estimate
33.75
26.92
24.02
22.16
18.237
95% LCL
32.56
25.54
22.55
20.65
16.686
95% UCL
34.99
28.37
25.57
23.77
19.933
* Although some scientists have attempted to explain certain occurrences of improved response with
increasing concentration in terms of nutrient selenium sufficiency-deficiency, the concentrations involved
in this study are too high to for selenium deficiency to be an explanation. The figure's apparent bi-phasic
measured response is thus best explained as being a chance outcome of noise.
C-56 Draft Document
-------
Colder Associates. 2009. Development of a Site-specific Selenium Toxicity Threshold for Dolly Varden
Char. Report to Northgate Minerals Corporation, PO Box 3519, Smithers, British Columbia. Report
Number 04-1421-101/2000.
Test Organism:
Exposure Route:
Test duration:
Study Design:
Effects Data:
Dolly Varden (Salvelnius malma)
Field collected.
Adult Dolly Varden char were collected from reference (North Kemess Creek),
high Se exposure (Upper Waste Rock Ponds and Creek) and moderate Se
exposure (lower Waste Rock Creek) sites during September 22 to 24, 2008. Eggs
were stripped from females and fertilized with milt from males collected from the
reference site. Fertilized eggs were taken to the laboratory for testing.
The test was terminated when 90% of the larvae reached swim-up, approximately
5 months after fertilization.
Approximately 30 fertilized eggs were added to each replicate rearing container.
The number of replicates per female parent ranged from one to four depending
on the number of eggs available. Embryos were maintained in 4 L containers
with 3.5 L dechlorinated tap water in a static-renewal system (3 renewals
times/week) at 5°C. The condition of the embyos and alevins were observed
daily and any dead individuals were counted and removed. Test termination
occurred over a 3-day period during February 11 to 13, 2009. The hatched larvae
were sacrificed using an overdose of the anesthetic, clove oil. Individual length
and weight were measured on each fry, and deformity analysis was performed on
fresh unpreserved larval fish using 40X magnification.
A graduated severity index (GSI) was used for deformity assessment (skeletal,
craniofacial, and finfold as well as edema). The narrative criteria were the same
as used by Holm et al. (2005) and Rudolph et al. (2008).
Alevin survival was not related to Se concentration in the eggs (Table 1). Almost
all of the mortality occurred during the egg stage. Only 4 alevins died during the
study, 1 from Fish #19 and 3 from Fish #2, both females collected at an exposed
site. The prevalence of deformities increased sharply after the selenium egg
concentration exceeded 50 mg/kg dw (Table 1, Figure 1). The proportion of
Dolly Varden larvae with any type of deformity (skeletal, craniofacial, and
finfold as well as edema) as a function of the log of the selenium concentration in
the eggs using TRAP (logistic equation) produced an ECi0 value of 56.22 mg/kg
dw eggs (Figure 1).
C-57 Draft Document
-------
Table 1. Selenium concentration in the eggs of Dolly Varden char and the survival of alevins to the
swim-up stage and the proportion of larvae without any type of deformity.
Fish
#
1
2
5
6
15
19
9
12
17
Sample
ID
WRC-
F105
WRC-F61
WRC-
F103
WRC-F83
WRC-
F104
WRC-F86
NK-F30
NK-F29
NK-F21
Location
Waste Rock Creek
Waste Rock Creek
Waste Rock Creek
Waste Rock Creek
Waste Rock Creek
Waste Rock Creek
North Kemess Creek
North Kemess Creek
North Kemess Creek
[Se]
eggs
mg/kg
dw
56.6
65.8
32.6
51.9
56.3
60.5
11
10.5
5.4
Survival of eggs to
up
Initial
120
120
29
120
60
120
30
46
90
End
71
81
29
115
48
115
1
15
86
swim-
%
59
68
100
96
80
96
3
33
96
Proportion of
larvae
without any
type of
deformity
0.89
0.58
0.97
0.97
0.90
0.72
a
1.00
0.91
SCD1 Redd#l
SCD2 Redd #2
Southern Collection
Ditch 10.3 30 18 60
Southern Collection
Ditch 24.7 40 32 80
1.00
1.00
C-5 8 Draft Document
-------
Figure 1. Proportion of Dolly Varden alevin without any type of deformity as a logistic function of
the logarithm of the selenium concentration in eggs (TRAP).
1.2r
O
o
CD
03
O
C
O
O
Q.
O
.6
r =0.93 3
1.0 1.2 1.4 1.6
Iog10 Egg Se (mg/kg dw)
2.0
Guess
Final
SE 95% LCL 95% UCL
LogXSO
Slope
YO
Total
Model
Error
1.844
4.152
0.975
ECx
50
20
10
5
1
DF
9
2
7
1.829
6.963
0.980
EC
67.42
60.12
56.22
52.85
46.11
SS
1.74E-01
1.63E-01
1.11E-02
0.007
1.252
0.017
95% LCL
64.92
57.96
53.00
48.64
40.13
MS
1.93E-02
8.13E-02
1.58E-03
1.812
4.003
0.939
95% UCL
70.01
62.35
59.64
57.43
52.99
F
51.429
1.845
9.924
1.021
P
0.99993
C-59 Draft Document
-------
AECOM. 2012. Reproductive success study with brown trout (Salmo trutta). Data quality assurance
report. Final. December 2012.
Formation Environmental. 2011. Brown Trout Laboratory Reproduction Studies Conducted in Support
of Development of a Site-Specific Selenium Criterion. Prepared for J.R. Simplot Company by Formation
Environmental. Revised October 2011.
Test Organism:
Exposure Route:
Test duration:
Study Design:
Brown trout (Salmo trutta)
Field collected.
Adult female and male brown trout were collected at three field sites from two
streams downstream of the Smokey Canyon mine. In addition, brown trout eggs
were obtained from two hatcheries as method controls.
Embryo-larval monitoring to 15 days post swim-up.
Eggs were collected from 26 ripe female brown trout at three field sites
downstream of the Smokey Canyon mine. These included one site on the highly
impacted Sage Creek (LSV2C) as well as two sites along Crow Creek (CC-150
and CC-350) downstream of the conflux with Sage Creek. The downstream -
most station along Crow Creek (CC-150) was intended to be a field control. Eggs
were fertilized in the field with milt collected from males collected at the same
site as females. Fertilized eggs were water hardened at the site using stream
water, then placed in oxygenated plastic bags and stored on ice in the dark
(cooler) for transportation to laboratory. Se was measured in adult fish (whole
body) and in eggs of field collected females. In addition, eggs were collected
from 8 ripe females obtained from the Saratoga National Fish Hatchery (SC) to
serve as method controls. Similar to field-caught fish, SC hatchery females were
stripped of eggs and fertilized by milt from males obtained from the same
hatchery. As a result of lower than expected hatch rates and fungal contamination
in some SC hatchery samples, additional hatchery fish were obtained (as already
fertilized eyed embryos) from the Spring Creek Trout Hatchery (SPC), which
were divided into four treatments.
Approximately 600 fertilized eggs from each female (or 600 eyed embryos for
SPC treatments) were placed in egg cups for hatching and monitoring. After
swim up, remaining fry were thinned to a target of 100 fry/treatment and
monitored for an additional 15 day post swim up feeding trial. Test termination
ranged from 83 to 88 days after hatch for all but the Spring Creek Hatchery egg
treatments, which occurred 50 days after the arrival of fertilized, eyed embryos
from that hatchery.
Endpoints measured in the laboratory study were fecundity, hatch, growth,
survival/mortality, and feeding success (growth) post swim up. Larval brown
trout were also evaluated for deformities (craniofacial, vertebral, fin) and edema.
For this study, deformities were combined and assessed as having at least one
deformity, or being fully free of deformities (i.e., normal).
C-60 Draft Document
-------
Effects Data: Se concentrations in eggs collected from 26 ripe females at 3 field locations
ranged from 6.2-12.8 mg Se/kg dw at CC150, 6.9-14.0 mg Se/kg dw at CC350,
and 11.2-40.3 mg Se/kg dw at LSV2C. Se concentrations in hatchery eggs ranged
from 0.76-1.2 mg Se/kg dw at the SC hatchery, and were 0.73 mg Se/kg dw at
the SPC hatchery. The Se whole body concentration in field collected fish ranged
from 7.2-22.6 mg/kg dw at LSV03, 4.7-8.4 mg/kg dw at CC150, and 5.5-9.2
mg/kg dw at CC350. Se whole body concentrations in SC hatchery fish ranged
from 2.5-4.3 mg/kg dw.
Because of concerns raised in a U.S. Fish and Wildlife (2012) review of the
Formation Environmental (2011) report, most notably with respect to the
consequences offish lost due to an overflow event during the 15 day post swim
up portion of the test, all endpoints were measured according to a both an
"optimistic" and a "worst-case" scenario. The "worst-case" scenarios were
introduced to examine the comment raised in the U.S. FWS (2012) review that
fish lost to overflow during the post swim up test were assumed to have been
dead or deformed. As an alternative to their proposal that all treatments that lost
fish to overflow should be discarded from the study, we examined the effect of
those individuals being either dead and/or deformed. Therefore, a total of six
EC20s (3 endpoints x 2 scenarios/endpoint) were calculated here.
The U.S. FWS (2012) review also noted fish that survived but failed to reach
swim up, which occurred among the offspring of the five females with the
highest egg selenium concentrations (LSV2C-003, -004, -005, -010, and -021)
would have likely died in the wild. We concur with this assertion, and treated all
fish that failed to reach swim up as dead, with respect to survival.
Three endpoints (percentage fully free from deformities (% normal), percentage
surviving from hatch through 15 day post swim up (% survival), and percentage
surviving from hatch through 15 days post swim up AND fully free of
deformities (% alive and normal)) were analyzed. Selenium concentrations and
respective counts are shown in Table 1 (% normal), Table 2 (% survival), and
Table 3 (% alive and normal). For all tables, each sample ID represents eggs
hatched from a single female fish, with the exception of the 4 SPC samples,
which were obtained as eyed eggs. Plots and ECi0s of each endpoint for both the
optimistic and worst case scenarios are shown in Figure 1. All analyses were
performed in TRAP (version 1.21) using tolerance distribution analysis and
assuming a triangular data distribution.
For both the worst-case and optimistic scenarios, ECi0s were lowest for
deformities, followed by survival, with the ECio for the combined endpoint being
the highest (Figure 1). For a given endpoint, ECi0s for the worst-case scenario
were lower than for the optimistic scenario. The final endpoint selected was %
normal, worst-case scenario (Figure Ib - top right), with an ECi0 of 15.91 mg
Se/kg dw egg.
In all of the above analyses, larvae hatched from eggs from both wild caught fish
and hatchery fish were combined in this analysis. The fish from the hatchery
were much larger and had many more eggs than the wild fish. In addition, the
hatchery fish contained a much lower concentration of selenium in their eggs
than the wild eggs. Furthermore, the fish from the Spring Creek Hatchery arrived
C-61 Draft Document
-------
as fertilized, eyed eggs, and reached swim up in 34 days, compared to 67-75 days
for eggs fertilized in the field or laboratory by Formation Environmental (2011).
Despite the differences between hatchery and wild-caught fish, the inclusion of
larvae hatched from eggs obtained from hatcheries produced similar ECio values.
Figure 2 provides a comparison of the worst-case deformity endpoint, with and
without the hatchery data. The ECi0 for wild fish only is 16.89 mg/kg egg dw
(Figure 2a); compared to an EC 10 of 15.91 mg/kg egg dw for both wild caught
and hatchery fish combined (Figure 2b). The ECio value of 15.91 mg/kg for the
wild+hatchery dataset is selected as the effect concentration because there does
not seem to be an apparent reason to exclude the hatchery fish as a reference for
larval survival.
Effect Concentration: 15.91 mg Se/kg dw in eggs
Table 1. Brown trout selenium concentrations and deformity data from hatch to test end. Worst
case counts assumed that all fish lost to the overflow event during the post swim up portion of the
study would have been deformed.
Sample
IDa
SC-001
SC-002
SC-003
SC-004
SC-005
SC-006
SC-007
SC-008
spc-oor
SPC-002C
SPC-005C
SPC-006C
CC-150-
009
CC-150-
011
CC-150-
012
CC-150-
013
CC-150-
015
CC-150-
016
CC-150-
017
CC-150-
018
Whole
body
Se, mg/kg
dw
3.6
4.1
3.7
4.3
3
3.1
2.7
2.5
8.4
5.6
6.7
5.9
6
7
5.6
4.7
EggSe
mg/kg
dw
0.76
0.94
0.83
0.92
1.2
1.2
1
0.96
0.73
0.73
0.73
0.73
12.8
8.4
8.5
8.4
9.1
7.5
6.6
6.9
#
Normal
63
72
131
46
23
457
93
283
427
371
400
427
106
87
156
137
210
13
99
195
# Assessed for
deformities.
"Optimistic
Case"
115
113
302
140
42
535
137
359
570
545
561
556
142
266
282
310
445
23
163
486
# Lost to # Assessed for
overflow deformities plus
during post # lost. "Worst
swim up test Case"
115
113
9 311
140
42
535
137
10 369
570
545
561
556
142
266
282
26 336
445
43 66
33 196
486
C-62 Draft Document
-------
Sample
IDa
CC-150-
020
CC-350-
006
CC-350-
007
CC-350-
008
LSV2C-
002
LSV2C-
003
LSV2C-
004
LSV2C-
005
LSV2C-
008
LSV2C-
010
LSV2C-
012
LSV2C-
016
LSV2C-
017
LSV2C-
019
LSV2C-
020
LSV2C-
021
Whole
body
Se, mg/kg
dw
7.2
9.2
5.5
8.5
8.9
13.8
17.9
13.6
9.6
22.6
7.2
9.2
13.2
8.6
11.3
20
EggSe
mg/kg
dw
6.2
14
6.9
9.5
12.8
40.3
36
26.8
17.7
38.8
13.2
13.4
20.5
12.5
11.2
28.1
#
Normal
453
120
68
269
483
2
16
8
147
5
217
440
110
267
240
8
# Assessed for
deformities.
"Optimistic
Case"
558
386
131
338
544
100
142
149
194
80
554
530
150
390
296
172
# Lost to # Assessed for
overflow deformities plus
during post # lost. "Worst
swim up test Case"
558
386
20 151
28 366
16 560
100
142
149
45 239
80
554
530
19 169
39 429
36 332
172
a SC - Saratoga National Fish Hatchery; SPC - Spring Creek Trout Hatchery; CC - Crow Creek; LSV -
Sage Creek
b Test end was 15 days after swim up.
0 Arrived as fertilized, eyed-eggs. No whole body Se measurement possible.
C-63 Draft Document
-------
Table 2. Brown trout selenium concentrations and survival data from hatch to test end. Worst case counts assumed that all fish lost to the
overflow event during the post swim up portion of the study would have been deformed.
Sample IDa
SC-001
SC-002
SC-003
SC-004
SC-005
SC-006
SC-007
SC-008
SPC-001C
SPC-002C
SPC-005C
SPC-006C
CC-1 50-009
CC-150-011
CC-150-012
CC-150-013
CC-150-015
CC-150-016
CC-150-017
CC-150-018
CC-1 50-020
CC-350-006
CC-350-007
CC-350-008
LSV2C-002
LSV2C-003
LSV2C-004
LSV2C-005
LSV2C-008
LSV2C-010
Whole
body
Se, mg/kg
dw
3.6
4.1
3.7
4.3
3
3.1
2.7
2.5
8.4
5.6
6.7
5.9
6
7
5.6
4.7
7.2
9.2
5.5
8.5
8.9
13.8
17.9
13.6
9.6
22.6
EggSe
mg/kg
dw
0.76
0.94
0.83
0.92
1.2
1.2
1
0.96
0.73
0.73
0.73
0.73
12.8
8.4
8.5
8.4
9.1
7.5
6.6
6.9
6.2
14
6.9
9.5
12.8
40.3
36
26.8
17.7
38.8
#Eggs
Hatche
d
144
138
340
189
70
564
188
396
598
585
589
593
173
288
314
402
479
89
223
522
584
432
181
407
584
404
309
287
263
108
Prop.
Survival
. Hatch
to swim
up
0.951
0.978
0.982
0.868
0.914
0.988
0.856
0.985
0.987
0.966
0.986
0.971
0.942
0.993
0.965
0.891
0.971
0.966
0.969
0.969
0.990
0.944
0.950
0.951
0.993
0.079
0.414
0.387
0.989
0.231
Prop
survival. Post
swim up.
"Optimistic
Case"
0.990
0.990
0.989
0.971
1.000
0.990
0.970
1.000
1.000
1.000
1.000
1.000
0.990
1.000
0.990
0.973
1.000
1.000
1.000
1.000
1.000
0.980
0.988
0.986
1.000
0.281
0.477
0.622
0.982
0.440
Prop
survival. Post
swim up.
"Worst
Case"
0.990
0.990
0.890
0.971
0.984
0.990
0.970
0.900
1.000
1.000
1.000
1.000
0.990
1.000
0.990
0.720
1.000
0.500
0.670
1.000
1.000
0.980
0.790
0.710
0.840
0.281
0.477
0.622
0.540
0.440
Prop survival.
Hatch to end.
"Optimistic
case"
0.942
0.968
0.971
0.842
0.914
0.978
0.830
0.985
0.987
0.966
0.986
0.971
0.933
0.993
0.955
0.866
0.971
0.966
0.969
0.969
0.990
0.926
0.938
0.938
0.993
0.022
0.197
0.240
0.971
0.102
Prop survival.
Hatch to end.
"Worst case"
0.942
0.968
0.874
0.842
0.900
0.978
0.830
0.886
0.987
0.966
0.986
0.971
0.933
0.993
0.955
0.641
0.971
0.483
0.649
0.969
0.990
0.926
0.751
0.675
0.834
0.022
0.197
0.240
0.534
0.102
Est. #
survived.
Hatch to end.
"Optimistic
case"
136
134
330
159
64
551
156
390
590
565
581
576
161
286
300
348
465
86
216
506
578
400
170
382
580
9
61
69
255
11
Est. #
survived.
Hatch to end.
"Worst case"
136
134
297
159
63
551
156
351
590
565
581
576
161
286
300
258
465
43
145
506
578
400
136
275
487
9
61
69
140
11
C-64 Draft Document
-------
Sample IDa
LSV2C-012
LSV2C-016
LSV2C-017
LSV2C-019
LSV2C-020
LSV2C-021
Whole
body
Se, mg/kg
dw
7.2
9.2
13.2
8.6
11.3
20
EggSe
mg/kg
dw
13.2
13.4
20.5
12.5
11.2
28.1
#Eggs
Hatche
d
591
570
217
471
357
424
Prop.
Survival
. Hatch
to swim
up
0.971
0.965
0.885
0.953
0.986
0.288
Prop
survival. Post
swim up.
"Optimistic
Case"
1.000
1.000
0.963
1.000
1.000
0.730
Prop
survival. Post
swim up.
"Worst
Case"
1.000
1.000
0.780
0.610
0.640
0.730
Prop survival.
Hatch to end.
"Optimistic
case"
0.971
0.965
0.852
0.953
0.986
0.210
Prop survival.
Hatch to end.
"Worst case"
0.971
0.965
0.690
0.582
0.631
0.210
Est. #
survived.
Hatch to end.
"Optimistic
case"
574
550
185
449
352
89
Est. #
survived.
Hatch to end.
"Worst case"
574
550
150
274
225
89
a SC - Saratoga National Fish Hatchery; SPC - Spring Creek Trout Hatchery; CC - Crow Creek; LSV - Sage Creek
b Test end was 15 days after swim up.
0 Arrived as fertilized, eyed-eggs. No whole body Se measurement possible.
C-65 Draft Document
-------
Table 3. Brown trout selenium concentrations and survival + deformity data (combined endpoint) from hatch to test end. Worst case
counts assumed that all fish lost to the overflow event during the post swim up portion of the study would have been deformed.
Sample IDa
SC-001
SC-002
SC-003
SC-004
SC-005
SC-006
SC-007
SC-008
SPC-001C
SPC-002C
SPC-005C
SPC-006C
CC-1 50-009
CC-150-011
CC-150-012
CC-150-013
CC-150-015
CC-150-016
CC-150-017
CC-150-018
CC-1 50-020
CC-350-006
CC-350-007
CC-350-008
LSV2C-002
LSV2C-003
LSV2C-004
LSV2C-005
Whole
body
Se, mg/kg
dw
3.6
4.1
3.7
4.3
3
3.1
2.7
2.5
8.4
5.6
6.7
5.9
6
7
5.6
4.7
7.2
9.2
5.5
8.5
8.9
13.8
17.9
13.6
EggSe
mg/kg
dw
0.76
0.94
0.83
0.92
1.2
1.2
1
0.96
0.73
0.73
0.73
0.73
12.8
8.4
8.5
8.4
9.1
7.5
6.6
6.9
6.2
14
6.9
9.5
12.8
40.3
36
26.8
#
Normal
63
72
131
46
23
457
93
283
427
371
400
427
106
87
156
137
210
13
99
195
453
120
68
269
483
2
16
8
# Normal
that were
dead at # Normal
assessment and alive
63
72
131
46
23
457
93
283
427
371
400
427
106
87
156
137
210
13
99
195
453
120
68
269
483
2 0
16 0
8 0
# Live fish
assessed
for
deformitie
s
115
113
302
140
42
535
137
359
570
545
561
556
142
266
282
310
445
23
163
486
558
386
131
338
544
0
0
0
# Fish lost to
overflow
# Fish died during post
during test swim up test
8
4
7 9
28
6
8
30
6 10
8
20
8
17
11
2
12
46 26
14
3 43
7 33
16
6
26
10 20
21 28
4 16
395
289
267
# Live fish
assessed + # died
during test.
"Optimistic case"
123
117
309
168
48
543
167
365
578
565
569
573
153
268
294
356
459
26
170
502
564
412
141
359
548
395
289
267
# Live fish
assessed + # died
during test + #
lost during post
swim up. "Worst
case"
123
117
318
168
48
543
167
375
578
565
569
573
153
268
294
382
459
69
203
502
564
412
161
387
564
395
289
267
C-66
Draft Document
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Sample IDa
LSV2C-008
LSV2C-010
LSV2C-012
LSV2C-016
LSV2C-017
LSV2C-019
LSV2C-020
LSV2C-021
Whole
body
Se, mg/kg
dw
9.6
22.6
7.2
9.2
13.2
8.6
11.3
20
EggSe
mg/kg
dw
17.7
38.8
13.2
13.4
20.5
12.5
11.2
28.1
#
Normal
147
5
217
440
110
267
240
8
# Normal
that were
dead at
assessment
5
8
# Normal
and alive
147
0
217
440
110
267
240
0
# Live fish
assessed
for
deformitie
s
194
0
554
530
150
390
296
0
# Fish died
during test
4
97
17
20
28
22
5
404
# Fish lost to
overflow
during post
swim up test
45
19
39
36
# Live fish
assessed + # died
during test.
"Optimistic case"
198
97
571
550
178
412
301
404
# Live fish
assessed + # died
during test + #
lost during post
swim up. "Worst
case"
243
97
571
550
197
451
337
404
a SC - Saratoga National Fish Hatchery; SPC - Spring Creek Trout Hatchery; CC - Crow Creek; LSV - Sage Creek
b Test end was 15 days after swim up.
0 Arrived as fertilized, eyed-eggs. No whole body Se measurement possible.
C-67
Draft Document
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Optimistic
Worst Case
100 -,
80
60
40
20
w
• ••*
-0.2 0.2 0.6 1.0 1.4 1,
log (mg Se/Kg egg dw)
100
80
60
40
20
VH-
>V*A
-0.2 0.2 0.6 1.0 1.4
log (mg Se/Kg egg dw)
1.8
w
100
80
60
40
20
-0.2 0.2 0.6 1.0 1.4
log (mg Se/Kg egg dw)
1.8
-0.2 0.2 0.6 1.0 1.4 1.8
log (mg Se/Kg egg dw)
ro
E
T3
C
ro
100
80
60
40
20
-0.2 0.2 0.6 1.0 1.4
log (mg Se/Kg egg dw)
1.8
ro
E
T3
C
ro
100
80
60
40
20
-0.2 0.2 0.6 1.0 1.4
log (mg Se/Kg egg dw)
1.8
Figure 1. Concentration-response relationships of brown trout deformities (a-b), survival (c-d),
and deformities+survival (e-f) in response to selenium concentrations in eggs. Each
endpoint was evaluated under optimistic and worst-case scenarios with respect to larval
fry lost during the 15-day post swim up test. ECi0s (in mg Se/kg egg dw) for each
endpoint-scenario combination were as follows: Deformities (18.36 - optimistic (a);
15.91 - worst case (b)); Survival (20.40 - optimistic (c); 16.79 - worst case (d));
Combined (21.16 - optimistic (e); 20.65 - worst case (f)).
C-68
Draft Document
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Wild Caught Only
100 n
80 -
CD 60 -
E
o
z
£S 40 -
20 -
-0.2 0.2 0.6 1.0 1.4
log (mg Se/kg egg dw)
1.8
Wild Caught Plus Hatchery
100 n
80 -
co 60 -
E
o
z
SS 40 -I
20 -
I •
• .•
• •
•
-0.2
0.2
0.6
1.0
1.4
1.8
log (mg Se/kg egg dw)
Figure 2. Proportion of brown trout larvae fully free from deformities, worst-case scenario, for
larvae hatched from eggs of wild-caught fish only (a), and from eggs of wild
caught+hatchery fish (b). The ECi0 for the worst-case deformity endpoint for the wild-
caught dataset was 16.89 mg Se/ kg egg dw, with a 95% confidence interval of 15.38-
18.55 mg Se/ kg egg dw). The ECio for the worst-case deformity endpoint for the wild
caught plus hatchery dataset was 15.91 mg Se/ kg egg dw, with a 95% confidence
interval of 14.77-17.13 mg Se/ kg egg dw).
C-69
Draft Document
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Besser, J.M., W.G. Brumbaugh, D.M. Papoulias, C.D. Ivey, J.L. Kunz, M. Annis, and C.G.
Ingersoll. 2012. Bioaccumulation and toxicity of selenium during a life-cycle exposure with desert
pupfish (Cyprinodon macularius}: U.S. Geological Survey Scientific Investigations Report 2012-5033,
30 p. with appendixes.
Test Organism:
Exposure Route:
Test Duration:
Study Design:
Desert pupfish (Cyprinodon macularius)
Dietary and waterborne. Pupfish were fed the oligochate, Lumbriculus
variegatus, which had been grown on a diet of selenized yeast.
180 days life cycle, 21 days Fl larvae, 58 days Fl juveniles and adults.
Desert pupfish (Cyprinodon macularius), a federally-listed endangered species,
were exposed simultaneously to waterborne and dietary selenium at six exposure
levels (controls and five selenium treatments) in a three-phase life cycle exposure
study. Aqueous exposures were prepared using sodium selenate and sodium
selenite salts at an 85%-15% proportion, respectively. Pupfish were fed the
oligochate, Lumbriculus variegatus, daily to satiation (25 to 30% rations based
on wet weights). Prior to being fed to the pupfish, the oligochaetes were exposed
to aqueous selenium and fed selenized yeast at appropriate concentrations to
attain the target dietary tissue concentrations. The measured concentrations in
water, oligochaetes (pupfish diet), and pupfish tissues for the control and five
treatments during the life cycle exposures.
Treatment
Control
Se-1
Se-2
Se-3
Se-4
Se-5
water
HS/L
nd
3.4
6.2
14
26
53
oligochaetes
mg/kg dw
1.6
5.1
7.3
14
24
52
pupfish, mg/kg dw
F0 WB eggs F! WB
0.75 1 1.2
2.5 3 3.4
3.4 4.4 3.7
6.7 8 6.7
12 13 12
24 27 31
The 85-day Phase 1 exposure was initiated with approximately five week old
juvenile pupfish (F0). Phase 1 consisted of two separate groups with one group
(started two weeks prior to the second group) used for determining survival,
growth and whole body selenium concentrations, and the other group used for
survival assessment and to provide adults for the main reproduction exposure.
Both groups in Phase 1 were similarly exposed to all six treatments, with each
treatment having 8 replicates and 10 fish in each replicate.
At the end of the 85-day Phase 1 exposure, the pupfish were reproductively
mature and were used for the Phase 2 exposure, the main reproduction study. A
preliminary reproduction study was conducted with adults from the first exposure
group of F0 pupfish. These fish were divided into two spawning groups and eggs
were collected on four dates during a 9-day period. The main purpose of the
preliminary study was to confirm the reproductive maturity of the pupfish, but
samples of larvae from this study were used for assessment of deformities. The
main reproduction study in Phase 2 was started with adults from the second F0
C-70
Draft Document
-------
exposure. These fish were sorted into spawning groups (1 male and 3 females) in
7-L exposure chambers, with eight replicate spawning groups per selenium
treatment. Spawning activity was monitored by removing (and replacing)
spawning substrates from each chamber three times a week (Monday-
Wednesday-Friday). There were 23 egg collection dates during a 60-day period.
All eggs were counted and eggs collected from eight Wednesdays were used for
hatching success, deformities and FI larval and juvenile growth and survival in
the 58-day Phase 3 exposure. Larvae were examined for developmental
endpoints including edema, delayed development, and skeletal, eye, craniofacial,
and fin deformities.
Effects Data: A summary of the endpoints by each treatment level is shown below.
Table 1. Summary of pupfish toxicity endpoints by exposure treatment (average across all
replicates). There were no statistically significant differences across controls and selenium
amendment treatments for any of the endpoints shown here (1-way ANOVA, a=0.05).
Endpoif Control Se-1 Se-2 Se-3 Se-4 Se-5
FO survival, day 28
FO survival, day 56
FO survival, day 85
FO survival, day 150
FO growth, day 28
FO growth, day 56
FO growth, day 85
FO growth, day 150
Fl survival, day 30
Fl survival, day 58
Fl growth, day 30
Fl growth, day 58
total number eggs
% reduction eggs
avg % deformities, main
avg % deformities, preliminary
100
100
100
91
213
535
935
1718
100
100
73
260
6845
NA
5.3
4.4
100
100
100
94
206
526
998
1763
100
100
73
264
6331
8
2.7
8.8
100
100
100
94
204
486
941
1776
100
93
76
286
4143
39
4.9
11.6
100
100
100
94
198
469
934
1755
100
90
78
286
4386
36
2.4
14.3
100
100
100
91
213
509
914
1673
98
95
77
288
3337
51
11.4
10.7
98
100
100
97
203
447
1053
1606
98
88
58
255
5225
24
8.1
21
'Endpoint units: survival, %; growth, mg wet weight; % reduction eggs is relative to the control.
The authors observed no significant differences in pupfish survival or growth
among treatments. The authors hypothesized the lack of statistically significant
acute effects was because the pupfish in this study were near their chronic
toxicity threshold, as suggested by the (non-significant) mean reductions in
growth (7% in F0 day 150) and survival (12% in FI day 58) in the highest
selenium treatment (Se-5), relative to controls (Table 1).
C-71 Draft Document
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Egg hatching and larval survival in all selenium treatments (not listed in Table 2)
were within 10 percent of control means, and differences among treatments were
not related to selenium exposure. The authors noted that the highest selenium
treatment, Se-5, did have the lowest larval survival (84%) and lowest combined
egg hatching and larval survival (76 percent). The means frequencies of
deformities were higher in the two highest Se treatments (Se-4 and Se-5, Table
1); however % deformities across treatment levels were not statistically
significant (1-way ANOVA, p=0.13; Beckon et al. (2012). However, overall
deformity rates were statistically significantly higher in a preliminary
reproduction than in the main reproduction test. Beckon et al. (2012)
hypothesized that the reason for the difference in deformity rates between the two
tests was related to the time the eggs were collected relative to the time the
respective spawning groups were isolated. Eggs were collected in the preliminary
reproductive study 1-9 days after the spawning groups were isolated, whereas
spawns used to characterize deformities in the main reproduction test were
collected at least 14 days after the onset of spawning. The larvae produced from
the earlier collected eggs may have been exposed to higher selenium
concentrations in the egg. The pattern of a gradual decrease in egg selenium
concentration over time was observed in the life cycle study.
Egg production varied considerably over the 23 collection dates (Table 2 and
Figure 1). Although each of the selenium treatments had a lower total number of
eggs relative to the control, one-way ANOVAs of cumulative egg production did
oo * j oo r^
not indicate significant differences among treatments on either a per-replicate
basis (p=0.34) or on a per-female basis (p=0.20). Similarly, repeated measures
ANOVA indicated no differences between treatments, but the authors indicated
significant differences among sampling dates and significant interactions of
treatment and date. Because of the lower number of eggs in the selenium
treatments and the significance of the interaction of treatment and time, the
authors concluded that pupfish egg production was adversely affected by
elevated selenium exposure and reported significant reductions in egg production
at treatment levels Se-2 through Se-5 (4.4 to 27 mg/kg dw Se in eggs). The
authors recognized that typically larval survival and deformities are the most
sensitive reproductive endpoint for selenium toxicity and not egg production and
suggested more study is needed to confirm the unusual sensitivity of pupfish egg
production to selenium.
C-72 Draft Document
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Table 2. Number of pupfish collected on each sampling date throughout the study, by treatment
level. Values represent the sum of all eggs collected on a given date for a given Se treatment.
Dav
2
4
7
9
11
14
17
21
23
25
28
30
32
35
37
39
42
44
51
53
56
58
60
Control
136
275
307
265
401
417
448
303
287
340
366
130
323
320
236
326
507
251
380
278
199
202
148
Se-1
112
173
273
252
136
359
456
664
205
308
273
164
304
427
176
151
140
133
359
63
478
329
396
Se-2
90
123
301
226
424
333
206
404
141
94
103
104
271
81
41
159
55
66
227
38
138
331
187
Se-3
67
142
283
169
319
246
163
204
143
143
101
52
78
150
113
184
193
152
338
197
195
410
344
Se-4
122
188
160
271
265
198
145
163
177
150
95
82
75
74
38
113
101
69
305
56
238
143
109
Se-5
94
162
432
283
380
401
232
400
175
228
181
132
151
223
38
140
140
137
370
188
222
320
196
C-73 Draft Document
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-S
c
o
OJ
Q.
s/i
ao
no
01
S3
£
3
z
700
600
500
400
300
200
100
•control
So-1
• Se-2
• Sc-3
Se-4
Se-5
10 20 30 40
Collection day
50
60
70
Figure 1. Pupfish egg production by sampling date
Several findings from the pupfish study put a clear demonstration of effect due to
selenium in question. The fact that the typical sensitive endpoints for selenium,
larval survival and deformities, were not demonstratively responsive to selenium
through the highest treatment level, the fact that the egg production data did not
show significance among treatments alone, and the fact that egg production
increased at the highest selenium treatment level provide sufficient doubt of a
clear effect due to selenium. These issues are discussed below.
Examination of the Repeated Measures Analysis:
Analysis Using the Full Dataset: The effects of selenium treatment and sampling date on pupfish egg
production (eggs per female per day) were reanalyzed. First, the data were
reanalyzed using repeated measures ANOVA. Results of the repeated measures
ANOVA analysis were qualitatively similar to those reported in Besser et al.
(2012) and are shown in the following table.
C-74
Draft Document
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Between Subjects
Source
Se treatment
Error
SumofSq. df
2,202.6 5
10,543.5 42
Mean Sq. F-rat. p-value
440,5 1.755 0.143
251.0
Within Subjects
Source Sum of Sq. df
Sampling Date 1,867.5 22
Se Treatment x Sampling Date 2,566.3 110
Error 15,771.8 924
Mean Sq. F-rat. p-value
84.89 4.973 <0.001
23.33 1.367 0.010
17.07
As with the results reported in Table 7 of Besser et al. (2012), there was no main
effect of Se treatment (note - for purposes of these analyses and associated text,
"Se treatment" is defined as the control plus the 5 treatments that received Se
amendments), but there was a statistically significant (p<0.05) effect of sampling
date and a significant date by Se treatment interaction. Results were qualitatively
similar because the p-values for Se treatment and sampling day were identical in
both analyses, yet the p-values for the day by Se treatment interaction term were
nearly identical.
A statistically significant sampling date effect means that there were significant
differences in overall egg production on different sampling dates. Daily egg
production per female ranged from 2.176 on day 2 to a high of 7.294 on day 11,
and was variable throughout the study. Of greater interest is the statistically
significant day x Se treatment interaction. What this means is, although there was
not an overall significant effect of Se treatment on egg production per female,
there was a significant Se treatment effect (p<0.05) on egg production per female
on at least one of the 23 sampling dates.
Analysis after Removal of Control Replicate Outlier: Repeated measures ANOVA analysis confirmed
the results reported in Besser et al. (2012). However, as shown on Figure 8b of
Besser et al. (2012), one replicate chamber (replicate g) within the control
treatment had only one surviving female pupfish from day 7 through the end of
the test (day 60), and that replicate also had the highest overall egg production
per female of any test chamber. All replicate chambers in all treatments began
with three female pupfish, and the replicate described above was the only one
with only one surviving female. All three females survived the 60 day test in the
majority of the replicate chambers. In order to determine whether the significant
date by Se treatment interaction was an artifact of this one test chamber, data
were reanalyzed after removing this replicate.
One requirement of repeated measures ANOVA is that the model cannot contain
any missing values. An alternative to repeated measures ANOVA when data are
missing, and the most commonly followed procedure under these circumstances,
is to analyze the data using a mixed model. This was the procedure followed
here.
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The results of a fully balanced mixed model (no missing data) should be identical
to repeated measures ANOVA. As an initial check, the full dataset was
reanalyzed as a mixed model. Sample chamber was the random effect parameter,
and Se treatment, sampling date, and Se treatment by sampling date were the
fixed effect parameters. As expected, the F-ratios for the effects of selenium
treatment, sampling date, and the sampling date by Se treatment interaction were
identical. Next, the data were reanalyzed after removing data from control
replicate g from all sampling dates. Results of this analysis are reported in the
table below.
Mixed Model - Fixed
Effect Numerator df Denominator df F-ratio p-Value
Se Treatment 5 902 1.087 0.366
Sampling Date 22 902 6.042 <0.001
Se Treatment x Sampling Date 110 902 1.310 0.023
T
The statistically significant interaction between Se Treatment and Sampling Date
persisted after removal of the potentially anomalous control treatment chamber
with one female pupfish. In other words, even after removing the one potentially
anomalous control replicate, there were still some individual sampling dates
where the effects of Se treatment were statistically significant (p<0.05).
Se Treatment x Sampling Date Interaction: When a significant interaction is
observed in a repeated measures ANOVA, the next recommended step in the
process is to examine each of the repeated measures (sampling dates) separately
to identify those dates where the significant difference in Se treatment level
occurred. When individual dates for the full dataset (including the replicate with
one surviving female) were analyzed separately, there were significant (p<0.05)
effects of Se treatment level on egg production on days 28, 35, 37, 42, and 53 (1-
way ANOVA, df5 42). There were no significant Se treatment effects on the
remaining 18 sampling dates. ANOVA results are summarized in the table
below.
Sampling Date F-ratio p-value
28 2.501 0.045
35 2.704 0.033
37 3.351 0.012
42 4.294 0.003
53 3.352 0.012
Because of the large number of comparisons (23 individual ANOVA models for
each sampling date), an alpha of 0.05 is inappropriate for this particular analysis.
This is because an alpha of p<0.05 means that a statistically significant result will
be observed 5% of the time due to chance alone (Type I error). In order to control
for the increased likelihood of a Type I error when making multiple comparisons,
the alpha level of 0.05 was adjusted using Sidak's correction (Abdi 2007). For 23
comparisons and an alpha of 0.05 for one comparison, the adjusted alpha using
Sidak's correction is as follows:
C-76 Draft Document
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1 - (1 - 0,05)2a = 0,0027
After adjusting alpha to account for the 23 separate sampling dates, there were no
sampling dates with a significant Se treatment effect (p<0.0027). As a result, it
was not necessary to perform post hoc means comparisons tests for any of the
individual sampling dates to determine which Se treatment levels were
significantly different from each other.
Each of the 23 sampling dates for the dataset where the replicate chamber from
the control treatment with one surviving female pupfish was excluded were also
analyzed using one-way ANOVA to determine which sampling dates had
significant Se treatment effects. Significant differences among Se treatment
levels at alpha 0.05 are shown in the table below.
Sampling Date F-ratio p-value
35 2.839 0.027
42 3.164 0.017
53 2.549 0.042
After adjusting alpha to account for the 23 separate sampling dates, there were no
sampling dates with a significant Se treatment effect (p<0.0027). As with the full
dataset, it was not necessary to perform post hoc means comparisons tests for any
of the individual sampling dates to determine which Se treatment levels were
significantly different from each other.
Summary of Repeated Measures Analysis: This analysis demonstrated that although there was a
significant Se treatment by sampling date interaction, regardless of whether or
not the control treatment chamber with one female pupfish was excluded,
differences among Se treatment levels were only observed for a small subset of
the 23 sampling dates. Furthermore, after adjusting alpha to account for multiple
comparisons, one-way ANOVA analyses conducted separately for each sampling
date to locate the source of the Se Treatment x Sampling Date interaction
determined that there were no statistically significant differences among Se
treatment levels on any sampling date, precluding the need to perform post hoc
comparison of means tests to identify significant differences among individual Se
treatments.
Combining Effect Metrics Using a Population Model: To improve the certainty of any conclusions to
be made about the sensitivity of pupfish to selenium, it is also worthwhile to
consider the biological (as opposed to statistical) significance of the observations.
But for total egg production, survival, and deformities, the concentration-
response curves did not show a sufficient concentration-related effect to calculate
an EC10. Nevertheless, because Besser et al. (2012) raised the issue of an
interaction of egg production with time, there is a particular concern that there
could be a delay in egg production that would reduce population growth rate,
even while total numbers of eggs were not significantly affected. This question
was evaluated by constructing a population model corresponding to data
available from the test.
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This modeling approach allows for combining and properly weighting effects on
egg production, timing of egg production, and survival. Percent hatch and percent
deformities were also considered in alternate calculations. Because the model is
only intended for combining the lab data into a unified concentration-response
curve, it cannot be interpreted as making real-world population predictions. The
relevant data were taken from spreadsheets Besser et al. (2012b and 2012c),
which were provided by Besser.
The reproduction and larval endpoints spreadsheet, Besser et al. (2012b),
presents egg production at 23 time points. This information thus allows for 23
adult life stages, each assigned its own fecundity. Another page of this
spreadsheet provides larval survival data, thus defining survival of the early life
stage. The juvenile and adult survival spreadsheet, Besser et al. (2012b), defines
a survival rate shared by these life stages.
For each treatment, the data from the test thus provide all the needed input for 25
life stages: (1) an embryo-larval stage with its own daily survival probability
(along with hatching and deformity percentages, when considered in alternative
calculations), (2) a non-reproducing juvenile stage sharing its treatment's daily
survival probability with the adult stages, and (3 - 25) 23 short-duration adult
stages each with its own egg production, but sharing its treatment's daily survival
probability with the treatment's other adult stages. Use of the data is detailed
below.
Egg Production: Egg production at the test's 23 observation time points is from
the spreadsheet Besser et al. (2012b), expressed as eggs per female per day. The
intent of Besser et al. (2012) was for each treatment to have eight replicates, and
each replicate was to have one male and three females. Only replicates matching
that design were used. Early in the test Control Replicate "g" ended up with only
one female, and was therefore not used here. Se-1 Replicate "h" and Se-3
Replicates "d" and "h" had been inadvertently stocked with two males and two
females, and were likewise not used here. Table 3 shows the time course of egg
production incorporated into the population model. For each treatment, model
fecundity, ml, for life stages i = 3 - 25, is the observed egg production divided by
2, in order to provide female eggs per female per day.
Percent Hatch: The spreadsheet Besser et al. (2012b) presents percent hatch for
eggs collected at selected time points. Within each treatment these were
averaged. In selenium reproductive studies percent hatch is often treated as a
noise variable unrelated to selenium exposure. Consequently, the population
growth calculations were run with and without including percent hatch. When
hatch was incorporated into the calculation, daily fecundity was reduced by
multiplying by percent hatch.
Deformities: The Besser et al. (2012b) spreadsheet also provides deformity
counts for the study's preliminary test and for its main test. Only the main test
results were used here. Counts were totaled for each treatment, and a percentage
calculated. Population growth calculations were performed both with and without
consideration of deformity percentage. For simplicity when considered, a worst
case assumption was made that deformed individuals do not contribute to the
C-78 Draft Document
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population. Percent deformity was thereby handled in manner parallel to percent
hatch, by multiplying daily fecundity by percent free of deformity.
Table 3. Life stage durations, and observed eggs per female per day at observation time points
for control and selenium treatments, only with replicates having the design three females and
one male. Model fecundity, m, is set at one-half the observed, to yield female eggs per female.
Repro Study
Obser-
vation Day
-
-
2
4
7
9
11
14
17
21
23
25
28
30
32
35
37
39
42
44
51
53
56
58
60
Assigned
Life Stage
Number
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
Total as £ (duration • eg
Life
Stage
Duration
35
85
2
2
3
2
2
3
3
4
2
2
3
2
2
3
2
2
3
2
7
2
3
2
2
;gs/f/d) =
Observed Eggs/Female/Day
Control
-
-
2.690
5.548
4.333
5.762
8.024
6.540
6.429
3.345
5.786
6.905
4.794
1.881
5.464
4.373
5.631
6.119
7.349
4.798
1.847
6.310
3.183
3.405
3.810
281.6
Se-1
-
-
2.571
4.048
4.302
5.524
3.238
4.905
7.143
7.881
4.643
7.286
4.317
3.881
7.286
7.310
4.417
3.917
2.222
3.274
2.139
1.512
7.317
7.810
8.226
294.3
Se-2
-
-
1.875
2.563
4.181
4.708
8.833
4.625
2.861
4.208
2.938
1.958
1.431
2.167
5.646
1.132
0.927
4.240
1.056
1.719
1.571
0.823
2.076
8.469
4.115
181.9
Se-3
-
-
1.319
2.153
3.185
3.639
4.528
2.296
1.481
1.764
3.806
2.792
1.306
1.403
1.444
2.880
1.556
3.556
2.500
3.194
2.532
5.403
2.491
9.597
6.347
174.7
Se-4
-
-
2.542
3.917
2.222
5.646
5.521
2.750
2.014
1.698
3.688
3.125
1.319
1.708
1.563
1.028
0.792
2.354
1.403
1.438
2.060
1.333
3.528
3.104
2.271
142.0
Se-5
-
-
1.958
3.375
6.000
5.896
7.917
5.569
3.222
4.167
3.646
4.750
2.514
2.750
3.146
3.097
0.792
2.917
1.944
2.854
2.202
3.917
3.083
7.656
4.271
220.1
Larval Survival: The Besser et al. (2012b) spreadsheet also has data for larval
survival after 14 and 21 days for eggs collected at three time points. The fraction
surviving 21 days was used here. For each treatment, the probability of the early
life stage (i=l) surviving each day equals the fraction surviving for 21 days,
raised to the 1/21 power: a\ = aL = (21-d Surv)1721, shown in Table 4.
Juvenile and Adult Survival: A second spreadsheet, Besser et al. (2012c), has
data on juvenile and adult survival after 30 and 58 days. The fraction surviving
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58 days was used (Table 4). Parallel to the handling of larval survival, for each
treatment the juvenile-adult daily survival probability, a]A = (58-d Surv)1758, as
shown in the table. This value applies to life stages i=2-25 (a2 through 1, there would be a slight youthful bias
within the life stage, such that slightly more than half would be only 1 day into
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the life stage and not ready to graduate, and slightly less than half would be in
their second day and ready to graduate. The above function adjusts for that.1
The projected population growth rate for each treatment was calculated as
follows. The 25x25 projection matrix was placed on an Excel spreadsheet. Each
cell in the diagonal was then modified to subtract the eigenvalue, A, which
represents the population growth rate. That is, each cell in the diagonal was
rewritten as ff^l-y,) - X. The determinant of the 25x25 matrix was then calculated
by function MDETERM. To obtain the population growth rate, Excel's Solver
was then tasked with finding a value for X that yielded a value of zero for the
matrix determinant. In this case,-10~18 < MDETERM < +10"18 was deemed
sufficiently close to zero. Introducing the constraint to look for X values between
1.01 and 1.04 was found helpful for Solver to find the dominant eigenvalue.
When Solver occasionally could not get the determinant within 10"18 of zero,
probably due to a solution oscillation that can occur because the input values y,
are expressed as a function of the solution output A, digits were removed from
Solver's best estimate for A, to provide a new starting value with which Solver
could complete the solution.
Effects on Projected Population Growth Rates: Table 5 and Figure 2 show the model results. Figures
2-B, -C, and -D are almost indistinguishable from Figure 2-A, because hatch and
deformity rates varied so little across treatments. Although population growth
rates at 4.4 - 27 mg Se/kg are less than at 1 - 3 mg Se/kg, the 6-fold increase in
concentration from 4.4 - 27 mg Se/kg yields no change in response.
Consequently, the results do not suggest a selenium-related effect, and no ECio
can be calculated. Based on the combined influences of egg production and
timing, and survival (with or without percentage hatch and deformities), pupfish
does not appear to be among the most sensitive species.
1 The formula for y is undefined (0/0) under the condition a= 1 and 1= 1, so it is not obvious from inspection how it behaves. This
function addresses a model artifact that is called numerical dispersion when it occurs in pollutant transport models. It prevents
overoptimistic rates of moving through the life stages, particularly in the 35-day and 85-day larval and juvenile stages, and allows
a 25-stage model of life duration 180 days to yield precisely the same growth rate as a 180-stage (one day per stage) model,
which was also constructed and checked for comparison. However, in this application where absolute growth rates have no
particular meaning and only relative differences between treatments are of interest, the function does not change the overall
perspective.
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Table 5. Model output: daily population growth rates as k (factor increase) and r (=ln X), for
models that account for survival, fecundity and its timing, and optionally also hatch and/or
deformities. Because k is responding to all the treatment parameters included in the model, its
treatment-to-treatment variations do not exactly track the variations in any single input.
Treat-
ment
Control
Se-1
Se-2
Se-3
Se-4
Se-5
Cone
1
3
4.4
8
13
27
Factors included in model:
All account for survival (<7L , <7JA ) and fecundity (m) and its timing
•k
1.0337
1.0346
1.0299
1.0285
1.0291
1.0294
r
0.0332
0.0340
0.0294
0.0281
0.0287
0.0290
Hatch
•k
1.0330
1.0338
1.0284
1.0277
1.0283
1.0288
R
0.0324
0.0333
0.0280
0.0273
0.0279
0.0283
deformity
•k
1.0334
1.0344
1.0295
1.0283
1.0283
1.0288
r
0.0328
0.0338
0.0291
0.0279
0.0279
0.0284
hatch &
•k
1.0326
1.0336
1.0281
1.0275
1.0276
1.0281
deform.
r
0.0321
0.0331
0.0277
0.0271
0.0272
0.0277
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1.2,
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o
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ai
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1248
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Egg concentration {mg
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&
n !-!
o
o
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Figure 2. Abbott-adjusted pupfish response as modeled population growth rate (solid-filled
symbols) and observed eggs per female per day, larval survival, and juvenile and adult
survival (open symbols). Where used in the population model (to modify fecundity),
hatch and deformity are shown as open symbols. Some open-symbol points are
obscured beneath solid-symbol points. (A) Upper left, egg production and survival only,
(B) upper right, adds in influence of percent hatch, (C) lower left, adds in influence of
deformities, and (D) lower right, adds in influence of percent hatch and deformities.
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Isolating the Influence of Timing of Egg Production: By combining survival with egg production and
its timing in the above analysis, the assessment obscures the influence of timing:
the issue that was the main reason for undertaking population modeling in the
first place. The concern is whether selenium exposure could delay reproduction,
thereby yielding reduced population growth. To help isolate the influence on the
timing of egg production, two population model runs were performed where all
treatments were assigned one of two daily survival rates (0.99 or 0.999) spanning
the full range of daily survival rates observed in the 21 and 58 day survival
calculations. That is, with survival held constant, the only factors varying across
treatments were egg production and timing.
The results are shown in the table below. The Abbott-adjusted results are plotted
in Figure 3. Although the relative differences in Figure 3 population growth rates
are subdued compared to the wider variation in egg production, this is merely a
consequence of the predicted population growth rate being more responsive to
survival than to reproduction. It is still apparent that the variations in total egg
production are affecting growth rate. The question to be addressed here is
whether increasing selenium concentration yields a decline in growth rate beyond
the pattern reflecting total egg production.
Population growth rates, as influenced only by differences in egg
production and timing
Treat-
ment
Control
Se-1
Se-2
Se-3
Se-4
Se-5
Cone
1
3
4.4
8
13
27
With only egg production (m) and its timing
variable across treatments
(7=0.999
•k
.0339
.0338
.0310
.0293
.0293
.0324
r
0.0334
0.0333
0.0306
0.0289
0.0288
0.0318
(7=0.99
•k
1.0246
1.0245
1.0217
1.0201
1.0200
1.0231
r
0.0243
0.0242
0.0215
0.0199
0.0198
0.0228
Inspection of Figure 3 indicates that when survival is assigned a constant value
across treatments, the pattern of population growth differences across treatments
does not suggest an additional selenium-accentuated factor depressing population
growth rate. Population growth at 13 and 27 mg Se/kg is slightly higher than
might be expected from total egg production, when compared to lower
concentrations. The lack of influence of selenium exposure on timing of egg
production is also illustrated by comparing each treatment's cumulative
proportion of egg production over the course of the test, as shown in Figure 4.
Although the treatments differ somewhat in the temporal pattern of their egg
production, there is no consistent relationship with selenium exposure.
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1 3
Ol
Ul
C '1 »
o *- *
a.
M
01
o:
~~ n s
-a u-a
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•4-"
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e
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t _ *
A A
» population growth, 0=0.999
1 population growth, o=0.99
'eees/fomale-day
L 2 4 8 16 32
Egg concentration (mg Se/kg dw)
Figure 3. Predicted population growth rate calculated considering
differences only in egg production and timing (having assigned
uniform survival rates across treatments).
o
1
o
li
an
an
o
o
'•£
o
0.
o
1
0.9
0.8
0.7
0.6
0.5
0.4
0.3
0.2
0.1
0
10 20 30 40 50
Observation Day
60
Figure 4. Cumulative pattern of egg production over time. (Control:
continuous line. Se-1: dot, dot, long dash. Se-2: long dashes. Se-
3: medium dashes. Se-4: short dashes. Se-5: dots.)
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Chronic Value:
In other selenium studies, egg production and percent hatch have not generally
been thought to be related to selenium exposure. Although Besser et al. (2012)
noted that repeated measures ANOVA indicated a potential interaction between
selenium treatment and egg production on particular sampling dates, a thorough
examination of the study data from multiple perspectives indicates no statistically
significant or biologically apparent effect of selenium on egg production, timing
of egg production, or percent hatch at or below the highest tested concentration
of 27 mg Se/kg (dw). Likewise there was no discernible effect on deformity
rates.
In the separate tests of Fl larval survival at 21 days and of Fl juvenile-adult
survival at 58 days, the highest treatment, 27 mg Se/kg (dw), displayed lower
survival than any other treatment. Although the reduction was not sufficient to be
statistically significant, Besser et al. (2012) suggest that this is indicative of a
threshold. Note that among toxicity tests in general, the 10% effect level of the
ECio might or might not be statistically significant from the perspective of
hypothesis testing.
Shown below are the survival rates for the 27 mg Se/kg treatment adjusted to the
control (Abbott-adjusted), or similarly adjusted to the average survival at all
lower treatments (some of which had better survival than the controls). Either
way the adjustment is done, results are similar. (These survival data, Abbott-
adjusted, are included in Figure 2.)
27 mg Se/kg treatment:
adjusted to control
adjusted to all lower treatments
Larval
Surv at 21
days
92.9%
89.1%
Juv-Adlt
Surv at 58
days
87.5%
91.6%
The effect level at 27 mg Se/kg was thus 7% - 13% in the above comparisons.
While the concentration response curve is not sufficiently defined to allow
confident assignment of an ECio, the data suggest a chronic value in the general
neighborhood of 27 mg Se/kg.
An effect level of 27 mg Se/kg egg for the pupfish in this study is consistent with
the findings of Saiki et al. (2012a) who evaluated selenium in two related species
in the Salton Sea, California. These authors measured 3.09 to 30.4 mg/kg whole
body Se levels in mosquitofish and sailfin molliesand based on a lack of a
negative relationship with the catch-per-unit-effort deduced these species were
not adversely affected by selenium. They extrapolated the finding of selenium
tolerance to the pupfish based on the results of another study (Saiki et al 2012b)
in which mosquitofish and sailfin mollies accumulated similar levels of selenium
to the pupfish. Note: the ratio of selenium in whole body to egg tissues in the
pupfish was approximately 1:1 in the Besser study (see first table in the pupfish
study summary above).
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Staub, B.P. W.A. Hopkins, J. Novak, J.D. Congdon. 2004. Respiratory and reproductive characteristics
of eastern mosquitofish (Gambusia holbrooki) inhabiting a coal ash settling basin. Arch. Environ.
Contamin. Toxicol. 46:96-101.
Test Organism:
Exposure Route:
Study Design:
Effects Data:
Chronic Value:
Eastern mosquitofish (Gambusia holbrooki)
Waterborne and Dietary - field exposed
Fish were collected from a contaminated ash basin (ASH) and a reference pond
(REF)
In July 1999, male eastern mosquitofish were collected from ASH and REF
(n=26, n=20, respectively) for measurement of standard metabolic rate (SMR). In
July 1999, gravid female eastern mosquitofish were collected from ASH and
REF and transported to a laboratory for testing. To ensure all females were
fertilized in the field, all offspring used in testing were limited to three weeks
after collection. (Eastern mosquitofish are live-bearers with a four week gestation
period.) Response variables compared between ASH and REF were (1) SMR of
males, (2) brood size of females, (3) percent of live offspring at parturition, and
(4) trace element concentration in females and offspring.
SMRs of males, brood size of females, and offspring viability were not
significantly different between sites. Average (n=5) concentrations of selenium in
females were 11.85 and 0.61 mg/kg dw in ASH and REF sites respectively. The
average concentrations of selenium in offspring were 15.87 mg/kg dw and below
detection in ASH and REF sites, respectively. The authors point out that the
selenium concentrations are an under-estimate of the field levels since the
females were allowed to depurate during their time in the laboratory prior to
parturition.
>11.85 mg Se/kg dw whole body
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Saiki, M.K., B.A. Martin, and T.M. May. 2004. Reproductive status of western mosquitofish inhabiting
selenium-contaminated waters in the grassland water district, Merced County, California. Arch. Environ.
Contamin. Toxicol. 47:363-369.
Test Organism:
Exposure Route:
Study Design:
Effects Data:
Chronic Value:
Western mosquitofish (Gambusia affinis)
Waterborne and Dietary - field exposed
Fish were collected from selenium-contaminated sites and reference sites in the
San Joaquin River watershed.
Western mosquitofish were collected in June and July 2001 from San Luis Drain
(SLD) at Gun Club Road (Se-contaminated site), North Mud Slough at Gun Club
Road (MSN1; reference site); North Mud Slough at State Highway 140 (MSNs;
Se-contaminated site); San Joaquin River at Lander Avenue (SJR; reference site).
20 gravid females from each site were held in the laboratory for two weeks to
quantify live and dead births and to make other measurements. Only 17 females
from SLD were collected. Live and dead fry were visually examined under low
magnification with a binocular microscope for evidence of external abnormalities
(teratogenic symptoms such as spinal curvature, missing or deformed fins, eyes
and mouths and edema).
The percentage of live births was high at both Se-contaminated sites (96.6 to
99.9%) and reference sites (98.8 to 99.2%). There were no obvious anomalies
(e.g., deformities, edema) observed during the study. The concentration of
selenium in 4 postpartum females from the site with the highest selenium
concentration, SLD, ranged from 13.0 to 17.5 mg Se/kg dw (geometric mean of
the high and low is 15.1 mg Se/kg dw. The concentration of selenium of western
mosquitofish collected at each site is in Table D-8.
>15.1 mg Se/kg dw whole body
Table D-8. Selenium in whole body samples of western mosquitofish from study sites
Site
SLD
MSN2
MSN1
SJR
N
8
24
20
22
[Se], mg/kg dw
18.1
9.31
2.72
0.907
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Coughlan, D.J. and J.S. Velte. 1989. Dietary toxicity of selenium-contaminated red shiners to striped
bass. Trans. Am. Fish Soc. 118:400-408.
Test Organism:
Exposure Route:
Test Treatments:
Test Duration:
Study Design:
Effects Data:
Chronic Value:
Striped bass (Morone saxitilis; adults from Lake Norman, NC, approximately
250 g each)
dietary only
Treated fish were fed selenium contaminated red shiners (1 g) from Belews Lake,
NC (9.6 mg Se/kg ww or 38.6 mg Se/kg dw based on a mean reported moisture
content of 75.1 percent). Control fish were fed golden shiners from a local bait
dealer (0.3 mg Se/kg ww or 1.3 mg Se/kg dw based on a mean reported moisture
content of 76.3 percent).
Test treatments were as described above. Two tanks contained treated fish (n =
20 fish total), and one tank offish served as the control (n = 10 fish). Each tank
received a continuous flow of soft well water (hardness and alkalinity approx. 30
mg/L as CaCO3) throughout the exposure.
80 days
During the experiment, all striped bass (n = 10 per tank) were fed to satiation
three times per day. Pre-weighed rations of live red shiners (treated fish) and
golden shiners (controls) were added to the tanks and allowed 5 hours to feed.
Uneaten prey was removed and weighed. Composite whole-body samples of
each prey fish were collected at regular intervals throughout the study for whole-
body tissue selenium analysis. The final selenium concentration in epaxial white
muscle was determined for surviving striped bass at the end of the test. Moribund
striped bass were sacrificed so as to obtain muscle tissue samples for selenium
analysis. Samples of liver and trunk kidney of these and the surviving striped
bass were dissected for observations of histopathology.
Striped bass fed selenium-laden red shiners exhibited changes in behavior
(lethargy, reduced appetite), negligible weight gain, elevated selenium
concentrations in muscle, histological damage, and death. Control fish ate and
grew well, and behaved normally. Average selenium ingestion was between 60
and 140 Og Se/fish per day until day 30. Appetite of the treated fish appeared to
be significantly reduced beyond this point compared to the appetite of the control
group. By day 78, all striped bass fed the Se-laden red shiners either had died or
were moribund and sacrificed for analysis. The final selenium concentration in
muscle of treated striped bass averaged from 3.5 (tank 1) and 4.0 (tank 2) mg/kg
ww, or 16.2 and 18.5 mg/kg dw, respectively, assuming 78.4 percent moisture
content in muscle tissue; default May et al (2000) value for all species. The final
selenium concentration in muscle of control striped bass fed uncontaminated
golden shiners averaged 1.1 mg/kg ww, or 5.09 mg/kg dw (assuming 78.4
percent moisture content in muscle tissue; default May et al (2000) value for all
species).
The chronic value for percent survival of striped bass relative to final selenium in
muscle tissue after being fed Se-laden red shiners is <16.2 mg/kg dw.
An EC20 value could not be calculated for this data set because the data did not
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meet the assumptions required for analysis.
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Bryson, W.T., W.R.Garrett, M.A. Mallin, K.A. MacPherson, W.E. Partin, and S.E. Woock. 1984.
Roxboro Steam Electric Plant 1982 Environmental Monitoring Studies, Volume II, Hyco Reservoir
Bioassay Studies. Environmental Technology Section. Carolina Power & Light Company.
28-day Embryo/Larval Study
Test Organism:
Exposure Route:
Study Design:
Effects Data:
Chronic Value:
Bluegill sunfish (Lepomis macrochirus; embryos and larvae)
dietary and waterborne - field exposure
Native adult bluegill were collected from Hyco Reservoir in Person County,
North Carolina and from a nearby control lake (Roxboro City Lake). Hyco
Reservoir is a cooling lake for Carolina Power & Light and receives the
discharge from the ash storage pond. No selenium values were given for Hyco
Reservoir, total selenium was not detected in the control lake (<1 :g/L). A mean
selenium for the ash pond effluent from a previous study was 53 :g/L (N=59;
range 35-80 :g/L).
All combinations of crosses between the Hyco and control fish were made using
gametes from the collected fish. Fertilized eggs were exposed in egg cups to 0,
20 and 50 percent ash pond effluent under flow-through conditions. Percent hatch
and swim-up successes were measured. Swim-up larvae were released to
exposure tanks where there were fed zooplankton collected from Hyco and the
control lake. Larvae were observed for 28 days at which time survival and weight
were measured.
Survival to the swim-up stage was different between larvae from Hyco females
fertilized with either male type and those larvae from control females fertilized
with either male type. All crosses involving a Hyco female resulted in larvae
exhibiting 100 percent mortality prior to reaching swim-up. Percent survival
from hatch to 28 days for larvae from control females exposed to control water
and fed control lake zooplankton was only 5 and 12 percent for the two replicates
so no meaningful comparisons can be made to the different dilution exposures or
diet exposure. The mean concentrations of selenium in the ovaries, female liver
and female muscle were 49, 130, and 84 mg/kg dw, respectively.
Effect level: <49, <130 and <84 mg Se/kg dw in adult ovaries, liver and muscle,
respectively
<49.65 mg Se/kg dw in whole body using the muscle to whole body equation
<84 mg Se/kg dw maternal muscle
<49 mg Se/kg dw ovary
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Ingestion Study
Test Organism:
Exposure Route:
Study Design:
Effects Data:
Chronic Value:
Bluegill sunfish (Lepomis macrochirus; 30-day old larvae)
Dietary and waterborne - field exposed adults
Juvenile bluegill from crosses with females in 0, 20 and 50 percent ash pond
effluent were transferred to control water and fed zooplankton from either Hyco
or the control lake. Selenium in Hyco and control zooplankton was 45 and 1.9
mg/kg dw, respectively. Duration was not given.
Survival and observations on pathology and morphology were made in the two
diet treatments.
Mortality in larvae fed control zooplankton was 23.7 percent, whereas mortality
in larvae fed Hyco zooplankton was 97.3 percent. There were no differences in
survival (for two diet treatments) in larvae that were raised for the 30 days prior
to the test in different effluent concentrations (0, 20 50 percent). The average
selenium concentrations in the larvae fed control and Hyco zooplankton were 1.9
and 24.7 mg/kg dw, respectively.
Effect level for larval survival: <24.7 mg Se/kg dw in larvae
None recommended for larval tissue.
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Bryson, W.T., W.R.Garrett, M.A. Mallin, K.A. MacPherson, W.E. Partin, and S.E. Woock. 1985a.
Roxboro Steam Electric Plant Hyco Reservoir 1983 Bioassay Report. Environmental Services Section.
Carolina Power & Light Company. September 1985.
28-day Embryo/Larval Study
Test Organism:
Exposure Route:
Study Design:
Effects Data:
Bluegill sunfish (Lepomis macrochirus; embryos and larvae)
dietary and waterborne - field exposed
Resident adult bluegill were collected from Hyco Reservoir in Person County,
North Carolina and from a nearby control lake (Roxboro City Lake). Hyco
Reservoir is a cooling lake for Carolina Power & Light and receives the
discharge from the ash storage pond. For embryo/larval study up to swim-up
stage, control fish were collected from the unaffected portion of Hyco.
Repeat of 1982 28-day Embryo/Larval Study. Three crosses between: Hyco
female and Hyco male; control female with Hyco male; and control female with
control male. Gametes were fertilized and maintained for the 28-day test in ash
pond effluent dilutions of 0, 20 and 50 percent. Percent hatch, percent swim-up
success and survival were measured to 28 days post hatch. Two treatments were
replicated and fed zooplankton collected from Hyco-affected and Hyco-
unaffected (control). Larvae were observed for 28 days at which time survival
and weight were measured.
Embryo/Larval Study up to Swim-up Stage. Five crosses were made between fish
collected from the affected and unaffected areas. Percent hatch, percent swim-up
and survival were measured until swim-up (approximately 3-4 days after hatch).
28-day Embryo/Larval Study. All larvae that hatched from eggs obtained from
Hyco females died prior to completing swim-up (see table below).
Effect level (larval survival): <30, <33 and <59 mg Se/kg dw for adult female
bluegill in ovaries, liver and muscle, respectively
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Summary of 28-day embryo larval study
%
effluent
0
20
20
50
0
20
50
0
20
20
50
Parent
source in
cross
MXF
HXH
HXH
HXH
HXH
HXC
HXC
HXC
CXC
CXC
CXC
CXC
% hatch
92
98
92
97
89
96
60
79
90
88
72
% swim-
up
0
0
0
0
87
96
84
95
96
97
92
%
survival,
28-days
0
0
0
0
18
34
58
40
36
25
42
Adult tissue, mg Se/kg dw
Gonad
M
33
33
33
33
33
33
33
nd
nd
nd
nd
F
30
30
30
30
2.2
2.2
2.2
2.2
2.2
2.2
2.2
Liver
M
43
43
43
43
43
43
43
37
37
37
37
F
33
33
33
33
4.4
4.4
4.4
4.4
4.4
4.4
4.4
Muscle
M
62
62
62
62
62
62
62
27
27
27
27
F
59
59
59
59
2.7
2.7
2.7
2.7
2.7
2.7
2.7
Chronic Value:
<36.49 mg Se/kg dw in whole-body using the muscle to whole body equation.
<59 mg Se/kg dw muscle
<30 mg Se/kg dw ovary
Embryo/larval study to swim-up. Percent swim-up of larvae from parents
collected in non-affected Hyco averaged 93 percent, whereas percent swim-up
from larvae collected from affected Hyco was 12 percent. Effect levels were
determined for adult female and larval tissues. Larval tissues were averaged
across effluent concentrations (geometric mean).
Effect level (percent swim-up):
Adult female ovaries: >9.1 mg/kg dw; <30 mg/kg dw
Adult female liver: >26 mg/kg dw, <33 mg/kg dw
Adult female muscle: >25 mg/kg dw, <59 mg/kg dw
Larvae: >12.8 mg/kg dw; < 165 mg/kg dw
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Summary of Embyo/Larval Study up to Swim-up - Affected vs Unaffected Hyco
date
of
fert.
6-24
6-27
6-28
6-28
6-29
7-14
7-26
7-27
Parents'
capture
location in
Hyco
affected
affected
affected
affected
affected
unaffected
unaffected
unaffected
Percent hatch
at % effluent
0
93
99
29
98
88
92
99
76
20
98
88
34
86
93
80
94
84
50
94
77
35
91
85
84
93
86
Percent swim-up
at % effluent
0
0
0
25
5
59
79
100
100
20
0
0
14
0
42
92
98
89
50
0
0
3
0
25
89
98
91
Selenium in tissue, mg/kg dw
Adult female
Ovary
30
30
30
30
30
9.1
9.1
9.1
Liver
33
33
33
33
33
26
26
26
Muse
59
59
59
59
59
25
25
25
Larvae
0: 130
20: 120
0: 130
20: 120
0: 130
20: 120
0: 130
20: 120
0: 130
20: 120
0: 19
20: 11
50: 10
0: 19
20: 11
50: 10
0: 19
20: 11
50: 10
Chronic Value:
The chronic value estimated for the percentage larvae reaching the swim-up stage
is presented as a range:
>25 mg Se/kg dw (unaffected area) and <59 mg Se/kg dw muscle (affected area)
>30 mg Se/kg dw (unaffected area) and <9.1 mg Se/kg dw ovary (affected area)
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Bryson, W.T., K.A. MacPherson, M.A. Mallin, W.E. Partin, and S.E. Woock. 1985b. Roxboro Steam
Electric Plant Hyco Reservoir 1984 Bioassay Report. Environmental Services Section. Carolina Power &
Light Company
Ingestion Study
Test Organism:
Exposure Route:
Test Treatments:
Test Duration:
Study Design:
Effects Data:
Chronic Value:
Bluegill sunfish (Lepomis macrochirus; juvenile- hatchery raised)
Dietary only
5 diets: Se form (nominal selenium concentration in base diet)
seleno-DL-cystine (5 mg/kg)
seleno-DL-cystine (10 mg/kg)
seleno-DL-methionine (5 mg/kg)
sodium selenite (5 mg/kg)
Hyco zooplankton (5 mg/kg)
60 days
Each treatment contained 40 fish which were maintained in a flow-through
system. Fish were fed at 3 percent of their body weight. Length and weight were
measured on days 30 and 60. Total selenium was measured in liver and whole-
body.
No decreased length or weight in any of the Se-diets relative to the control.
all values are whole-body
seleno-DL-cysteine: >2.16 mg Se/kg dw
seleno-DL-cysteine-2X: >3.74 mg Se/kg dw
seleno-DL-methionine: >2.46 mg Se/kg dw
sodium selenite : >1.21 mg Se/kg dw
Hyco zooplankton: >2.35 mg Se/kg dw
Because none of the selenium-spiked diet formulations affected growth of
juvenile fish at the concentrations tested, the chronic value selected for this study
is >3.74 mg Se/kg dw for the seleno-DL-cysteine-2X formulation.
Source and Exposure Embryo-Larval Study
Test Organism: Bluegill sunfish (Lepomis macrochirus; Adults from Hyco and a control lake)
Exposure Route: Dietary and waterborne - field exposure
Test Treatments:
Test Duration:
Four treatments:
Hyco-collected fish exposed to Hyco water in flow through spawning tanks.
Hyco-collected fish in control water in flow through spawning tanks.
Control fish exposed to Hyco water in flow through spawning tanks.
Hyco-collected fish in control water in flow through spawning tanks.
Adult fish were in spawning tanks 4-7 months
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Study Design:
Effects Data:
Eggs from each treatment were observed for percent hatch and percent swim-up.
Fish collected from the control lake did not spawn. Percent hatch and percent
swim-up from Hyco fish in Hyco and control water are given in the table below.
The percent hatch and percent swim-up were >83 and >83 for all the Hyco fish
suggesting no effect for these endpoints.
Source of
parents
Hyco
Hyco
Control
Control
Se in parental
liver tissue,
mg/kg dw
18.6
18.6
13.8
13.8
Water type
for eggs and
larvae
Hyco
well water
Hyco
well water
N
16
10
a
12
Percent hatch
86.6
83.8
a
86.0
Percent
swim-up
91.1
95.5
83.3
97.4
a percent hatch unknown.
Chronic Value: The chronic value forthis study is >18.6 mg Se/kg dw liver tissue.
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Gillespie, R.B. and P.C. Baumann. 1986. Effects of high tissue concentrations of selenium on
reproduction by bluegills. Trans. Am. Fish. Soc. 115:208-213.
Test Organism:
Exposure Route:
Test Treatments:
Study Design:
Effects Data:
Chronic Value:
Bluegill sunfish, wild-caught (Lepomis macrochirus; adults; embryos and larvae)
dietary and waterborne - field exposure
High selenium adult fish were collected (electrofishing and with Fyke nets) from
Hyco Reservoir. Low selenium adult fish were collected from Roxboro City
Lake, Roxboro, NC.
All possible combinations of bluegill parents from Hyco Reservoir and Roxboro
City Lake were artificially crossed in June and July, 1982 and 1983, respectively.
Fertilization success was assessed by stripping subsamples of 100 to 500 eggs per
female and combining them with 2 ml of sperm. All zygotes were reared in
Roxboro City Lake water and percent fertilization was estimated 2-3 hours later
as the proportion of mitotically active zygotes. To estimate hatching success,
gametes were combined as before and subsamples of 100 to 300 embryos per
cross were transferred to egg cups and maintained in closed aquaria receiving re-
circulated Roxboro City Lake water. Percent hatch (approx. 2d at 22 to 25°C)
was based on the number of yolk-sac larvae. In 1982, about 200 embryos from 8
crosses were observed and preserved at intervals up to 40 h after fertilization, and
about 450 larvae were preserved at intervals of 40 to 180 h after fertilization. In
1983, about 1,800 larvae were observed and preserved from 40 to 150 hr from
crosses involving females from Hyco Reservoir, and about 40-300 hr for crosses
involving females from Roxboro City Lake (10 crosses total).
No significant differences were found in percent fertilization or in percent hatch
among parent combinations from the 18 crosses made in June 1982 and July
1983. In contrast, larvae from all crosses involving a Hyco female were
edematous; 100 percent of the larvae were abnormal in 7 of 8 crosses. Note: This
outcome was observed when the same female from Hyco Reservoir was crossed
with males from either Hyco Reservoir or Roxboro City Lake. The range of
selenium concentrations in the ovaries of Hyco Reservoir females used for the
cross experiments was from 5.79 to 8.00 (GM = 6.945 mg/kg ww; n=7). The
reported concentrations of selenium in ovaries and carcasses of females collected
from Hyco Reservoir in 1982 and 1983 were 6.96 and 5.91 mg/kg ww (n=22 and
28, respectively). The reported concentrations of selenium in ovaries and
carcasses of females collected from Roxboro City Lake in 1982 and 1983 were
0.66 and 0.37 mg/kg ww (n=14 and 19, respectively). The mean selenium
concentration in bluegill larvae (n=222) from artificial crosses of parents from
Hyco Reservoir was 28.20 mg Se/kg dw.
<46.30 mg Se/kg dw ovary using 85 percent moisture for ovaries measured in
study.
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Doroshov, S., J. Van Eenennaam, C. Alexander, E. Hallen, H. Bailey, K. Kroll, and C. Restrepo.
1992. Development of Water Quality Criteria for Resident Aquatic Species of the
San Joaquin River; Part II, Bioaccumulation of Dietary Selenium and its Effects
on Growth and Reproduction in Bluegill (Lepomis macrochirus). Final Report to
State Water Resources Control Board, State of California. Contract Number 7-
197-250-0.
Test Organism:
Exposure Route:
Test Duration:
Study Design:
Effects Data:
Bluegill sunfish (Lepomis macrochirus); Population A: selenium
bioaccumulation observations used 113 g (range 30-220 g) obtained from
Rainbow Ranch Fish Farm, California. Population B: spawning performance
observations used 106 g (range 65-220 g) females and 164 g (range 80-289 g)
males obtained from Chico Game Fish Farm.
Dietary only
Dietary
Seleno-L-methionine added to trout chow; the three nominal dietary
concentrations of 8, 18 and 28 mg/kg seleno-L-methionine were measured at 5.5,
13.9, and 21.4 mg/kg Se (moisture content 13 to 16%).
140 days
Population A fish and Population B females were fed nominal dietary treatments
8, 18 and 28 mg/kg seleno-L-methionine; Population B males were fed untreated
diets until the start of spawning. Population A fish were sampled on days 0, 30,
58, 86 and 114 for Se measurement. At least 3 females were sampled each event.
Fish remaining after day 114 were transferred to an outdoor pond fed untreated
diet and sampled on day 144 for depuration analysis.
On day 120 Population B males and females were paired for natural spawning
which had limited success. Fish were maintained in treatment tanks and females
were monitored for egg ripeness. When ripe, females were induced to ovulate
and ova were fertilized in vitro with semen stripped from males. Fertilized eggs
were sampled for fertilization success, Se content, and two live sub-samples for
bioassay, one a 30-day embryo-larval test and another for larval development
during first 5 days after hatching.
Larval development: after hatching, 100 larvae were transferred to beakers and
samples were examined daily for normal, abnormal and dead were recorded.
Larval bioassay: 90 fertilized eggs from each female were placed in groups of
approximately 30 eggs. Larvae and fry were fed rotifers and brine shrimp nauplii
through the 30 day observation.
Treatment effects were only observed on early development bioassays. In the 5-
day larval bioassay, systemic edema and underdeveloped lower jaw were
apparent in all larvae in the 21.4 mg/kg dietary treatment by day 3 and complete
mortality by day 5, except for two progenies where 10% of the larvae appeared
normal. No abnormalities were observed in control and 5.5 mg/kg treatment. 3 of
the 6 progenies in the 13.9 mg/kg treatment exhibited 10 to 20% larvae with
similar abnormalities (in table below). The average proportion of larvae with
edema were 5% in 13.9 and 95% in 21.9 mg/kg, both of these were statistically
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different from the control (0% edema).
In the 30-day larval survival bioassay, statistical difference was only in the
highest test treatment for survival and growth measurements, length and weight
(see table below). The proportion of abnormal larvae was higher in the selenium-
treated diets but was not significantly different from the control. The percent of
abnormal larvae in the 13.9 mg/kg treatment (7.2%) was only slightly higher than
the control (6.3%).
Authors present the effect level for bluegill at the 13.9 mg/kg dietary treatment
(NOEC 5.5 mg/kg) based on proportions of edema and delayed resorption of the
yolk sac. The latter endpoint is based on significantly greater yolk area and oil
globule area in the 13.9 and 21.4 mg/kg treated eggs.
The most sensitive endpoint, percent edema, as a function of selenium in egg dw
and adult muscle dw, was fitted to a logistic curve from which EC estimates were
calculated (see Figures 1 and 2). The ECi0 and EC2o values are given in the
following table.
Effect level
EC10
EC20
Egg, mg Se/kg dw
20.05
22.43
Maternal muscle, mg Se/kg dw
11.51
12.85
Chronic Value:
ECio value (edema) at 20.05 mg Se/kg egg dw or 11.51 mg Se/kg muscle dw
Chronic value is 20.05 :g Se/g eggs dw.
Selenium Concentrations (mg/kg dw) in Bluegills from Population A Day 113 of Bioaccumulation
Dietary
treatment
Ovarv
Female liver
Testis
Male liver
Control
2.17(0.05^1
2.51(0.32)
2.65 (0.21)
4.10(0.37)
5.5 mg/kg dw
10.89(1.83^1
NA
9.87
14.32
13.9 mg/kg dw
26.17(0.07^1
22.75 (2.96)
16.38(0.71)
24.28 (4.54)
21.4 mg/kg dw
40.32 (2.44^1
40.68(2.14)
29.70 (5.02)
52.47 (5.23)
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Selenium Concentrations (mg/kg dw) in Bluegill Parents (Population B) Used in Larval Toxicity
Tests
Dietary
treatment
Male liver
Testis
Female liver
Female muscle
Ovary
Eggs
Larvae
Fry
Control
4.07 (0.23)
1.87(0.11)
4.00 (0.26)
1.47(0.14)
2.23(0.11)
2.81 (0.14)
NA
1.48(0.11)
5.5 mg/kg dw
6.94 n. 58)
3.64 (0.47)
12.33 (1.09)
5.80 (0.79)
6.34 (0.47)
8.33 (0.63)
NA
1.25 (0.02)
13.9 mg/kg dw
20.46 (3.46)
9.96 (0.45)
25.98 (4.28)
10.41 (2.02)
14.10(2.62)
19.46 (3.83)
NA
1.37(0.06)
21.4 mg/kg dw
31.63(1.75)
15.25 (0.45)
47.60(4.11)
23.64 (2.04)
30.63 (3.23)
38.39(3.14)
35.30(4.16)
1.46(0.03)
5-day Larval Development Toxicity Test, average (SD)
Dietary
treatment
Edema. %
Control
0
5.5 mg/kg dw
0
13.9 mg/kg dw
5(2)*
21.4 mg/kg dw
95.7 (2.7)*
Results from 30-day Embryo-larval Toxicity Test, average (SD)
Dietary treatment
Larval survival, %
Larval length, mm
Larval weight, mg
Abnormalities in
larvae, %
Control
71 (8.5)
19.1(1.2)
114(24)
6.3 (7.9)
5.5 mg/kg dw
51.9(26.5)
19.9(1.2)
133 (27)
15.0(5.8)
13.9 mg/kg dw
64.4 (3.4)
19.3 (0.8)
119(16)
7.2(3.1)
21.4 mg/kg dw
2.5 (3.5)*
16.6 (2.5)*
81 (37)*
25.0 (43.3)
* statistically significantly different from control
C-101
Draft Document
-------
Thirty day toxicity test mortalities and tissue selenium concentrations in respective females, "n"
is number of eggs on Day 0, "r" is mortality on Day 30, "p" is proportions.
Progenies
08-2C
18-4C
5.5-1S
5.5-2S
5.5-6S
13.9-1S
13.9-3S
13.9-6S
21.4-1S
21.4-2S
21.4-3S
21.4-4S
21.4-5S
21.4-6S
21.4-7S
n
89
85
85
90
85
90
87
87
88
90
86
88
90
86
88
r
17
17
64
42
19
29
34
31
87
89
79
88
90
86
82
P
0.191
0.200
0.753
0.467
0.224
0.322
0.391
0.356
0.989
0.989
0.919
1.000
1.000
1.000
0.932
[Se], mg/kg dw (female)
Ovary
1.95
2.38
7.72
5.55
4.06
3.94
21.82
20.40
29.90
45.82
27.24
23.18
32.64
37.63
18.02
Liver
4.04
5.03
14.89
7.06
10.49
7.54
34.74
36.82
38.02
33.96
59.01
62.71
55.25
48.14
36.10
Muscle
2.25
0.95
7.07
5.80
1.41
2.75
15.44
16.58
NA
31.10
17.28
27.40
24.00
24.66
17.42
Eggs
3.54
3.25
11.49
8.31
6.18
8.55
22.06
30.20
44.02
36.31
25.21
52.18
42.40
38.47
30.12
C-102
Draft Document
-------
Bluegill sunfish (Doroshov et al 1992)
5
c
a>
o
120
100
80
60
40
20
.8 1.0 1.2 1.4
Log([Se] egg mg/kg dw)
Parameter Summary (Logistic Equation Regression Analysis)
Parameter Guess FinalEst StdError 95%LCL 95%UCL
LogX50 1.3830 1.4340 0.0283 1.0748 1.7933
S 1.3889 4.167 0.775 -5.681 14.014
YO 100.00 100.47 3.09 61.19 139.75
% Effect
50.0
20.0
10.0
5.0
Effect Concentration Summary
Xp Est 95%LCL 95%UCL
27.16 11.88 62.12
22.43 8.34 60.31
20.05 6.30 63.82
18.086 4.732 69.131
06/29/2009 20:28
MED Toxic Response Analysis Model. Version 1.03
C-103
Draft Document
-------
Bluegill sunfish (Doroshov et al 1992)
•s
120
100
80
| 60
40
20
.2 .4 .6 .8 1.0 1.2
Log([Se] adult muscle mg/kg dw)
Parameter Summary (Logistic Equation Regression Analysis)
Parameter
LogXSO
S
YO
Guess FinalEst
1.1522 1.1907
1.6314 4.240
100.00 100.04
StdError
0.0005
0.012
0.04
95%LCL 95%UCL
1.1840 1.1974
4.082 4.397
99.58 100.49
Effect Concentration Summary
% Effect
50.0
20.0
10.0
5.0
Xp Est
15.512
12.851
11.511
10.400
95%LCL
15.274
12.629
11.287
10.172
95%UCL
15.754
13.077
11.740
10.634
06/29/2009 20.33
MED Toxic Response Analysis Model, Version 1.03
C-104
Draft Document
-------
Hermanutz et al. 1992. Effects of elevated selenium concentrations on bluegills (Lepomis macrochirus)
in outdoor experimental streams. Environ. Tox. & Chem. 11: 217-224
Hermanutz et al. 1996. Exposure of bluegill (Lepomis macrochirus) to selenium in outdoor experimental
streams. U.S. EPA Report. Mid-Continent Ecology Division. Duluth, MN.
Tao, J., P. Kellar and W. Warren-Hicks. 1999. Statistical Analysis of Selenium Toxicity Data. Report
submitted for U.S. EPA, Health and Ecological Criteria Div. The Cadmus Group.
Test Organism:
Exposure Route:
Study Design:
Bluegill (Lepomis macrochirus; 3 to 4-year old adults)
Dietary and waterborne followed by dietary only
Dietary and waterborne
Selenite was added to artificial streams which entered the food web; thus, fish
were also exposed to selenium in the diet.
Dietary only
Recovering streams exposed bluegill to selenium in prey organisms. Selenite
addition to water was ceased (selenium in water was below detection level).
Eight Monticello artificial streams were used for three separate studies between
1987 and 1990.
Stream
Dates
BGa put in station 0-2
BG transferred to sta. 6
End of study
1
2
3
4
5
6
7
8
Study I
9-1-87
5-16-88
8-22-88
Unused
Unused
10 :g/L
30 :g/L
Control
30 :g/L
Control
10 :g/L
Study II
10-88
5-89
8-89
Control
2.5 :g/L
10 :g/L
Recovering
Control
Recovering
2.5 :g/L
10 :g/L
Study III
11-89
5-90
7-90
Control
Recovering
Recovering
Recovering
Control
Recovering
Recovering
Recovering
BG = Bluegill
C-105
Draft Document
-------
A schematic diagram of an artificial stream is provided below. For each study, a random sample of 22-50
adult bluegill were transferred from stations 0-2 (provided temperatures above 4°C during winter) to
station 6 (most suitable for nests) during mid-May for spawning. Spawning activity was monitored in the
streams. Embryo and larval observations were made in situ and in the laboratory from fertilized eggs
taken from the streams and incubated in the lab.
Schematic Design of One of the Artificial Streams in the Monticello Study
Station Number
inlet
I
Adults from fill to
mid-Mav
Adult barrier
Adult barrier
Adult barrier
Adults from mid -
May to end of study
C-106
Draft Document
-------
Effects on Progeny - Study Ia
Egg cup observations
treatment
control
control
10 :g/L
10 :g/L
30 :g/L
stream
5
7
3
8
4
ovary Se (mg/kg ww)
Up
NA
0.47
4.29
4.72
3.71
down
0.53
0.01
2.53
6.37
NA
geomean
0.53
0.07
3.29
5.48
3.71
ovary Se
(mg/kg
dw)b
2.21
0.29
13.73
22.85
15.46
Geomean
ovary Se
(mg/kg
dw)
0.79
17.71
15.46
% hatch
mean ± SD
93.3 ±9.1
71.5 ±22.5
60.3 ±25. 8
% survival
to 4th day
mean ± SD
69.7 ± 13.9
28. 8 ±23.1
9.1 ± 12.9
% edema
mean ± SD
0.1 ±0.2
80 ± 1.0
50.3 ±64.1
% lordosis
mean ± SD
1.8 ±2.6
11.6± 15.9
6.3 ± 1.8
% hemorr
mean ± SD
0.1 ±0.3
28.5 ±40.6
26.8 ±20.2
Nest observations
treatment
control
control
10 :g/L
10 :g/L
30 :g/Lc
stream
5
7
3
8
4
ovary Se (mg/kg ww)
up
NA
0.47
4.29
4.72
3.71
down
0.53
0.01
2.53
6.37
NA
geomean
0.53
0.07
3.29
5.48
3.71
ovary Se
(mg/kg
dw)b
2.21
0.29
13.73
22.85
15.46
Geomean
ovary Se
(mg/kg
dw)
0.79
17.71
15.46
# active
nests
mean ± SD
6.5 ±2.1
5.0 ±4.2
1.0 ± 1.4
# embryos
Collected
mean ± SD
1441 ±205
1282 ±457
361±510
% dead
Embryos
mean ± SD
0.9 ±0.03
3.2 ±2.9
0.4
# larvae
Collected
mean ± SD
3947 ± 1888
1169± 1093
157 ±222
% dead
Larvae
mean ± SD
3.0 ± 1.1
17.0±21.3
12.1
a Selenium concentrations in table were taken from Hermanutz et al. (1996); effect values were taken from Hermanutz et al (1992).
b used 76% moisture for egg/ovary in bluegill (average of Gillespie and Bauman 1986 and Nakamoto and Hassler 1992) to convert egg/ovary ww
to dw
0 No active nests, embryos, or larvae found in one of the 30 (ig/L streams. Therefore, N = 1 for % dead embryos and dead larvae in the 30 (ig/L
treatment
C-107
Draft Document
-------
Effects on Progeny - Study IIa
Egg cup observations
treatment
control
control
2.5 :g/L
2.5 :g/L
10 :g/L
10 :g/L
recSO
:g/L
recSO
:g/L
stream
1
5
2
7
3
8
4
6
No. of
trials
6
5
0
4
3
2
0
6
%
hatch
93.0
96.4
NA
81.4
83.3
91.1
NA
92.9
%
survival
to 3rd
day
75.2
71.5
NA
71.6
57.7
57.1
NA
73.0
%
edema
0
0
NA
0
100
100
NA
17.4
% lordosis
0
0
NA
0
11.1
18.2
NA
0
% hemorr
0
0
NA
3.6
49.3
41.1
NA
11.5
% healthy"
97.8
97.9
NA
92.2
0
0
NA
70.7
ovary Se (mg/kg ww)
up
1.02
1.09
2.02
6.96
5.87
down
0.76
0.76
1.82
3.36
8.1
12.6
13.2
avg
0.89
0.91
1.82
2.61
8.10
9.36
8.80
ovary Se
(mg/kg dw)c
3.72
3.79
7.58
10.86
33.75
39.02
36.68
Nest Observations
Treatment
control
control
2.5 :g/L
2.5 :g/L
10 :g/L
10 :g/L
R 30 :g/L
R 30 :g/L
Stream
1
5
2
7
3
8
4
6
#
active
Nests
6
9
1
5
2
3
0
8
#
embryos
Collected
2458
1329
0
1462
672
931
NA
646
% dead
embryos
0.94
0
0
0
0.32
NA
0
# larvae
collected
3252
3435
2497
4717
5376
750
NA
6782
% dead
larvae
0.03
1.05
0.20
0.08
0.50
0.40
NA
7.8
#samples
w larvae
7
13
3
8
9
4
NA
16
%
edema
0
0
4.1
0
81.4
50
NA
27.3
%
lordosis
0
0
25
0
5.0
14.7
NA
0
%
hemorr
0
0
77.6
52
55.5
26.7
NA
17.1
ovary Se (mg/kg ww)
up
1.02
1.09
2.02
6.96
5.87
Down
0.76
0.76
1.82
3.36
8.1
12.6
13.2
Avg
0.89
0.91
1.82
2.61
8.10
9.36
8.80
ovary Se
(mg/kg dw)c
3.72
3.79
7.58
10.86
33.75
39.02
36.68
a Selenium concentrations in table were taken from Hermanutz et al. (1996); effect values were taken from Tao et al. (1999).
b Among live larvae that survived up to third day after first larvae hatched; assumes the observations of multiple abnormality types always co-
occurred in the same organism. This may overestimate the actual % healthy when this assumption is violated.
0 used 76% moisture for egg/ovary in bluegill (average of Gillespie and Bauman 1986 and Nakamoto and Hassler 1992) to convert egg/ovary ww
C-108
Draft Document
-------
to dw
R = recovering stream
C-109 Draft Document
-------
Effects on Progeny - Study III"
Egg cup observations
treatment
control
control
R2.5 :g/L
R2.5 :g/L
R 10 :g/L
R 10 :g/L
R 30 :g/L
R 30 :g/L
Stream
1
5
2
7
3
8
4
6
number of
trials
2
3
3
6
3
5
% hatch
92
76.7
87.3
87.2
75.3
92
% survival
to 3rd day
58.6
69.2
66
76.5
74.5
78
% edema
0
0
0
0
0
% lordosis
0
0.9
0
0
0
% hemorr
0
0.8
0
0
0
ovary Se
(mg/kg ww)
1.2
0.93
1.84
1.97
6.25
2.44
3.82
ovary Se (mg/kg
dw)b
5.0
3.88
7.67
8.21
26.04
10.17
15.92
Nest observations
treatment
control
control
R2.5 :g/L
R2.5 :g/L
R 10 :g/L
R 10 :g/L
R 30 :g/L
R 30 :g/L
stream
1
5
2
7
3
8
4
6
# active nests
2
2
5
5
2
4
9
# samples
with larvae
5
3
5
2
4
4
13
% edema
0
0
0
0
0
0
0
% lordosis
0
0
0
0
0
0
0
% hemorr
0
0
0
0
0
0
0
ovary Se
(mg/kg ww)
1.2
0.93
1.84
1.97
6.25
2.44
3.82
ovary Se (mg/kg
dw)b
5.0
3.88
7.67
8.21
26.04
10.17
15.92
a The NOAEC for the study are from recovering 30 Og Se/L treatment.
b used 76% moisture for egg/ovary in bluegill (average of Gillespie and Bauman 1986 and Nakamoto and Hassler 1992) to convert egg/ovary ww
to dw
R = recovering stream
C-110
Draft Document
-------
Effects Data: The Hermanutz bluegill data from Study I&II were analyzed by
combining the nest and egg cup observations for percent hemorrhage, percent
lordosis, and percent edema in response to Se concentrations in parental ovaries
(mg/kg dw). Study I effects data were obtained from Hermanutz et al. (1992),
and corresponding Study I ovary Se concentrations were obtained from
Hermanutz et al. (1996). Study II effects and exposure data were obtained from
Hermanutz et al. (1996). Adult survival was not considered because it was very
low in both studies II and III. Prior to analysis, all percentages were transformed
(100-% value of response) so that the response variables decreased with
increasing Se. Endpoints and values used to parameterize TRAP are included in
the table below. All endpoints in the table below were analyzed using a three
parameter logistic equation model in TRAP with log transformed Se
concentrations. Aside from %larval survival, data from nest and egg cup
observations in Study II were combined with egg cup data from study I into a
single analysis. For percent survival, data from egg cup observations in Study I
and II were combined into a single analysis. No percent larval survival
measurements were made during nest observations in Study II. Nest observation
data for the relevant endpoints were not collected during Study I. Finally, neither
data from the two recovering streams in Study II, nor any of the Study III data
were included in these analyses. As stated on page 37 in the criteria document,
the recovery streams do not reflect the type of system to which water quality
criteria are most commonly applied; those receiving existing waterborne
pollutant discharges.
Of the endpoints evaluated, % edema was the endpoint that was most conducive
to regression analysis. Percent edema was also the most sensitive endpoint, with
an EC10 of 12.68 mg/kg ovary Se dw, and a 95% confidence interval of 8.47-
18.97 mg/kg ovary Se dw. TRAP results for %edema are shown below. The
EC 10 for % lordosis was 19.38 mg/kg ovary Se dw, but because of the large
standard error surrounding the slope and inflection point parameters of the
model, the corresponding confidence intervals (0.06-6103 mg/kg ovary Se dw)
were extremely large. Coupled with the fact that the incidence of lordosis was
low in both studies (less than 20% at the highest Se concentrations, with a
maximum value of 25%), this endpoint was determined to be less appropriate
than %edema. Model convergence for % hemorrhage could only be achieved in
TRAP at an unrealistic y-intercept value of 150, indicating a negative 50%
incidence of hemorrhage at a Se concentration of zero. Finally, model
convergence for %larval survival could not be achieved in TRAP.
Chronic Value: The chronic value for bluegill was calculated as the ECi0 value of 12.68 mg/kg
Se dw (larval edema in repose to Se concentration in the parental ovaries).
C-111 Draft Document
-------
Combined Nest and Egg Cup Observations for TRAP Analysis (Study I&II)
Study
II
II
II
II
II
II
II
II
II
II
II
II
I
I
I
Stream
1
1
5
5
2
7
7
7
3
3
8
8
5,7
3,8
4
Data
Source
Egg Cup
Nest
Egg Cup
Nest
Egg Cup
Nest
Egg Cup
Nest
Egg Cup
Nest
Egg Cup
Nest
Egg Cup
Egg Cup
Egg Cup
Se Treatment
Control
Control
Control
Control
2.5 Mg/L
2.5 ng/L
2.5 Mg/L
2.5 Mg/L
10 Mg/L
10 ng/L
10 ng/L
10 Mg/L
Control
10 Mg/L
30 ng/L
Ovary Se
(mg/kg dw)
3.72
3.72
3.79
3.79
7.58
7.58
10.86
10.86
33.75
33.75
39.02
39.02
0.79
17.71
15.46
%Edema
0
0
0
0
NA
4.1
0
0
100
81.4
100
50
0.1
80
50.3
%Lordosis
0
0
0
0
NA
25
0
0
11.1
5.0
18.2
14.7
1.8
11.6
6.3
% Hemorrhage
0
0
0
0
NA
77.6
3.6
52
49.3
55.5
41.1
26.7
0.1
28.5
26.8
%Survival
75.2
71.5
71.6
57.7
57.1
69.7
28.8
9.1
C-112
Draft Document
-------
Incidence of larval bluegill edema as a function of the logarithm of the selenium concentration in
parental ovaries.
120 r
100
co
CD
03
•a
o
o
80
60
40
20
.2 .4 .6 .8 1.0 1.2
Log Ovary Se(mg/kg dw)
1.4
1.6
1.8
Model Parameters (Logistic Regress ion Nonlinear Regression Model)
Initial Guess Final Estimate S.E. 95% LCL 95% UCL
logX50
S
Y-intercept
1.2
6.29
100
1.1905
6.2904
99.93
2.52E-02
4.6679
6.0437
1.135
-3.9836
86.628
1.2459
16.564
113.23
Effect Concentration Summary
% Effect ECx 95% LCL 95% UCL
50
20
10
5
15.504
13.657
12.68
11.842
13.646
10.232
8.4743
7.1031
17.615
18.229
18.974
19.743
C-113 Draft Document
-------
Coyle, J.J., D.R. Buckler and C.G. Ingersoll. 1993. Effect of dietary selenium on the reproductive
success of bluegills (Lepomis macrochirus). Environ. Toxicol. Chem. 12:551-565.
Test Organism:
Exposure Route:
Test Duration:
Study Design:
Bluegill sunfish (Lepomis macrochirus; two-year old pond-reared adult fish and
resultant fry)
Dietary and waterborne
Dietary
Seleno-L-methionine added in an aqueous solution to Oregon moist pellets;
moisture content of diet was 25 percent.
Waterborne
Flow through,10 Og Se/L nominal, 6:1 ratio of selenate:selenite, 98 percent
purity, adjusted to pH 2 with HC1 to prevent bacterial growth and change in
oxidation states of Se(FV) and Se(VI).
140 days
The experiment consisted of a test control and food control (see Test Treatment
table below) with fish (n=28 initially) in the four remaining treatments fed one of
the four seleno-methionine diets in combination with 10 Og Se/L in water.
Spawning frequency, fecundity, and percentage hatch were monitored during the
last 80 days of the exposure period. Survival of resulting fry (n=20) was
monitored for 30 days after hatch. Adults and fry were exposed in separate,
modified proportional flow-through diluters. Fry were exposed to the same
waterborne selenium concentrations as their parents. Adults were fed twice daily
ad libitum. Whole-body selenium concentrations in adult fish were measured at
days 0, 60, and were calculated from individually analyzed carcass and gonadal
tissue (ovaries and testes) at day 140. Eggs not used in percentage of hatch
determinations were frozen and analyzed for total selenium.
Measured Se in:
water
(Og Se/L)
diet
(mg Se/kg dw)
Test Treatments
1
(test control)
0.56
0.76
2
(food control)
8.4
0.76
3
10.5
4.63
4
10.5
8.45
5
10.1
16.8
6
11.0
33.3
C-l 14 Draft Document
-------
Effects Data:
There was no effect of the combination of highest dietary selenium concentration
(33.3 mg/kg dw) in conjunction with exposure to a waterborne selenium
concentration of 11.0 Og/L on adult growth (length and weight), condition factor,
gonad weight, gonadal somatic index, or reproductive endpoints (i.e., spawning
frequency, number of eggs per spawn, percentage hatch) during the 140-day
exposure. The mean corresponding whole-body selenium concentration in adults
exposed to this waterborne and dietary selenium combination was 19 mg/kg dw.
Survival of fry from the exposed adults was affected by 5 days post-hatch.
Concentrations of whole-body selenium in adult tissue at day 60 were used to
determine effects in the fry because eggs were taken for the larval tests beginning
at day 60 of the adult exposure.
Effects on Adults
Se in diet,
mg/kg dw
0.8
0.8
4.6
8.4
16.8
33.3
Se in water,
:g/L
0.5
7.9
10.5
10.5
10.1
10.1
whole -body
Se (140 d),
mg/kg dw
0.8
1.0
3.4
6.0
10
19
replicate
A
B
A
B
A
B
A
B
A
B
A
B
total no.
spawns
15
10
12
11
20
12
2
9
13
13
14
4
eggs/spawn
14,099
5,961
9,267
9,255
9,782
13,032
10,614
7,995
10,797
9,147
8,850
8,850
hatchability,
%
94.5
90.5
89.5
84.5
86.5
96.5
96.5
90
83
91.5
80
80
C-115 Draft Document
-------
Effects on Larvae
Se in diet, mg/kg
dw
0.8
0.8
4.6
8.4
16.8
33.3
Se in water, :g/L
0.5
7.9
10.5
10.5
10.1
10.1
egg, mg/kg dw
1.8
1.8
7.3
13
23
42
adult whole -body
(60 d), mg/kg dw
0.9
0.9
2.9
4.9
7.2
16
mean survival,
%
92
93
90
95
87
7
Chronic Value:
EC2o and ECio estimates using logistic equation with log transformation of
exposure:
effect level
EC20
ECio
egg, mg Se/kg dw
26.30
24.10
whole body, mg Se/kg dw
8.954
7.936
C-116 Draft Document
-------
Coyle et al. 1993 bluegill larval survival - TRAP logistic
SS
CO
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110 r
100
90
80
70
60
50
40
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20
10
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-
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\
\
\
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0 ,2 .4 .6 .8 1.0 1.2 1.4 1.6 1.8
Log([Se]egg mg/kg dw)
Parameter Summary (Logistic Equation Regression Analysis)
Parameter Guess FinalEst StdError 95%LCL 95%UCL
LogXSO 1.4626 1.4990 0.0137 1.4554 1.5427
S
YO
1.8812 5.041 0.529 3.358 6.723
92.50 92.50 1.05 89.15 95.85
Effect Concentration Summary
% Effect Xp Est 95%LCL 95%UCL
50.0 31.55 28.54 34.89
20.0 26.93 23.75 30.54
10.0 24.55 21.19 28.45
5.0 22.54 19.02 26.72
06/29/2009 14:04
MED Toxic Response Analysis Mode!, Version I 03
C-117 Draft Document
-------
Coyle et al. 1993 bluegill larval survival - TRAP logistic
110
100
80
- 70
CO
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CO
1 «
30
20
10
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0 .2 .4 .6 .8 1.0 1.2 1.4
Log([Se]adult whole body mg/kg dw)
Parameter Summary (Logistic Equation Regression Analysis)
Parameter Guess FinalEst StdError 95%LCL 95%UCL
LogXSO 1.0102 1.0408 0.0202 0.9764 1.1051
S 1.9483 3.851 0.452 2.412 5.290
YO 92.50 92.52 1.17 88.81 96.23
Effect Concentration Summary
% Effect
50.0
20.0
10.0
5.0
Xp Est
10.985
8.929
7.910
7.074
95%LCL
9.472
7.398
6.342
5.484
95%UCL
12.739
10.777
9.865
9.125
06/29/2009 14:10
MED Toxic Response Analysis Model, Version t.03
C-118 Draft Document
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Cleveland, L. et al. 1993. Toxicity and bioaccumulation of waterborne and dietary selenium in juvenile
bluegill sunfish (Lepomis macrochirus}. Aquatic Toxicol. 27:265-280.
Test Organism:
Life Stage:
Exposure Route:
Study Design:
Effects Data :
Bluegill sunfish (Lepomis macrochirus)
juvenile (5 months - waterborne exposure; 3 months - dietary exposure)
waterborne (60-d) and dietary (90-d) - separate exposures
waterborne - 6:1 selenate:selenite at 0.17, 0.34, 0.68, 1.38, 2.73 mg/L; dietary -
seleno-L-methionine in Oregon moist at 1.63, 3.25, 6.5, 13, 26 mg Se/kg dw)
Fish were exposed using a flow-through diluter. Each test consisted of an
exposure and a depuration phase. Whole body tissue measurements were made at
31 and 60 days of waterborne exposure and at 31, 59 and 90 days of dietary
exposure. Mortality and condition factor, K (weight x 105/length3), were reported
at selected intervals.
The waterborne exposure (see table below) was determined to have an EC2o =
4.07 mg Se/kg dw (1.96-8.44 mg/kg 95% CL). However, because it was a water-
only exposure, it was not considered in the derivation of the FCV. These data
nevertheless provide evidence that exposure route influences the tissue
concentration toxicity threshold, although the mechanistic explanation for this
phenomenon is lacking.
A mortality effect level for the dietary exposure could not be calculated because
the highest selenium whole body concentration (13.4 mg Se/kg dw) only had
17.5% mortality. The middle selenium concentration did have 22.5% mortality.
Cleveland et al. reported a significant decrease in K between 4.7 and 7.7 mg/kg
dw (see table below).
Waterborne Exposure Study
Measured selenium in
water (:g/L)
20 (control)
160
330
640
1120
2800
60-d measured
selenium in whole
body (mg/kg dw)
1.1
2.8
4
5.3
9.8
14.7*
60-d mortality (%)
10
12.5
22.5
52.5
70
97.5
Condition factor (K)
1.5
1.5
1.6
1.5
1.6
NA
*a 30-d measurement because all fish were dead at 60 days in this concentration.
C-119 Draft Document
-------
Dietary Exposure Study
Measured selenium in
food (mg/kg ww)
0.68 (control)
2.3
3.5
6.6
12.7
25
90-d measured
selenium in whole
body (mg/kg dw)
1
2.1
3.3
4.7
7.7
13.4
90-d mortality (%)
5
7.5
10
22.5
15
17.5
Condition factor (K)
1.3
1.3
1.3
1.3
1.2
1.2
Chronic Value:
Given (a) the very slight reduction in K (1.3 to 1.2 between 4.7 and 7.7 mg Se/kg
dw WB, with no further reduction at 13.4 mg Se/kg dw WB) and uncertain
relevance of growth data, and (b) no apparent concentration-related effect on
mortality between 4.7 and 13.4 mg Se/kg dw WB, the NOAEC is interpreted to
be 13.4 mg Se/kg dw for this study; and the chronic value is >13.4 mg Se/kg dw
whole body.
C-120 Draft Document
-------
Lemly, A.D. 1993a. Metabolic stress during winter increases the toxicity of selenium to fish. Aquatic
Toxicol. 27:133-158.
Test Organism:
Exposure Route:
Test Duration:
Study Design:
Effects Data :
Chronic Value:
Bluegill sunfish (Lepomis macrochirus; juvenile 50-70 mm)
Waterborne and dietary
Water
1:1 selenite:selenate in stock at pH 2; metered in to reach 5 :g/L
Diet
seleno-L-methionine in TetraMin (5 mg/kg dw)
180 days
Fish were exposed (treatment and control) under intermittent flow-through
conditions for 180 days. Tests were run at 4E and 20EC with biological
(histological, hematological, metabolic and survival) and selenium measurements
made at 0, 60, 120 and 180 days. Fish were fed at a rate of 3% body weight per
day. All treatments were initiated at 20EC and then decreased in the cold
treatment at a rate of 2EC per week for 8 weeks to reach 4EC and then
maintained at that temperature for the remainder of the 180 days.
In the 20EC test, fish accumulated 6 mg/kg dw selenium (whole-body) with no
significant effect on survival (4.3% and 7.4% mortality in control and treatment,
respectively). In the 4EC test, fish exposed to selenium accumulated 7.9 mg/kg
dw (whole-body) selenium and had significant mortality after 120 (33.6%) and
180 days (40.4%) relative to control (3.9%). Several hematological
measurements were significantly different in both the warm and cold selenium
exposures relative to controls. Both warm and cold selenium treatments also had
greater O2 consumption than controls. Fish lipid content in the cold Se treatment
decreased more than the cold control; lipid content did not decrease in either the
warm control or the warm Se treatment (see summary tables below). The results
suggest significant mortality occurs in juvenile bluegill during winter months
when tissue concentrations reach 7.91 mg/kg dw and lipid levels decrease to 6
percent.
20EC, >6 mg Se/kg whole-body; 4EC, <7.91 mg Se/kg dw whole body
Comments: See "Comparison of the Cold-Temperature Bluegill Juvenile-Survival Studies" in
this appendix after presentation of the Mclntyre et al. (2008) study.
C-121 Draft Document
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Mean Concentration of Selenium in Tissues, Cumulative Survival*, Percent Lipid Content and Oxygen Consumption in Juvenile
Bluegill
day
0
60
120
180
cold - Se control
Sea
1
1
1.1
1.4
Surv.
%
100
97.1
97.1
97.1
lipid,
%
13.2
12.5
11.5
10.5
02b
98
58
57
57
cold + Se
Sea
1
5.8
7.9
7.9
Surv.
%
100
92.9
66.4
59.6
lipid,
%
13.2
10
6
6
02b
98
63
81
78
warm - Se control
Sea
1
1.2
1.1
1.2
Surv.
%
100
95.7
95.7
95.7
lipid,
%
13.2
13.3
13.4
13.6
02b
98
98
100
100
warm + Se
Sea
1
5.8
6
6
Surv.
%
100
100
96.7
92.6
lipid,
%
13.2
13.3
13.4
13.5
02b
98
103
120
120
a whole body Se tissue concentration, mg/kg dw
b oxygen consumption, mg/kg/hr
* Cumulative Survival: In this experiment, 240 juvenile bluegill were placed in three 400-L fiberglass tanks, 80 in each, and exposed to
each control and treatment for a period of 180 days. Ten fish were removed at random from each treatment replicate on days 0, 60, 120,
and 180 for selenium, histological, hematological, and metabolic measurements.
Replicate and Average Whole-body concentrations (mg/kg dry weight) of selenium in juvenile bluegill *
replicat
e
c+Se
w+Se
c-Se
w-Se
dayO
1
0.87
1.17
0.89
0.99
2
1.21
0.96
3
0.95
0.90
mean
1.01
1.01
0.89
0.99
day 60
1
6.30
5.61
0.97
1.12
2
5.49
6.19
3
5.76
5.43
mean
5.85
5.74
0.97
1.12
day 120
1
8.36
6.37
1.01
0.99
2
7.31
5.92
3
7.85
5.50
mean
7.84
5.93
1.01
0.99
day 180
1
7.53
5.48
1.10
0.96
2
8.01
5.72
3
8.19
6.02
mean
7.91
5.74
1.10
0.96
C-122
Draft Document
-------
* Each value is for a composite sample made from 5 fish.
The Kaplan-Meier estimator was used to calculate survival at time t
S(t) =
where r(t,) is the number offish alive just before time th i.e. the number at risk, and dt is the number of deaths in the interval /, = [th ti+l].
The 95% confidence interval for such estimate (Venables and Ripley 2002) was computed as
where
±k
" H(t)
and j #
C-123
Draft Document
-------
The following table lists the estimates of survival in the cold + Se treatment at 60, 120 and 180 days. The term n.event is the number of
deaths at a given interval; n.risk is the number of organisms alive at the beginning of the interval; survival is computed by the Kaplan-
Meier estimator.
Time
60
120
180
n.risk
210
165
88
n. event
15
47
9
survival
0.929
0.664
0.596
std.err
0.0178
0.0350
0.0381
lower 95% CI
0.884
0.590
0.517
upper 95% CI
0.956
0.728
0.666
Hematological Measurements in Juvenile Bluegill Sunfish (indicates significantly different from control)
Warm Exposure
blood parameter
total erythrocyte, 106/ml
% mature
nuclear shadows, 104/ml
total leucocytes, 104/ml
% lymphocytes
% neutrophils
hematocrit, %
MCHC (mean corpuscular hemoglobin
cone.)
Cold Exposure
dayO
warm-Se
2.95
85
0.95
17.22
23
15
37
23
warm+Se
2.92
86
0.86
17.41
25
13
36
25
dayO
day 60
warm-Se
2.96
86
0.97
16.90
20
14
37
25
warm+Se
2.93
93*
2.05*
17.55
23
15
29*
19*
day 60
day 120
warm-Se
2.99
86
0.83
16.73
19
17
36
25
warm+Se
2.95
94*
2.38*
17.62
26
19
29*
18*
day 120
day 180
warm-Se
2.96
85
0.91
17.05
21
17
38
25
warm+Se
2.89
94*
2.30*
17.36
22
16
28*
17*
day 180
C-124
Draft Document
-------
blood parameter
total erythrocyte, 106/ml
% mature
nuclear shadows, 104/ml
total leucocytes, 104/ml
% lymphocytes
% neutrophils
hematocrit, %
MCHC (mean corpuscular hemoglobin
cone.)
MCV (mean corpuscular volume)
cold-Se
2.91
84
0.86
16.48
17
13
39
26
182
cold+Se
2.93
82
0.84
16.88
16
12
37
25
171
cold-Se
2.97
87
0.83
16.79
16
15
40
25
188
cold+Se
2.90
95*
2.30*
16.91
17
11
30*
18*
146*
cold-Se
3.01
85
0.89
16.80
19
15
41
22
180
cold+Se
2.95
96*
2.49*
16.74
15
12
28*
17*
135*
cold-Se
3.00
85
0.90
16.96
19
12
39
23
185
cold+Se
2.99
97*
2.36
16.63
18
14
27*
17*
130*
C-125
Draft Document
-------
Mclntyre et al. 2008. Effect of Selenium on Juvenile Bluegill Sunfish at Reduced Temperatures. US
EPA, Health and Ecological Criteria Division. EPA-822-R-08-020
Test Organism:
Exposure Route:
Test Duration:
Study Design:
Bluegill sunfish (Lepomis macrochirus}; juvenile; average length 47 mm,
average weight 1 g
Waterborne and dietary
Water
1:1 selenite:selenate; For exposure systems (ES) 1 and 3, fish were exposed to a
control and a series of 6 nominal concentrations, 1.25, 2.5, 5, 10, 20 and 40 :g
Se/L. For ES2, fish were exposed to a control and one nominal concentration, 5
:g Se/L.
Diet
For ESI and ESS, fish were fed a series of six concentrations of selenium and a
background control in Lumbriculus variegatus. The measured selenium
concentrations in the L. variegatus treatments in ESI were: 2.3 (control), 4.5, 5.3,
7.5, 14.2, 25.7 and 34.9 mg Se/kg dw; in ES3: 2.2 (control), 4.2, 5.0, 7.2, 15.2,
25.4 and 46.7 mg Se/kg dw. Fish were fed worms at a rate of 4% of the current
biomass in each fish tank. Selenium was accumulated in L. variegatus by feeding
the worms in separate tanks a series of six concentrations of selenized-yeast
diluted with nutritional yeast: 1.7, 3.3, 6.7, 13.3, 26.7 and 53.5 mg Se/kg dw.
Control worms were fed nutritional yeast only. Each tank was additionally
exposed to the associated aqueous concentration selenium, e.g., the worms fed
the 1.7 mg Se/kg dw selenized yeast were exposed to 1.25 :g Se/L, the worms fed
the 3.3 mg Se/kg dw selenized yeast were exposed to 2.5 :g Se/L, and so on.
For ES2, fish were fed TetraMin spiked with seleno-L-methionine at a nominal
concentration of 5 mg/kg dw and at a rate of 3% of the current biomass in each
tank.
182 days
Juvenile bluegill were exposed concurrently to selenium using three separate
exposure systems, ESI, ES2 and ES3. In ESI and ES3, 100 fish were exposed to
each of 6 selenium treatments (low through high treatments are referred to as
Treatments 1 through 6) and two controls in 200 L carboys under flow-through
conditions. Each treatment consisted of an aqueous selenium concentration and
an associated dietary selenium concentration, e.g., the fish in the lowest ESI
treatment were exposed to 1.25 :g Se/L and fed worms containing 4.5 mg Se/kg
dw (see Exposure Route for other treatment concentrations). Temperature was
controlled in each system through the immersion of the carboys in a temperature-
controlled water bath and by controlling the temperature of the dilution water
being added to the carboys. The temperature in ESI was maintained at 20°C for
the first 30 days of exposure, and then decreased 2°C/week until it reached 4°C
(test day 79) at which point temperature was maintained until test termination
(test day 182). The only difference between ESI and ES3 was temperature was
decreased 2°C/week until it reached 9°C (test day 65) at which point temperature
was maintained until test termination (test day 182).
C-126 Draft Document
-------
The exposure of ES2 was similar to ESI and ESS in that 100 juvenile bluegill
were exposed to treatment in 200 L carboys under flow-through conditions. The
ES2 selenium treatment consisted of two replicates of 5 :g Se/L waterborne and 5
mg Se/kg dw diet (Tetramin). Two controls were maintained with ES2. The
temperature regime for ES2 was identical to ES 1 .
Observations on fish behavior and mortality were checked daily. Total selenium
was measured in each fish tank weekly and selenium speciation was measured
monthly in each fish tank. Whole body total selenium was measured in the
worms from each tank (2 replicate 5 g samples) on test days 0, 30, 60, 112 and
182 and in the bluegill from each tank (3 replicates of 3 -fish composites - total 9
fish) on test days 0, 7, 30, 60, 112 and 182. The standard length and weight of
each fish was measured on each sample day. Lipid content was measured in fish
at day 0 and from each treatment at test termination.
Effects Data: Selenium increased in bluegill as the exposure concentrations increased (see
following table). No meaningful mortality was observed in ES2. The number of
fish that died in ES2 during the 1 82 day test were two fish in one treatment
replicate and none in the other treatment replicate; no deaths were reported in
ES2 controls. Significant mortality of juvenile bluegill was observed in ESI and
ES3. After 182 days, a total of 24 and 68 fish died in Treatments 5 and 6,
respectively in ESI; and a total of 38 and 61 fish died in Treatments 5 and 6,
respectively in ES3. See table below for mortalities in all treatments. Estimates of
bluegill survival were adjusted for the removal of individuals from the test
population. Individuals were removed from the experiments before test
completion, for sampling tissue concentrations or because they suffered
accidental deaths unrelated to selenium toxicity. For such data, it was necessary
to account for the reduction in number of individuals at risk of death due to
selenium over time. If r(t,) is the number of individuals at risk just before time tt
and dt is the number of deaths in the interval, It = [tt, ti+\), then survival (S) at time
t can be estimated as
The product (P) was calculated for each period in which one or more deaths
occur. The equation is the Kaplan-Meier estimator (Venables and Ripley 2002).
This correction was applied to calculate the proportion of survival in treatments
with ten or more deaths (10% mortality). The table below provides the adjusted
proportion and surviving bluegill in each treatment along with the concentration
of selenium in bluegill at test termination. The values in this table were used to
calculate the EC20 and ECi0 values using the TEAM software. Growth and lipid
content of the bluegill was not negatively affected by the selenium exposures.
C-127 Draft Document
-------
Measured total selenium concentrations in bluegill sunfish for all treatments and controls in Exposure System 1, 2 and 3.
Total Selenium in Whole Body Bluegill Tissue, mg/kg dw
ES
1
Test Day
0
7
30
60
112
182
ES
3
Test Day
0
7
30
60
112
182
ES
2
Test Day
0
7
30
60
112
182
Control
Average (SD)
1.93 (0.21)
2.43(0.31)
2.10(0.21)
2.11 (0.02)
1.98(0.04)
2.08(0.10)
Control
Average (SD)
1.93 (0.21)
2.50(0.10)
2.24 (0.41)
2.70 (0.22)
2.16(0.14)
1.67(0.21)
Control
Average (SD)
1.93 (0.21)
2.19(0.19)
2.49(0.15)
1.53 (0.03)
1.57(0.01)
1.38(0.06)
Treatment 1
Average (SD)
1.93 (0.21)
2.48(0.11)
2.85 (0.10)
2.70 (0.20)
3.16(0.11)
2.56(0.21)
Treatment 1
Average (SD)
1.93 (0.21)
2.60 (0.29)
2.44 (0.26)
2.88 (0.08)
2.49(0.10)
3.20 (0.27)
5A
Average (SD)
1.93 (0.21)
3.55 (0.25)
7.05 (0.76)
8.23 (1.55)
8.97(1.28)
9.41 (1.63)
Treatment 2
Average (SD)
1.93 (0.21)
2.43 (0.18)
3.10(0.04)
3.07 (0.05)
3.41 (0.08)
3.15 (0.25)
Treatment 2
Average (SD)
1.93 (0.21)
2.38(0.10)
2.70(0.16)
3.04 (0.39)
3.10(0.12)
3.83 (0.47)
5B
Average (SD)
1.93 (0.21)
3.08 (0.50)
7.51 (1.18)
8.09 (0.67)
9.45 (1.73)
10.61 (0.38)
Treatment 3
Average (SD)
1.93 (0.21)
2.64 (0.06)
2.94(0.13)
3.69 (0.25)
3.99 (0.26)
4.02 (0.21)
Treatment 3
Average (SD)
1.93 (0.21)
2.82 (0.20)
3.13(0.10)
3.79 (0.24)
3.64(0.16)
5.48 (0.24)
Treatment 4
Average (SD)
1.93 (0.21)
2.72 (0.07)
4.24 (0.22)
5.21 (0.30)
6.42 (0.05)
6.72 (0.09)
Treatment 4
Average (SD)
1.93 (0.21)
3.19(0.33)
3.95 (0.16)
5.54(0.21)
6.54(0.21)
9.38 (0.63)
Treatment 5
Average (SD)
1.93 (0.21)
3.27 (0.27)
6.62 (0.23)
8.62 (0.45)
11.60(0.43)
10.71 (0.55)
Treatment 5
Average (SD)
1.93 (0.21)
4.29 (0.20)
6.06 (0.36)
9.50(0.91)
11.50(0.25)
16.01 (0.30)
Treatment 6
Average
(SD)
1.93 (0.21)
4.27 (0.44)
10.21 (0.36)
12.66 (0.45)
Treatment 6
Average
(SD)
1.93 (0.21)
6.13 (0.62)
11.07(0.92)
15.14(0.96)
17.24 (0.30)
C-128 Draft Document
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Total number of deaths in ESI and ES3 Treatments throughout the experiment's duration (182
days). Both ESI and ES3 had two control tanks.
Treatment ESI ESS
Control (#1, #2) 0,7 1,1
1 5 0
2 1 1
3 00
4 33
5 24 38
6 68 61
The concentration of selenium in bluegill and the adjusted proportion of surviving fish at the end of
the 182 day exposure.
ESI ES3
Treatment [Se]tlssue, mg/kg dw surv
control 2.08 0.962
1 2.56 0.988
2 3.15 0.984
3 4.02 1.000
4 6.72 0.962
5 10.71 0.497
6 12.66 0.075
[Se]tlssue, mg/kg dw
1.67
3.20
3.83
5.48
9.38
16.01
17.24
surv
0.988
1.000
0.988
1.000
0.960
0.435
0.168
Chronic Value:
The NOAEC for bluegill in ES2 was calculated as the geometric mean of the
concentration of bluegill in the two replicates at the end of the exposure period,
9.992 mg Se/kg dw whole body. The chronic value for ES2 is therefore >9.992
mg Se/kg dw whole body. The EC2o and ECi0 values for ESI and ES3 are given
in the following table.
EC2o mg Se/kg dw
ECio mg Se/kg dw
ESI (4°C)
Whole body
9.78
9.27
ES3 (9°C)
Whole body
14.64
14.00
C-129 Draft Document
-------
Comparison of the Cold-Temperature Bluegill Juvenile-Survival Studies of Lemly (1993a) and
Mclntyre et al. (2008)
The Lemly (1993a) and Mclntyre et al. (2008) cold-temperature juvenile bluegill studies are summarized
on the previous pages. This discussion compares and contrasts these studies.
Both studies indicated that juvenile bluegill are more sensitive to selenium at lower temperature than at
higher temperature. For a 4°C temperature regime, the EC10 of 9.27 mg Se/kg dw WB obtained with
Mclntyre 's selenized yeast-worm -fish dietary bioaccumulation system is somewhat similar to the
threshold of 5.85 mg Se/kg dw WB estimated from the time course of bioaccumulation and mortality in
Lemly' s single treatment with seleno-L-methionine in TetraMin. These chronic values differ by a factor
of 1.58.
The difference in diet does not appear to explain the modest difference in results; however, since
Mclntyre's other 4°C experiment (Exposure System ES2), which used Lemly's seleno-L-methionine in
TetraMin diet, experienced no significant toxicity, whereas Lemly's similarly exposed fish experienced
40 percent mortality by the end of the test. In addition to the difference in observed mortalities,
bluegill in the 4°C selenium exposure decreased in both lipid content and body condition over the 180
days whereas no decreases in these measurements were observed in the Mclntyre et al. study, although
the fish used in both studies were of comparable size and body condition at test initiation: 47 mm average
standard length (range 44 to 54 mm) and a body condition index (100 x fish weight/standard length) of
3.2 in ES2 compared to 50 to 70 mm total length and a body condition factor of 3.9 in Lemly.
There are several possible reasons why such results could differ between studies. (1) ES2 maintained
exposure at 20°C for the first 30 days of exposure before decreasing the temperature compared to 7 days
in the Lemly study. (2) Lemly measured O2 consumption by removing and reintroducing test fish to the
test tanks, which was not done by Mclntyre et al. (3) The two studies differed in photoperiod - Lemly
"began with a 16: 10 h light/dark photoperiod which was gradually reversed to 10: 16" (sic) whereas
Mclntyre et al. used a fixed photoperiod of 16:8. (4) Some genetic differences between the tested batches
of organisms may be expected, reflecting different origins, despite the similarities in their starting size
and condition.
The modification to maintain 20°C for 30 days was to allow a longer period of time for the fish to
accumulate selenium during a warmer condition prior to decreasing the temperature. This did result in
shortening the exposure in ES2 at 4°C by 19 days (103 days at 4°C) compared to 122 days at 4°C in
study. However, as the majority of deaths in Lemly^is study occurred between in the middle 60
days of the 180-day test, the slightly shorter cold period in the Mclntyre study would not explain the
differences in mortalities.
As stated above, Lemly removed fish (N = 15) from each treatment for oxygen consumption
measurement and then returned these fish to the exposure tanks. There is the possibility that the fish
removed from the cold plus selenium treatment were sufficiently stressed by the exposure conditions that
the additional handling stress contributed to the mortality observed in this treatment. Between test days 60
and 180, 56 fish died Lemly's cold plus selenium treatment. Even if stress due to handling affected all the
fish used in the oxygen consumption measurements (up to 30 fish), it does not explain all the mortality
that was observed and therefore does not explain the difference between the two studies.
C-130 Draft Document
-------
It may then be questioned whether the fixed photoperiod alone could account for the differences in the
results of the two studies. More explicitly, did the longer light period in Mclntyre et al. photoperiod allow
the fish to feed more than the fish exposed to the shorter light period in the Lemly study, such that lipid
and body condition in the Mclntyre et al. fish were maintained and therefore not susceptible to "winter
stress syndrome." The effects of photoperiod on fish and other ectotherms are well-documented.
Temperature-independent seasonal changes in fish have been reported for growth and food conversion
efficiency (Biswas and Takeuchi 2003; Jonassen et al. 2000; Simensen et al. 2000), feeding behavior
(Volkoff and Peter 2006), metabolic rate (Evans 1984), and reproduction (Koger et al. 1999; Scott 1979).
Some of these studies have found conflicting results on the effect of photoperiod on growth (Fuchs 1978;
Jonassen et al. 2000; Simensen et al. 2000). Coupled with temperature being a dominant factor in
controlling physiological functions in temperate-zone fish as indicated by a 3 to 4-fold fluctuation in
metabolic activities over 10°C (Brett 1970; Fry 1971), it is difficult to use literature findings to explain
the difference in the two bluegill studies.
Observational recordings of the feeding behavior in Mclntyre et al. noted that in both control replicates
and in both treatment replicates the feeding of the juvenile bluegill went from active to not active on test
day 78 when temperatures were decreased from 6.6 to 5.8°C. The feeding observations are reflected in a
gradual slight decrease in the body condition factor (K) after test day 60 in the figure below. Although
food intake was not quantified during the study, the lack of growth indicated in K suggests feeding
markedly decreased as the temperature declined, as shown in the figure. Body condition decreased much
more in the Lemly's cold plus selenium exposed fish after test day 60 (approximately 50%) but K in his
cold-without-selenium exposure decreased only slightly, similar to Mclntyre et al. Therefore it is not
possible to determine if the greater decrease in K and in lipid content in Lemly's cold plus selenium
treatment was due to decreased feeding because of a shorter photoperiod or because the bluegill fish
population used in his study were more sensitive to selenium in cold conditions. Mclntyre et al. obtained
bluegill from Osage Catfisheries in Missouri whereas Lemly collected fish from ponds (assumed to be
near Blacksburg, Virginia, not stated in paper). The fish obtained from Missouri, a location with colder
winters than Virginia, may have been better adapted for withstanding colder winter temperatures than
Lemly's fish and therefore were less sensitive to "winter stress syndrome" as induced by selenium
exposure. Similarly, different populations of a species can have varying sensitivities to stressors.
Furthermore, the relative difference in the Lemly and Mclntyre et al. results is slightly lower than Delos
(2001) found to be typical when equivalent toxicity tests of the same species are compared. There should
thus be no expectation that the two study results should agree more closely than they do.
C-131 Draft Document
-------
CO
LL
g
'•^
T3
O
5 -
4 -
3 -
1 -
Condition Factor (K)
Temperature (°C)
22
- 20
- 18
- 16
- 14
- 12
- 10
- 8
- 6
- 4
O
o
CD
3
Q.
CD
50
100
150
200
Test Day
Relationship between body condition factor (K) and temperature in juvenile bluegill fed a diet of Se-
enriched TetraMin in the Mclntyre et al. (2008) study.
Both Lemly (1993) and Mclntyre et al. (2008) showed reduced survival of juvenile bluegill exposed to
elevated selenium under lab-simulated winter conditions, albeit at somewhat different concentrations. But
only Lemly, not Mclntyre et al., found the decreased survival to be accompanied by loss of lipid and body
condition. Such loss is not generally corroborated by field evidence (Janz 2008). Several studies have
measured growth and energy storage indicators in juvenile fish just prior to and just after winter at
reference sites and sites with elevated selenium in northern Canada (Bennett and Janz 2007a, b; Kelly and
Janz 2008; Driedger et al 2009; Weber et al. 2008). The growth (length, weight, condition factor, muscle
J£NA:DNA ratio, muscle protein) and energy storage (whole body lipids, whole body triglycerides, liver
triglycerides, liver glycogen) indicators for five fish species (northern pike, burbot, fathead minnow,
creek chub, white sucker) measured just after winter were similar or greater than those measured just
before winter at the selenium exposed sites. The slimy sculpin did show a decrease in whole body
triglycerides, but the reduction was similar at exposed and reference sites.
C-132 Draft Document
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Carolina Power & Light. 1997. Largemouth Bass Selenium Bioassay- Report. Carolina Power & Light
Company, Environmental Services Section, 3932 New Hill, North Carolina. December 1997
Test Organism:
Exposure Route:
Test duration:
Study Design:
Effects Data:
Largemouth bass (Micropterus salmoides)
Laboratory; dietary exposure only; DL-selenomethionine added to an artificial
diet. Adult largemouth bass obtained from a commercial supplier were fed
several months prior to spawning a series of selenium concentrations in the
artificial diet.
Embryo-larval monitoring through swim-up stage.
Dietary exposure studies were conducted in 1995 and in 1996. In 1995, the
measured dietary Se concentrations were 0.9 (control), 2.9, 7.5 and 11.2 mg
Se/kg dw; in 1996, they were 26.7, 53.1 and 78.4 mg Se/kg dw. Parent fish were
fed to satiation twice per day. Approximately 100 eggs from each spawn were
transferred to each of 2 to 4 incubation cups. Eggs and larvae were monitored for
mortality and deformities up to the larval swim-up stage. Selenium was measured
in the liver, muscle and gonad tissues of the parent fish. All live deformed larvae
at swim-up stage were considered as mortalities in the analyses.
Over the two year period, 56 successful spawns were obtained across all dietary
treatments. Live larval fish with deformities (kyphosis, scoliosis, jaw gap, and
lordosis) and edema at swim-up stage were considered mortalities for data
analysis. The average concentration of selenium in ovaries ranged from 3.1
mg/kg dw in the control to 77.6 mg/kg dw in the high dietary treatment (Table 1).
Larval survival generally decreased as the selenium concentration in the ovary
increased (Table 1; Figure 1). A plot of the percent survival of larval largemouth
bass as a function of the logarithm of selenium concentration in the parental
female ovary using TRAP produced an EC 10 of 20.35 mg Se/kg dw (Figure 1).
Effect
Concentration:
20.35 mg/kg dw in ovaries
C-133 Draft Document
-------
Table 1. Selenium concentrations in the diet, ovary and muscle tissues and the percent mortality
and deformities.
Measured Se in
diet fed to parents,
mg/kg dwa
0.9±0.1
(0.7- 1.3)
2.9 ±0.5
(2.1-3.8)
7.5 ±0.6
(6.3 - 8.4)
11.2±1.4
(9.3-14.1)
26.7 ±1.7
(23.6-29.5)
53.1 ±4.8
(45.5-61.9)
Spawn No.
6
12
13
26
34
35
3
4
10 (2F)
13
14
9
12
15
18
1
2
5
7
8
16
19
6
11
17
2
5
11
16
17
19
36
37
51
52
22
25
30
31
32
41
48 (2F)
50 (2F)
55
Se in parent ovary, mg/kg dw
Individual
5.38
7.34
3.51
5.74
1.58
1.36
2.09
1.85
2.11
1.86
1.40
9.59
8.03
9.73
7.66
8.43
25.15
15.31
1.20
6.78
8.25
10.20
35.44
15.08
24.59
37.14
44.67
34.26
35.58
33.48
48.24
35.81
37.88
32.95
59.89
46.22
70.45
81.62
54.99
53.96
51.48
84.31
32.87
73.33
Average
3.1
8.8
10.8
25.0
40.0
61.0
Larval survival, %
Individual
75.5
99.7
96.2
88.9
99.5
96.8
98.8
100
97
97.1
98.4
84.9
100
98.5
95.9
75
63.9
90.6
79.1
95
96.8
100
91.5
77.9
96.7
91.2
0
75.9
0
9.9
0
6.3
0
0
0
0
0
0
0
0
0
0
0
0
Average
95.3
94.8
85.8
88.7
18.3
0
C-134 Draft Document
-------
Measured Se in
diet fed to parents,
mg/kg dwa
78.4 ±4.3
(73.2-87.0)
Spawn No.
4(2F)
7
8
10
18
21
24
28
38
44
47
49
Se in parent ovary, mg/kg dw
Individual
66.81
56.98
86.49
65.99
72.35
71.89
62.44
99.02
52.37
102.82
88.15
105.29
Average
77.6
Larval survival, %
Individual
66
0
0
0
0
0
0
0
0
0
0
0
Average
5.5
± standard error; range of concentrations in parentheses.
C-135 Draft Document
-------
Figure 1. Percent larval survival as a function of the logarithm of the selenium concentration in
the parental ovaries.
1MB all data
120 r
100
60
_ro 40
.5 1.0 1.5 2.0
Log(0vary Se, mg/kg dw)
Parameter Summary (Logistic Equation Regression Analysis)
Parameter Guess FinalEst StdError 95%LCL 95%UCL
LogXSO 1.4000 1.4828 0.0309 1.4207 1.5449
s
YO
6.000 3.154
97,00 93.37
1.138
4,44
0.8/2 b.435
84.46 102.28
Effect Concentration Summary
% Effect
50,0
20.0
10.0
5.0
Xp Est
30.40
23.60
20.35
17.758
95%LCL
26.35
17,65
13.77
10.920
95%UCL
35.07
31.56
30,09
28,876
!>ic anponsz AnalvJU Model Virs.n 1 m
C-136 Draft Document
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C-137 Draft Document
-------
APPENDIX D: Other Data
D-l Draft Document
-------
Selenite
Additional data on the lethal and sublethal effects of selenium on aquatic species are presented in Table
D-l. Bringmann and Kuhn (1959a,b, 1976, 1977a, 1979, 1980b, 1981), Jakubczak et al. (1981), and
Patrick et al. (1975) reported the concentrations of selenite that caused incipient inhibition (defined
variously, such as the concentration resulting in a 3% reduction in growth) for algae, bacteria, and
protozoans (Table D-l). Although incipient inhibition might be statistically significant, its ecological
importance is unknown. Albertano and Pinto (1986) found the growth of three red algal species was
inhibited at selenite concentrations that ranged from 790 to 3,958 ug/L.
Selenate
Dunbar et al. (1983) exposed fed D. magna to selenate for seven days and obtained an LC50 of 1,870
ug/L. This value is in the range of the 48-hr EC50s in Table D-l.
Watenpaugh and Beitinger (1985a) found that fathead minnows did not avoid 11,200 ug/L selenate
during 30-minute exposures (Table D-l). These authors also reported (1985b) a 24-hr LC50 of 82,000
ug/L for the same species and they found (1985c) that the thermal tolerance of the species was reduced by
22,200 ug/L. Westerman and Birge (1978) exposed channel catfish embryos and newly hatched fry for
8.5 to 9 days to an unspecified concentration of selenate. Albinism was observed in 12.1 to 36.9% of the
fry during the five years of such exposures. Pyron and Beitinger (1989) also investigated fathead
minnows, and after a 24-hr exposure, no effect on reproductive behavior was found at 36,000 ug/L, but
when adults were exposed to 20,000 ug/L selenate for 24-hr, edema was observed for their larvae.
The respiratory rate of the eastern oyster, Crassostrea virginica, was unaffected by exposure to selenate at
400 ug/L for 14 days (Fowler et al. 1981). Embryos of the striped bass were quite tolerant to selenate in
dilute salt water (Klauda 1985a, b). There was a 93% successful hatch of embryos at 200,000 ug/L, but
50% of 72-day-old juveniles died after four days at 87,000 ug/L. Exposure of juvenile fish for up to 65
days to concentrations of selenate between 39 and 1,360 ug/L caused developmental anomalies and
pathological lesions.
D-2 Draft Document
-------
Table D-l. Other Data on Effects of Selenium on Aquatic Organisms
Species
Hardness
(mg/L as
Chemical CaCCX) Duration
Effect
Concentration*
Reference
FRESHWATER SPECIES
Selenium (IV)
Green alga,
Scenedesmus
quadricauda
Green alga,
Selenastrum
capricornutum
Green alga,
Selenastrum
capricornutum
Green alga,
Selenastrum
capricornutum
Alga,
Chrysochromulina
breviturrita
Red alga,
Cyanidium
caldarium
Red alga,
Cyanidioschyzon
merolae
Red alga,
Galdieria
sulphuraria
Algae (diatoms),
Mixed population
Bacterium,
Escherichia coli
Bacterium,
Pseudomonus putida
Protozoan,
Entosiphon sulcatum
Protozoan,
Microreqma
heterostoma
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Selenious
acid
Selenious
acid
Selenious
acid
Selenious
acid
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
96hr
72hr
72 hr
72 hr
30 days
20 days
20 days
20 days
18 days
-
16hr
72 hr
28 hr
Incipient
inhibition (river
water)
Decreased dry
weight and
chlorophyll a
BCF = 12-21b
BCF= 11,164C
Increased growth
Inhibited growth
Inhibited growth
Inhibited growth
Inhibited growth
Incipient
inhibition
Incipient
inhibition
Incipient
inhibition
Incipient
inhibition
2,500
75
10-100
150
320
3,958
3,140
790
11,000
90,000
11,400
(11,200)
1.8
(1.9)
183,000
Bringmann and
Kuhn 1959a,b
Foe and Knight,
Manuscript
Foe and Knight,
Manuscript
Foe and Knight,
Manuscript
Wehr and Brown
1985
Albertano and
Pinto 1986
Albertano and
Pinto 1986
Albertano and
Pinto 1986
Patrick et al.
1975
Bringmann and
Kuhn 1959a
Bringmann and
Kuhn 1976;
1977a; 1979;
1980b
Bringmann 1978;
Bringmann and
Kuhn 1979;
1980b; 1981
Bringmann and
Kuhn 1959b
Protozoan,
Sodium
! hr Incipient
62
Bringmann and
D-3 Draft Document
-------
Species
Chilomonas
paramecium
Protozoan,
Uronema parduezi
Snail,
Lymnaea stagnalis
Cladoceran,
Daphnia magna
Cladoceran,
Daphnia magna
Cladoceran,
Daphnia magna
Cladoceran,
Daphnia magna
Cladoceran (<24 hr),
Daphnia magna
Cladoceran
(5th instar),
Daphnia magna
Cladoceran,
Daphnia magna
Cladoceran
(preadult),
Daphnia pulex
Ostracod,
Cyclocypris sp.
Amphipod,
Hyalella azteca
Amphipod
(2 mm length),
Hyalella azteca
Amphipod
(2 mm length),
Hyalella azteca
Amphipod
(2 mm length),
Hyalella azteca
Midge (first instar),
Chironomus riparius
Chemical
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Selenious
acid
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Hardness
(mg/L as
CaCCK'l Duration
20 hr
7.5 days
48 hr
214 24 hr
214 24 hr
329 48 hr
96 hr
14 days
48 hr
21 days
48 hr
220d 48 hr
42 24 hr
100.8 48 hr
329 14 days
133 48 hr
133 10 days
133 24 days
134 48 h
Effect
inhibition
Incipient
inhibition
LT50
EC50 (river
water)
LC50
EC50
(swimming)
EC50 (fed)
EC50 (fed)
LC50 (fed)
LC50 (fed)
Did not reduce
oxygen
consumption or
filtering rate
LC50
LC50 (fed)
LC50
LC50
(fed)
LOEC
reproduction
(static-renewal)
LC50
Concentration8 Reference
Kuhnl981;
Bringmann et al.
1980
118
3,000
2,500
16,000
9.9
710
430
430
685
160
680
1,200
>498
70
623
312
200
7,950
Bringmann and
Kuhn 1980a;
1981
Van Puymbroeck
etal. 1982
Bringmann and
Kuhn 1959a,b
Bringmann and
Kuhn 1977a
Bringmann and
Kuhn 1977b
Halter etal. 1980
Adams and
Heidolphl985
Johnston 1987
Kimball,
Manuscript
Reading and
Buikemal980
130,000 Owsley 1984
Halter etal. 1980
Brasher and Ogle
1993
Brasher and Ogle
1993
Brasher and Ogle
1993
Ingersoll et al.
1990
D-4 Draft Document
-------
Hardness
(mg/L as
Species
Midge (first instar),
Chironomus riparius
Coho salmon (fry),
Oncorhynchus
kisutch
Rainbow trout (fry),
Oncorhynchus
mykiss
Rainbow trout (fry),
Oncorhynchus
mykiss
Rainbow trout,
Oncorhynchus
mykiss
Rainbow trout,
Oncorhynchus
mykiss
Rainbow trout,
Oncorhynchus
mykiss
Rainbow trout
(juvenile),
Oncorhynchus
mykiss
Rainbow trout
(juvenile),
Oncorhynchus
mykiss
Rainbow trout
(juvenile),
Oncorhynchus
mykiss
Rainbow trout
(juvenile),
Oncorhynchus
mykiss
Rainbow trout
(juvenile),
Oncorhynchus
mykiss
Rainbow trout,
Oncorhynchus
mykiss
Rainbow trout,
Oncorhynchus
mykiss
Chemical
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
CaCCK'l Duration
40-48 48 h
325 43 days
334 21 days
334 21 days
330 5 days
325 48 days
325 96 days
4wk
4wk
4wk
42 wk
42 wk
135 9 days
135 96 hr
9 days
Effect
LC50
LC50
LC50
Reduced growth
LC50
LC50
LC50
MATC
survival
MATC
survival
BCF = 23
MATC growth
(dietary only
exposure)
MATC survival
(dietary only
exposure)
LC50
LC50
(fed)
Concentration8
14,600
160
460
250
2,700
2,750
500
280
200
4.7
Hg/g dw
(whole-body)
100
>9.96
Hg Se/g dw
(food)
5.34
Hg Se/g dw
(food)
7,020
7,200
5,410
Reference
Ingersoll et al.
1990
Adams 1976
Adams 1976
Adams 1976
Adams 1976
Adams 1976
Adams 1976
Gissel-Nielsen
and Gissel-
Nielsen 1978
Gissel-Nielsen
and Gissel-
Nielsen 1978
Gissel-Nielsen
and Gissel-
Nielsen 1978
Goettl and
Davies 1978
Goettl and
Davies 1978
Hodson et al.
1980
Hodson et al.
1980
Rainbow trout,
Sodium
135
96 hr LC50
8,200
Hodson et al.
D-5 Draft Document
-------
Hardness
(mg/L as
Species
Oncorhynchus
mykiss
Rainbow trout,
Oncorhynchus
mykiss
Rainbow trout,
Oncorhynchus
mykiss
Rainbow trout
(fertilized egg),
Oncorhynchus
mykiss
Rainbow trout
(embryo),
Oncorhynchus
mykiss
Rainbow trout,
Oncorhynchus
mykiss
Rainbow trout
(sac fry),
Oncorhynchus
mykiss
Rainbow trout
(sac fry),
Oncorhynchus
mykiss
Rainbow trout
(egg )=
Oncorhynchus
mykiss
Rainbow trout
(embryo),
Oncorhynchus
mykiss
Rainbow trout
(sac-fry),
Oncorhynchus
mykiss
Rainbow trout
(swim-up fry)
Oncorhynchus
mykiss
Northern pike,
Esox lucius
Goldfish,
Carassius auratus
Chemical
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Selenium
dioxide
CaCO^) Duration
9 days
135 41 days
135 50 wk
135 44 wk
120 hr
90 days
272 90 days
272 90 days
96 hr
96 hr
96 hi
96 hr
10.2 76 hr
157 14 days
Effect
(fed)
LOAEC
(Reduced hatch
of eyed embryos)
Decreased iron in
blood and red
cell volume
BCF = 33.2
BCF = 21.1
Did not reduce
survival or time
to hatch
Chronic value for
survival
LC50
MATC
survival
BCF = 17.5
BCF = 3.5
BCF = 3.1
BCF = 3.0
BCF= 13.1
BCF =1.6
BCF = 80.3
BCF = 20.2
LC50
LC50
Concentration8
6,920
26
53
53
10,000
14
55.2e
31.48
0.4
45.6
0.4
45.6
0.4
45.6
0.4
45.6
11,100
6,300
Reference
1980
Hodson et al.
1980
Hodson et al.
1980
Hodson et al.
1980
Klaverkamp et al.
1983b
Mayer etal. 1986
Hunnetal. 1987
Hunnetal. 1987
Hodson et al.
1986
Hodson et al.
1986
Hodson et al.
1986
Hodson et al.
1986
Klaverkamp et al.
1983a
Cardwell et al.
1976a,b
D-6 Draft Document
-------
Species
Goldfish,
Carassius auratus
Goldfish,
Carassius auratus
Goldfish,
Carassius auratus
Goldfish,
Carassius auratus
Fathead minnow,
Pimephales promelas
Fathead minnow,
Pimephales promelas
Fathead minnow,
Pimephales promelas
Fathead minnow,
Pimephales promelas
Creek chub,
Semotilus
atromaculatus
Bluegill,
Lepomis
macrochirus
Bluegill,
Lepomis
macrochirus
Bluegill (juvenile),
Lepomis
macrochirus
Bluegill (juvenile),
Lepomis
macrochirus
Largemouth bass
(juvenile),
Micropterus
salmoides
Yellow perch,
Percaflavescens
African clawed frog,
Xenopus laevis
African clawed frog,
Xenopus laevis
Chemical
Sodium
selenite
Sodium
selenite
Selenium
dioxide
Selenium
dioxide
Selenium
dioxide
Sodium
selenite
Sodium
selenite
Selenious
acid
Selenium
dioxide
Sodium
selenite
Selenium
dioxide
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Hardness
(mg/L as
CaCO,)
-
-
-
_
157
329
329
220d
_
318
157
16
25
and
200
25
and
200
10.2
-
-
Duration
10 days
46 days
7 days
48 hr
9 days
96 hr
14 days
8 days
48 hr
48 days
14 days
323 days
120 days
120 days
10 days
7 days
1-7 days
Effect
Mortality
Gradual anore
and mortality
LC50
Conditional
avoidance
LC50
LC50
(fed)
LC50
(fed)
LC50
(fed)
Mortality
LC50
LC50
MATC larval
survival
(dietary only
exposure)
No mortality
No mortality
LC50
LC50
Cellular damaj
Concentration8
5,000
2,000
12,000
250
2,100
1,000
12,500
19.75
Hg Se/g dw
(food)
10
Reference
Ellis 1937; Ellis
etal. 1937
Ellis etal. 1937
Weir and Hine
1970
Weir and Hine
1970
Cardwell et al.
1976a,b
Halter etal. 1980
600 Halter etal. 1980
420
312,000 Kim etal. 1977
Kimball,
Manuscript
400 Adams 1976
Cardwell et al.
1976a,b
Woock et al.
1987
Lemly 1982
Lemly 1982
4,800 Klaverkamp et al.
1983a,b
1,520 Browne and
Dumont 1980
Dumont 1980
Selenium (VI)
D-7 Draft Document
-------
Hardness
(mg/L as
Species
Alga,
Chrysochromulina
breviturrita
Rotifer,
Brachionus
calyciflorus
Snail,
Lymnaea stagnalis
Cladoceran,
Daphnia magna
Cladoceran
(juvenile),
Daphnia magna
Cladoceran
(5th instar),
Daphnia magna
Cladoceran
(5th instar),
Daphnia magna
Amphipod
(2 mm length),
Hyalella azteca
Amphipod
(2 mm length),
Hyalella azteca
Amphipod
(2 mm length),
Hyalella azteca
Amphipod
(1-11 days old),
Hyalella azteca
Amphipod
(1-11 days old),
Hyalella azteca
Midge (first instar),
Chironomus riparius
Midge (first instar),
Chironomus riparius
Rainbow trout
(embryo, larva),
Oncorhynchus
mykiss
Goldfish
(embryo, larva),
Carrassius auratus
Chemical
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
CaCCK'l Duration
30 days
120 96 hr
6 days
129.5 7 days
48hr
48 hr
90 hr
133 48 hr
133 10 days
133 24 days
18 10 days
(S04=3.4)
124 10 days
(S04=32)
134 48 h
40-48 48 h
104 28 days
(92-110)
195 7 days
Effect
Increased
growth
EC20 Growth
(dry weight)
LT50
LC50
(fed)
LC50
(fed)
LC50
(fed)
42% of
organisms had
visible changes in
gut morphology
LC50
LC50
(fed)
LOEC
reproduction
(static renewal)
LC50
(fed)
LC50
(fed)
LC50
LC50
EC50 (death and
deformity)
EC50 (death and
deformity)
Concentration8
50
42.36
(Hg/g dw)
15,000
1,870
550
750
250
2378
627
>700
43
371
16,200
10,500
5,000
(4,180)
(5,170)
8,780
Reference
Wehr and Brown
1985
Dobbsetal. 1996
Van Puymbroeck
etal. 1982
Dunbar et al.
1983
Johnston 1987
Johnston 1987
Johnston 1989
Brasher and Ogle
1993
Brasher and Ogle
1993
Brasher and Ogle
1993
Borgmann et al.
2005
Borgmann et al.
2005
Ingersoll et al.
1990
Ingersoll et al.
1990
Birge 1978;
Birge and Black
1977; Birge etal.
1980
Birge 1978
D-8 Draft Document
-------
Species
Goldfish,
Carassius auratus
Fathead minnow,
Pimephales promelas
Fathead minnow,
Pimephales promelas
Fathead minnow,
Pimephales promelas
Fathead minnow,
Pimephales promelas
Fathead minnow,
Pimephales promelas
Fathead minnow,
Pimephales promelas
Fathead minnow,
Pimephales promelas
Fathead minnow,
Pimephales promelas
Channel catfish
(embryo, fry),
Ictalurus punctatus
Narrow-mouthed
toad
(embryo, larva),
Gastrophryne
carolinensis
Chemical
Sodium
selenate
Sodium
selenate
Sodium
selenate
-
-
-
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Hardness
(mg/L as
CaCCK'l Duration
24 hr
337.9 48 days
338 48 days
51 30min
24 hr
24 hr
44-49 7 days
160-180 24 hr
160-180 24 hr
90 8.5-9
days
195 7 days
Effect
BCF= 1.42
BCF=1.15
BCF= 1.47
BCF = 0.88
BCF= 1.54
LC50
LC50
No avoidance
LC50
Reduced thermal
tolerance
Chronic value -
growth
Chronic value-
growth
Chronic value-
survival
No effect on
reproductive
behavior
Edema in larvae
produced from
adults exposed to
Selenium VI
Induced albinism
EC50 (death and
deformity)
Concentration8
0.45
0.9
1.35
2.25
4.5
2,000
1,100
11,200
82,000
22,200
1,739
561
2,000
36,000
20,000
_
90
Reference
Sharma and
Davis 1980
Adams 1976
Adams 1976
Watenpaugh and
Beitinger 1985a
Watenpaugh and
Beitinger 1985b
Watenpaugh and
Beitinger 1985c
Norberg-King
1989
Pyron and
Beitinger 1989
Pyron and
Beitinger 1989
Westerman and
Birge 1978
Birge 1978;
Birge and Black
1977; Birge etal.
1979a
Organo-selenium
Bluegill (juvenile),
Lepomis
macrochirus
Bluegill (juvenile),
Lepomis
macrochirus
Seleno-L-
methionine
Seleno-L-
methionine
16 323 days
283 90 days
MATC larval
survival
(dietary only
exposure)
EC20 survival
(dietary only
exposure)
20.83
Hg Se/g dw
(food)
>13.4
Hg/g dw
(food)
Woock et al.
1987
Cleveland et al.
1993
D-9 Draft Document
-------
Species
Bluegill
(2 yr and adult),
Lepomis
macrochirus
Bluegill
(2 yr and adult),
Lepomis
macrochirus
Redear sunfish
(adult),
Lepomis microlophus
Phytoplankton,
Mixed population
Cladoceran
(<24 hr),
Daphnia magna
Cladoceran
(<24 hr),
Daphnia magna
Midge (<24-hr),
Chironomus riparius
Bluegill (juvenile),
Lepomis
macrochirus
Bluegill (juvenile),
Lepomis
macrochirus
Species
Anaerobic
bacterium,
Methanococcus
vannielli
Bacterium,
Vibriofisheri
Chemical
Selenium
Selenium
Selenium
Selenium
Selenite-
Selenate
mixture
Selenite-
Selenate
mixture
Selenite-
Selenate
mixture
Selenite-
Selenate
mixture
Selenite-
Selenate
mixture
Chemical
Sodium
selenite
Sodium
selenite
Hardness
(mg/L as
CaCO,) Duration Effect
field NOEC
deformities
field NOEC
deformities
field LOEC Adverse
histopathological
alterations
Selenium Mixtures
field Reduced growth
rates
138 21 days MATC
growth
138 21 days MATC
productivity
138 30 days MATC
emergence
283 60 days NOEC survival
283 60 days EC20 survival
Salinity
(g/kg) Duration Effect
SALTWATER SPECIES
Selenium (IV)
HOhr Stimulated growth
5 min 50% decrease in
light output
(Microtox> )
Concentration8
53.83
ug Se/g dw
(liver)
23.38
ug Se/g dw
(ovaries)
<38.15
ug Se/g dw
18
115.2
ugSe/L
21.59 ug/gdw
(whole-body)
503.6
340
4.07
ug/g dw
(whole body)
Reference
Reashetal. 1999
Reashetal. 1999
Sorensenl988
Riedeletal. 1991
Ingersoll et al.
1990
Ingersoll et al.
1990
Ingersoll et al.
1990
Cleveland et al.
1993
Cleveland et al.
1993
Concentration
{ugM
79.01
68,420
Reference
Jones and
Stadtman
1977
Yuetal. 199:
D-10 Draft Document
-------
Species
Green alga,
Chlorella sp.
Green alga,
Platymonas
subcordiformis
Green alga,
Dunaliella
primolecta
Diatom,
Skeletonema
costatum
Diatom,
Chaetoceros
muelleri
Diatom,
Phaeodactylum
tricornutum
Diatom,
Thallassiosira
aestivalis
Brown alga,
Fucus spiralis
Red alga,
Porphyridium
cruentum
Chemical
Sodium
selenite
Sodium
selenite
Sodium
selenite
Selenium
dioxide
Selenium
dioxide
Selenium
dioxide
Selenium
oxide
Sodium
selenite
Sodium
selenite
Salinity
(g/kg) Duration
32 14 days
32 14 days
32 20 days
5 days
6 days
8 days
29-30 72 hr
60 days
32 27 days
Effect
5- 12% increase in
growth
23% increase in
growth
Increased growth;
induced glutathione
peroxidase
BCF = 18,000
BCF = 16,000
BCF = 10,000
BCF = 337,000
BCF = 65,000
BCF = 5,000
BCF = 109,000
BCF = 27,000
BCF = 7,000
No effect on cell
morphology
1355% increase in
growth of thalli
Increase growth;
induced glutathione
peroxidase
Concentration
10-10,000
100-10,000
4,600
0.06
0.79
3.6
0.06
0.79
3.6
0.06
0.79
3.6
78.96
2.605
4,600
Reference
Wheeler et al.
1982
Wheeler et al.
1982
Gennity et al.
1985a,b
Zhang et al.
1990
Zhang et al.
1990
Zhang et al.
1990
Thomas et al.
1980a
Fries 1982
Gennity et al.
1985a,b
Selenium (VI)
Bacterium,
Vibrio fisheri
Green alga,
Chlorella sp.
Green alga,
Chlorella sp.
Green alga,
Dunaliella
primolecta
Green alga,
Dunaliella
primolecta
Green alga,
Dunaliella
primolecta
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
15 min
32 14 days
32 4-5 days
32 14 days
32 14 days
32 4-5 days
50% decrease in
light output
(Microtox> )
No effect on rate of
cell
100% mortality
No effect on rate of
cell population
growth
71% reduction in
rate of cell
population growth
100% mortality
3,129,288 Yuetal. 1997
10-1,000 Wheeler etal.
1982
10,000 Wheeler etal.
1982
10-100 Wheeler etal.
1982
1,000 Wheeler etal.
1982
10,000 Wheeler etal.
1982
D-ll Draft Document
-------
Species
Green alga,
Platymonas
subcordiformis
Green alga,
Platymonas
subcordiformis
Green alga,
Platymonas
subcordiformis
Green alga,
Platymonas
subcordiformis
Brown alga,
Fucus spiralis
Red alga,
Porphridium
cruentum
Red alga,
Porphyridium
cruentum
Eastern oyster
(adult),
Crassostrea
virginica
Striped bass
(embryo),
Morone saxatilis
Striped bass
(larva),
Morone saxatilis
Striped bass
(juvenile),
Morone saxatilis
Striped bass
(juvenile),
Morone saxatilis
Chemical
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Salinity
(g/kg) Duration Effect
32 14 days No effect on rate of
cell population
growth
32 14 days 16% decrease in
rate of cell
population growth
32 14 days 50% decrease in
rate of cell
population growth
32 4-5 days 100% mortality
60 days 160% increase in
growth rate of thalli
32 14 days 23-35% reduction
in rate of cell
population growth
32 4-5 days 100% mortality
34 14 days No significant
effect on respiration
rate of gill tissue
7.2-7.5 4 days 93% successful
hatch and survive
4.0-5.0 4 days LC50 (control
survival= 77%)
3.5-5.5 9-65 days Significant
incidence of
development
anomalies of lower
jaw
3.5-5.5 45 days Significant
incidence of severe
blood
cytopathology
Concentration
(ug/L)a
10
100
1,000
10,000
2.605
10-1,000
10,000
400
200,000
13,020
39-1,360
1,290
Reference
Wheeler et al.
1982
Wheeler et al.
1982
Wheeler et al.
1982
Wheeler et al.
1982
Fries 1982
Wheeler et al.
1982
Wheeler et al.
1982
Fowler et al.
1981
Klauda
1985a,b
Klauda
1985a,b
Klauda
1985a,b
Klauda
1985a,b
a Concentration of selenium, not the chemical. Units are Og selenium/L of water unless noted otherwise.
b Converted from dry weight to wet weight basis (see Guidelines)
0 Growth of algae was inhibited
d From Smith etal. (1976).
e Calculated from the published data using probit analysis and allowing for 8.9% spontaneous mortality.
D-12 Draft Document
-------
Other Data - Endangered Species
Two similar studies were conducted in 1996 and 1997 to determine effects of site water and site food,
both contaminated with selenium, on the endangered species, razorback sucker, Xyrauchen texanus
(Hamilton et al. 2001a,b; published later in a peer-reviewed journal in 2005, see Hamilton et al. 2005
a,b,c). Both studies show marked effects of selenium on survival of razorback sucker larvae exposed to
contaminated food and to a lesser extent, contaminated water. Although the data convincingly
demonstrate effects to larval survival from exposure to contaminated food, interpretation of the results in
the context of chronic criterion derivation is complex because of inconsistencies between: 1) levels of
selenium in the food and larvae relative to larval survival; 2) the time to larval mortality relative to
selenium in the diet and selenium in the larvae; and 3) levels of other inorganic contaminants in food and
water (possible organic contaminants were not measured). A summary of each of these two studies is
presented below.
Evaluation of Contaminant Impacts on Razorback Sucker held in Flooded Bottomland Sites Near Grand
Junction , Colorado - 1996 (Hamilton et al. 2001a; also Hamilton et al. 2005 a,b,c)
This study was initiated with 5-day old razorback sucker larvae spawned from adults (first time spawners)
which were previously held (9 months) in three different locations along the Colorado River that
contained varying levels of selenium: Horsethief (the designated reference site which receives water
pumped directly from the Colorado River near Fruita, CO, and where dissolved selenium concentrations
in water ranged from < 1.6 to 3.9 (ig/L during the period of exposure), Adobe Creek (low level selenium
contamination - dissolved selenium concentrations in water ranged from 1.5 to 11.6 (ig/L; avg. = 3.8
(ig/L), and North Pond (high level selenium contamination - dissolved selenium concentrations in water
ranged from 3.8 to 19.6 (ig/L; avg. = 9.5 (ig/L). The selenium content in eggs from three Horsethief
females ranged from 5.8 to 6.6 mg Se/kg dw, and the selenium content in adult muscle plugs at spawning
was from 3.4 to 5.0 mg Se/kg dw. The selenium content in the eggs from three Adobe Creek females
ranged from 38.0 to 54.5 mg Se/kg dw, and the selenium content in adult muscle plugs at spawning was
from 11.5 to 12.9 mg Se/kg dw. The selenium content in the eggs from three North Pond females ranged
from 34.3 to 37.2 mg Se/kg dw, and the selenium content in adult muscle plugs at spawning was from
14.1 to 17.3 mg Se/kg dw. The selenium content in eggs from one of three hatchery brood stock females
was 7.1 mg Se/kg dw, and the selenium content in muscle plugs of two of three hatchery brood stock
females at spawning ranged from 2.6 to 13.8 mg Se/kg dw. The razorback sucker larvae spawned from
fish hatchery brood stock (older, previously spawned females) and held in Colorado River (Horsethief)
water were used as an additional reference group of test fish.
D-13 Draft Document
-------
The experimental groups were subdivided into those receiving reference water (hatchery water; 24-Road
Fish Hatchery) or site water (Table D-2). They were further subdivided into those receiving a daily ration
of reference food (brine shrimp) or zooplankton (predominantly cladocerans and copepods) collected
from each site where their parents were exposed for the previous 9 months. A total of 60 larvae from each
of the four adult sources (Horsethief, Adobe Creek, North Pond, Brood Stock held in different ponds at
Horsethief) were exposed to each treatment (2 replicates x 3 spawns x 10 fish/beaker). The larvae were
held in beakers containing 800 ml of test water. Fifty percent of the test water was renewed daily.
Table D-2. Treatment conditions during the 30-day larval study
Source of Larvae
Horsethief Adults
Adobe Creek Adults
North Pond Adults
Hatchery raised Adults
Treatments
Reference food: Reference
water
Reference food: Site water
Site food: Reference water
Site food: Site water
Reference food: Reference
water
Reference food: Site water
Site food: Reference water
Site food: Site water
Reference food: Reference
water
Reference food: Site water
Site food: Reference water
Site food: Site water
Reference food: Reference
water
Reference food: Site water
Site food: Reference water
Site food: Site water
Sein
food
(mg/kg dw)
2.7
2.7
5.6
5.6
2.7
2.7
20
20
2.7
2.7
39
39
2.7
2.7
5.6
5.6
Dissolved Se in
water
(Og/L)
<1.6
0.9
<1.6
0.9
<1.6
5.5
<1.6
5.5
<1.6
10.7
<1.6
10.7
<1.6
0.9
<1.6
0.9
D-14 Draft Document
-------
Growth, survival and development were evaluated amongst treatment groups for up to 30 days in the
treatment conditions. Each treatment group was fed once daily after renewal. Test waters were collected
every day from each site as grab samples for the renewal. A small portion of this water was retained at 3-
and 7-day intervals for an analysis of total and dissolved selenium concentrations. At approximately 2-
day intervals, aquatic invertebrates and brine shrimp not used for feeding were sieved from the media for
selenium analysis. The number of live fish was recorded daily. After the 30-day exposure period, the
surviving fish were sacrificed and measured for total length. At this same time, approximately four fish
from each treatment, when available, were collected as a composite sample and analyzed for total
selenium.
After 30 days of exposure in the reference food-reference water treatment, survival of razorback sucker
larvae from brood stock and Horsethief adults (89 and 87 percent, respectively) was slightly higher than
those from Adobe Creek adults (84 percent) and North Pond adults (75 percent). Corresponding selenium
concentrations in larval whole-body tissue were 3.6, 3.3, 7.7 and 9.7 mg Se/kg dw, respectively. Survival
was similar or slightly reduced in larvae from all four sources after 30 days of exposure in the reference
food-site water treatments; corresponding selenium concentrations in larval whole-body tissue were 5.2,
5.1, 12.7 and 15.2 mg Se/kg dw, respectively. In contrast, none of the larvae spawned from parents from
Horsethief, Adobe Creek, or North Pond survived to 30 days when fed zooplankton collected from the
three sites, irrespective of the water type they were exposed to (i.e., reference or site). Only the larvae
from brood stock adults, which were fed zooplankton from the Horsethief site for this treatment, survived,
and even these larvae suffered substantial mortality (40 and 60 percent respectively). The mean selenium
concentrations in whole-body tissue of larvae from brood stock adults after the 30-day exposures were 5.4
mg Se/kg dw (site food-reference water treatment) and 6.9 mg Se/kg dw (site food-site water treatment).
Several inconsistencies were observed that indicate selenium may not be solely responsible for the effect
on larval survival. Larval survival in the Adobe Creek treatment group exposed to reference water (<1.6
(ig/L) and reference food (2.7 mg Se/kg dw ) was 84 percent, similar to survival of larvae from brood
stock (89 percent). The selenium concentration in the larvae from this Adobe Creek treatment group after
30 days was higher (7.7 mg/kg dw) than that of the brood stock fish (5.4 mg Se/kg dw) in the reference
water (<1.6 (ig/L) and site food (5.6 mg Se/kg dw) treatment, which had a 30-day survival of 62 percent.
Also, the time to 50 percent mortality between the site food treatments, where most mortality occurred,
was not related to selenium concentration in the diet or in the larvae.
D-15 Draft Document
-------
Although the larvae from brood stock held at Horsethief and the larvae from the first-time spawning
adults held at Horsethief that were used for the 9 month exposure received the same site food, no larvae
from the latter group survived the 30 day exposure. Concentrations of selenium in the larvae of these two
treatment groups were essentially the same between days 6 and 12 of the exposure (8.1 to 8.9 mg Se/kg
dw). During this same general time frame (6 to 7 days of exposure), larvae from Adobe Creek and North
Pond adults apparently tolerated up to 32 and 39 mg Se/kg dw in tissue, respectively, without any
increase in mortality when exposed to reference food and reference water. Larvae grown out under
hatchery conditions from adults in the Horsethief and Adobe Creek treatments also did not differ in total
deformities compared to larvae from brood stock. There was also no difference between treatments
(brood stock, Horsethief, Adobe Creek, and North pond) in percent egg viability, percent hatchability,
percent embryos with deformities, and percent mortality of deformed embryos and larvae from a separate
test initiated with eggs in the same study (Hamilton et al. 2005b).
Evaluation of Contaminant Impacts on Razorback Sucker held in Flooded Bottomland Sites Near Grand
Junction , Colorado - 1997 (Hamilton et al. 200lb)
In a similar 30-day larval study conducted by the authors in the following year (1997), razorback sucker
larvae from a single hatchery brood stock female (11 mg Se/kg dw muscle) were subjected to the sixteen
different combined water and dietary exposure conditions described in the earlier (1996) study. The
female parent was held at Horsethief Canyon State Wildlife Area before spawning. The larvae were held
in beakers containing 800 ml of test water as before; fifty percent of the test water was renewed daily.
Specific treatment conditions for the 1997 30-day larval study are listed in Table D-3.
Table D-3. Treatment conditions during the 30-day larval study
Water Treatments
Reference food (brine shrimp):
Reference water (24-Road Hatchery)
Reference food: Site water (Horsethief)
Reference food: Site water (Adobe Creek)
Reference food: Site water (North Pond)
Horsethief food: Reference water
Horsethief food: Site water (Horsethief)
Sein
food
(mg/kg dw)
3.2
6.0
32.4
52.5
3.2
6.0
Se in
water
(Og/L)
< 1
1.6
3.4
13.3
< 1
1.6
D-16 Draft Document
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Water Treatments
Horsethief food: Site water (Adobe Creek)
Horsethief food: Site water (North Pond)
Adobe Creek food: Reference water
Adobe Creek food: Site water (Horsethief)
Adobe Creek food: Site water (Adobe Creek)
Adobe Creek food: Site water (North Pond)
North Pond food: Reference water
North Pond food: Site water (Horsethief)
North Pond food: Site water (Adobe Creek)
North Pond food: Site water (North Pond)
Sein
food
(mg/kg dw)
32.4
52.5
3.2
6.0
32.4
52.5
3.2
6.0
32.4
52.5
Sein
water
(Og/L)
3.4
13.3
< 1
1.6
3.4
13.3
< 1
1.6
3.4
13.3
After 30 days of exposure in this yearns study, there was also good survival of razorback sucker larvae
fed reference food (brine shrimp) and held in reference water or water from Horsethief (83 and 81
percent, respectively). The survival of these larvae was significantly greater than survival of larvae fed
brine shrimp and held in water from North Pond (52 percent). Corresponding selenium concentrations in
larval whole-body tissue after 10 days were 6.3, 6.7, and 11 mg Se/kg dw, respectively. The average
concentrations of selenium in the water for the three treatments were <1, 1.6, and 13.3 (ig Se/L. After 30
days the mean selenium concentrations in these larvae were 5.2, 5.2, and 16 mg Se/kg dw, respectively.
Survival was markedly reduced (0 to 30 percent survival) in the remaining treatments where larvae were
fed zooplankton from the various sites. Complete mortality was experienced by larvae exposed to
Horsethief food and reference water treatment after 30 days.
Similar to the previous study, several inconsistencies in results suggested that selenium may not have
been solely responsible for the effect on larval survival. The most notable inconsistency was that the
greatest effect on larval survival (percent survival or time to 50 percent mortality) was from exposure to
Horsethief food, the food with the lowest selenium contamination.
The authors of the above two studies (Hamilton et al. 2001a,b) make a strong argument that some of the
inconsistency in response observed in their studies between larvae fed reference and site diets may be
D-17 Draft Document
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related to the difference in arsenic concentration between the two diets. The arsenic concentration
measured in the brine shrimp used in the reference diet was 24 mg total As/kg dw (measured in the
second larval study) versus between 6 and 7.5 mg total As/kg dw measured in the zooplankton from the
various sites. In their publication (Hamilton et al. 2005c), the authors cite several studies reporting an
ameliorating effect of arsenic against the toxicity of a variety of forms of selenium in various animals
(Dubois et al. 1940, Hoffman et al. 1992, Klug et al. 1949, Levander 1977, Moxon 1938, Thapar et al.
1969). In terms of the survival of larvae from Horsethief, Adobe Creek and North Pond adults when fed
the reference diet, the authors propose that the arsenic concentrations in the brine shrimp diet may have
resulted in an antagonistic interaction with selenium and reduced adverse effects in larvae. Such
hypothesis is questionable, because their studies included diets spiked with inorganic arsenic salts,
whereas the arsenic in brine shrimp (and other natural diets), is most likely predominantly organic arsenic
(US EPA 2003). Additionally, in a separate but related study by the same authors (Hamilton et al. 2005d),
larval razorback sucker spawned from one female at the Ouray Native Fish Facility were fed zooplankton
from six sites (SI, S3, S4, S5, SR, and NR) adjacent to the Green River, Utah at four different initial ages
(5, 10, 24, and 28 day old larvae) for 20 to 25 days. The selenium concentrations in zooplankton from the
SI reference site ranged from 2.3 to 3.5 mg Se/kg dw (dissolved Se in water <0.6 to <1.1 (ig/L). The
concentrations in zooplankton from sites S3 and S4 were slightly higher (range 2.4 to 6.7 mg Se/kg dw;
water, 0.3-0.8 (ig/L), substantially elevated at S5 (12- 26 mg Se/kg dw; water, 0.6-3.1 (ig/L), and highest
at SR and NR (44-94 mg Se/kg dw; water, 14-107 (ig/L). All larvae in the test initiated when they were 5
days old (study 1) died after 25 days of exposure. Median time to death was shortest in fish fed
zooplankton from the reference site (SI) and longest for SR and NR. Interestingly, the concentration of
arsenic measured in zooplankton collected from SI was 12 mg As/kg dw, half that of the brine shrimp
used in the above study (Hamilton et al. 200 Ib), which did not appear to antagonize the toxicity of the
selenium in the diet in this test. In this and the previous two studies, additional inorganic contaminants
such as vanadium and strontium were elevated in the zooplankton fed to the larval razorback sucker.
Other Data - Chronic Studies with Fish Species
Some chronic studies met the requirements of an acceptable chronic test but were excluded from Table 1
for a variety of reasons. Summaries of these studies are provided below.
D-l 8 Draft Document
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Vidal, D., S.M. Bay and D. Schlenk. 2005. Effects of dietary selenomethionine on larval rainbow trout
(Oncorhynchus mykiss). Arch. Environ. Contamin. Toxicol.49:71-75.
Test Organism:
Exposure Route:
Test Duration:
Study Design:
Effects Data:
Chronic Value:
Rainbow trout (Oncorhynchus mykiss)
Dietary only
Selenomethionine was added to dry fish food; the measured dietary
concentrations were 4.6, 12 and 18 (ig Se/g dw. The measured selenium in the
control diet was 0.23 (ig Se/g dw.
90 days
Each of the three dietary treatments and control had 5 replicates, each replicate
contained 12 to 16 larval rainbow trout that were 27 days old at initiation. Each
fish was fed an average of 10 mg/d for 30 days; 25 mg/d on days 30-60; and 40
mg/d thereafter. Fish were sampled on days 30, 60 and 90 for length, weight,
selenium, hepatic GSH and thiobarbituric acid-reactive substances (TEARS)
measurements.
The authors reported significant decreases in weight and length after the 90-day
exposure (Table D-4). There were no significant differences in the hepatic lipid
peroxidation and hepatic GSH to GSSH ratios among the treatments. The authors
found significant differences in weight and length in the 4.6 and 12 (ig Se/g dw
dietary treatments, but not the 18 (ig Se/g dw treatment. Based on larval trout
body burden, the authors reported an LOEC of 1.20 (ig/g ww, the concentration
of Se in fish fed the 12 (ig Se/g dw dietary treatment. The Se concentration in
larval rainbow trout associated with the lowest dietary treatment that showed
significant decreases in larval weight and length was 0.58 (ig Se/g ww or 2.06 (ig
Se/g dw based on 71.8% moisture in whole body rainbow trout (NCBP).
The data from this study was not used to calculate a chronic value for selenium
due to several inconsistencies. The significant decreases in length and weight
observed in the two lowest concentrations were not observed in the highest
dietary treatment. The Se concentrations in the larval rainbow trout were
irregular with the 60-day concentrations being considerably higher than the 90-
day concentrations. The authors explain this observation to rapid growth in the
fish causing dilution of the Se body burden. However, the increase in fish weight
from 30 to 60 days was similar to the 60 to 90 day increase and the 60 day Se
concentrations increased from day 30. Also, the Se concentration in the control
fish went from below detection on day 0 to 0.46 (ig/g ww on day 30; to 1.24 (ig/g
ww on day 60; and to 0.31 (ig/g ww on day 90. The 60-day measured Se in the
control fish (1.24 (ig/g ww) was more than twice the concentration of Se in the
fish with lowest concentration showing effects (0.58 (ig/g ww).
D-19 Draft Document
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Table D-4. Mean (SD) rainbow trout growth after four SeMet dietary treatments
test day
0
30
60
90
Treatment,
Mg/g dw
control
control
4.6
12
18
control
4.6
12
18
control
4.6
12
18
weight, g
0.37(0.30)
1.33(0.92)
1.25(0.21)
1.33(0.30)
1.31(0.37)
2.96 (0.92)
2.33 (0.63)
2.52(0.38)
2.59 (0.24)
5.17(1.09)
3.45 (0.35)*
3.45 (0.35)*
3.82(0.62)
fork length,
cm
3.14(0.41)
4.66(0.41)
4.84 (0.29)
5.09(0.46)
4.97 (0.50)
6.91 (0.56)
6.69 (0.67)
6.88(0.35)
6.92 (0.24)
7.70(0.33)
6.93(0.19)*
6.84(0.68)*
7.37 (0.62)
[Se] whole body,
(lg/g WW
ND
0.46 (0.20)
1.05(0.77)
1.81(1.04)
1.60(0.93)
1.24(0.54)
1.70(0.72)
1.83(0.94)
2.62(1.22)
0.31(0.20)
0.58(0.21)
1.20(0.21)*
1.41 (0.27)*
[Se] whole body,
Hg/g dw**
ND
1.63
3.72
6.42
5.67
4.40
6.03
6.49
9.29
1.09
2.06
4.25
5.00
* Significantly different than the control.
** ww converted to dw using 71.8% moisture for whole body rainbow trout (NCBP).
Deng, X. 2005. Early life stages of Sacramento splittail (Pogonichthys macrolepidotus) and selenium
toxicity to splittail embryos, juveniles and adults. Doctoral dissertation, University of California, Davis.
Test Organism:
Exposure Route:
Test duration:
Study Design:
Sacramento splittail (Pogonichthys macrolepidotus)
Dietary only
Four concentrations of selenium in the fish diet (0.6, 17.3, 33.0, and 70.1 mg/g)
were created by mixing different proportions of selenized and Torula yeast. A
different batch of selenized yeast was used in the adult exposure.
24 weeks
Fourteen adult fishes were placed in each circular tank (92 cm diameter, 33 cm
height) and fed one of the four diets. Each diet was provided to fishes in three
tanks. The twelve tanks were arranged in three rows. Each row had all four
treatment concentrations with randomly assigned positions. Thus, the experiment
had a randomized block design. Adult splittail fishes were obtained from the
Tracy Pump Station (U.S. Bureau of Reclamation, Tracy, CA). After 12 and 24
weeks of exposure, blood samples were collected, the liver, gonad, kidney and
D-20 Draft Document
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Effects Data:
white muscle were dissected, and liver and gonad were weighed to calculate
hepatosomatic and gonadosomatic indices. Stages of ovarian and testicular
development were determined from histological studies.
No mortality occurred throughout the experiment. Fish in control, 17.3, and 33.0
mg/g treatments exhibited normal behavior. Fish exposed to 70.1 mg/g in did not
consume as much food as fishes exposed to lower selenium concentrations, and
displayed abnormal behaviors. Splittail adults were less sensitive to dietary
selenium than juveniles. Relative to control, no changes in body weight, total
length, GSI, and condition factor were observed in fishes exposed to selenium
concentrations in food up to 33 mg/g. In general, tissue concentrations in fishes
exposed to selenium were higher than in the control, but differences in selenium
concentrations among them were often small and not significant (Table D-6).
Percentages of ovaries with atretic follicles increased with higher concentrations
of selenium in their diet: 30% in control, 45.5% in the 17.3 mg Se/g, and 100% in
the 33.0, and 70.1 mg/g treatments. The average concentration of selenium in
ovaries offish exposed to 17.3 mg/g in their diet was 6.5 mg/g. This low effect
level, though, is disputable because of the very low number of ovaries analyzed,
the occurrence of atresia in 30% of ovaries in control, and the lack of significant
differences in concentrations of selenium in ovaries among treatments exposed to
elevated levels of this element.
Table D-6. Mean concentration of selenium in ovaries (SE)J
[Se] in ovary (mg/g dw)
Diet Concentration (mg Se/g)
0.6
4.4
(0.57)
17.3
6.5
(1.0)
33.0
8.3
(0.14)
70.1
8.9
(0.46)
' Values estimated from Figure 4 in Deng (2005) (pg. Ill)
de Rosemond, K. Liber and A. Rosaasen. 2005. Relationship between embryo selenium concentration
and early life stage development in white sucker. Bull. Environ. Contamin. Toxicol. 74: 1134-1142.
Test Organism:
Exposure Route:
Test duration:
Study Design:
White Sucker (Catostomus commersoni)
Field collected.
In June, 2002, eggs were collected from 4 females from Island Lake (exposed
site); milt was obtained from 2 males. Island Lake is downstream from Cluff
Lake uranium mine located in northern Saskatchewan. Selenium concentrations
in Island lake range from 1 to 11 (ig/L and in recent years have been typically 4-5
(ig/L. No fish/eggs were collected from a reference site.
Through the end of yolk absorption by the larvae; 33 days post-fertilization.
Individual batches of eggs were fertilized in the field with milt and water-
hardened. Eggs were air transported to the laboratory in Saskatoon for testing.
D-21 Draft Document
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Effects Data:
Effect
Concentration:
200 eggs were randomly selected from each clutch and then separated into
groups of 100 which were placed into individual test chambers (n = 8).
On test day 30 (3 days prior to test termination), all fish larvae that exhibited
macroscopic deformities (e.g., kyphosis, lordosis, scoliosis and edema) were
removed, photographed and preserved. At test termination, (day 33), 40 larvae
from each female whites sucker were evaluated for deformities using a
microscope.
Although all four females were collected from the exposed site, selenium
concentrations in eggs were grouped into two low (Fish 2 and 3 in Table D-7)
and two high (Fish 1 and 4 in Table D-7). Larval mortality and developmental
deformities were not related to selenium concentrations in eggs (Table D-7). The
data suggest that embryo/larval effects are not observed at concentrations in eggs
reaching 40.3 mg/kg dw (geometric mean of the two high selenium
concentrations in eggs). However, because a reference condition with low
selenium exposure was not established, it is not appropriate to estimate an effect
concentration for this study. Note: the average percent moisture for the four
clutches of eggs was 92.6%.
NA
Table D-7. Embryo/larval endpoints for eggs from four female white sucker collected from Island
Lake in June 2002.
Measurement
Successfully hatched larvae3
Deformed larvaeb
Dead larvae0
Macroscopic deformities , %
Embryologicald
Developmental6
Microscopic deformities, %
Developmental
Total developmental deformities, %8
[Se] eggs mg/kg wwh
[Se] eggs mg/kg dwh
Fishl
161
21
6
6.8
6.2
7.5
13.7
2.7
33.6
Fish 2
140
25
14
6.4
11.4
5
16.4
0.7
9.4
Fish 3
176
16
6
5.7
3.4
2.5
5.9
0.6
8.4
Fish 4
141
13
4
1.4
7.8
7.5
15.3
3.2
48.3
a Initial number was 200 per fish
b Total number of deformed larvae throughout study; includes embryological and macroscopic
deformities
0 Total number of larvae that died throughout study.
d Percent of curled deformities that appeared in embryonic fish; deformities were evident immediately
after embryos hatched.
e Percent of deformities that were designated developmental; deformities became evident as larvae grew
and absorbed yolk sac (after experimental day 15).
D-22 Draft Document
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f Percent of microscopic developmental deformities that were evident in the 40 fish examined per
female white sucker.
8 The estimated percentage of offspring that had microscopic and macroscopic developmental
deformities combined.
h Selenium concentration measured in a subsample of embryos collected on test day 0.
D-23 Draft Document
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Ogle, R.S. and A.W. Knight. 1989. Effects of elevated foodborne selenium on growth and reproduction
of the fathead minnow (Pimephales promelas). Arch. Environ. Contam. Toxicol. 18:795-803.
Test Organism:
Exposure Route:
Test Treatments:
Test Duration:
Study Design:
Effects Data:
Fathead minnows (Pimephales promelas; juvenile, 59 to 61 d old)
Dietary only
Purified diet mix spiked with inorganic and organic selenium: 25 percent
selenate, 50 percent selenite, and 25 percent seleno-L-methionine, homogenized
in dextrin.
Completely randomized block design (2 blocks); 4 replicates per block (n = 8
replicates total per treatment). Actual mean total selenium levels in each
exposure treatment were: 0.4 (control), 5.2, 10.2, 15.2, 20.3, and 29.5 mg/kg dw.
Fish used in the first randomized block (F2 generation fish) were progeny from FI
generation originally obtained from the Columbia National Fishery Research
Laboratory, some of which were used in an initial range-finding experiment. Fish
obtained from a commercial supplier were used in the second randomized block.
The prepared diet was extruded into 1.5 mm pellets which were air-blown dried
to 5 percent moisture content and crushed and sieved so that only particles
retained by an 11.8 mesh/cm sieve were used in the study. The amount of
selenium in water that leached from the food during the experiment averaged
only 0.8
105 days, F2 generation (block one) and commercial fish (block two);
14 days F3 generation
Ten fish were randomly placed in each cell per block (n = 8x10, or 80 fish total
per treatment). Fish were fed twice daily at 6 percent body weight per day, with
wastes and uneaten food removed 30 min. after each feeding. Test tanks were
flushed with two tank volumes of fresh test water after each feeding (solution
renewal). Growth (as wet weight) was determined every two weeks by bulk
weighing, and one fish from two of the cells per treatment in a given block (n = 4
total per treatment) was removed for selenium (whole-body) analysis. After 105
days of exposure, a single male and female fish from each treatment replicate (n
= 4 breeding pairs per treatment in a given block, or 8 breeding pairs per
treatment total) were placed in 250 ml beakers and inspected for spawning
activity for 30 days following the first spawning event for that pair (each pair
being one replicate). Gonads and muscle tissue were dissected for selenium
analysis from these fish at the end of the 30 days spawning period. The spawning
substrates were inspected daily for eggs to determine fertility and viability.
Samples of not more than 50 eggs from each spawn were incubated in flowing,
aerated water and inspected for percent hatch determination. Ten larvae from
each incubated brood were transferred to separate glass test chambers and
maintained (48 h renewal; fed brine shrimp twice daily) for 14 days to determine
percent larval survival.
There was no effect of selenium on any of the reproductive parameters measured
at the dietary concentrations tested. Percent hatch and percent larval survival
were very high (>87.4 percent) and essentially equal for all of the treatments.
Growth of pre -spawning adults was affected by the selenium exposure (Table D-
5).
D-24 Draft Document
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Table D-5. Effects on Fathead Minnow Growth after 98 days of Exposure to Dietary Selenium
Measured mean selenium in
diet, mg/kg dw
0.4
5.2
10.2
15.2
20.3
29.5
Whole-body selenium,
mg/kg dw
1.76
2.78
3.42
5.40
6.58
7.46
Mean fish weight,
g ww
1.30
1.24
1.20
1.21
1.09
0.94
Chronic Value:
An EC value could not be calculated for these data because the data did not meet
the minimum requirements for analysis.
D-25 Draft Document
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GEI Consultants. 2008. Maternal Transfer of Selenium in Fathead Minnows, with Modeling of Ovary
Tissue to Whole Body Concentrations.
Test Organism:
Exposure Route:
Test duration:
Study Design:
Effects Data:
Fathead Minnow (Pimephales promelas)
Field collected.
Gravid adult fathead minnows were collected from creeks with a wide range of
surface water selenium concentrations near the city of Denver, CO during the
2006 summer breeding season.
Sites
Low selenium exposure:
• Sand Creek at Colfax. In 2002, aqueous selenium averaged 0.9 (ig/L.
Moderate to high selenium exposure:
• Sand Creek downstream of refinery
• East Tollgate Creek
• Mainstem Tollgate Creek
Control fish - no field exposure
• Laboratory-reared fish from Aquatic BioSystems
Embryo-larval test was 48 hours post hatch.
Field collected adult fish were either field dissected for selenium measurement in
paired tissues or transported live back to the laboratory in coolers with site water.
Fish were transported to the laboratory where mating pairs were bred in
individual chambers containing spawning substrates. Eggs were removed from
the spawning substrate and reared in a standard Falcon dish with lab water. Eggs
were screened under a dissecting microscope for viability. Dead eggs were
removed and numbers recorded on a datasheet. Three separate breeding
experiments were conducted.
Upon hatching, larvae were moved to standard bioassay cups containing lab
water and maintained in the laboratory incubator at 25°C. Larvae were
maintained via static conditions in exposure cups for 48 hours post-hatch without
food to ensure full absorption of the yolk sac before they were fixed in formalin.
Deformity assessment was performed on fixed embryos using a dissection
microscope. Test endpoints consisted of egg production, fertilization success,
mortality, and deformities (includes edema and skeletal, craniofacial and finfold
malformations). The authors used a graduated severity index (GSI) for
deformities in which larvae were scored 0 (normal), 1 (slight), 2 (moderate), and
3 (severe) based on the level of defect.
All fish successfully spawned except those collected from Sand Creek
downstream from the refinery. These fish had visible parasites and were only
used in the ovary-to-whole body selenium analysis. A suite of metal and
metalloids were measured in fish samples from each location. Fish collected from
East Tollgate Creek had higher concentrations of 9 of the 15 metals that were
D-26 Draft Document
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measured in fish from at least one site. Aluminum and iron showed the highest
difference with an approximate 10-fold increase in the East Tollgate Creek fish.
Only the first brood of each mating pair was used for the analysis because effects
appeared to be muted in subsequent broods. The lower response in the second
brood was thought to be due to clearing of selenium in the oocytes. There was
poor correlation between egg fertilization (R2 = 0.13) and embryo mortality (R2 =
0.18) data with whole body selenium concentrations in the adult fish (see Table
D-9 for summary data; see Table D-10 for individual brood data). Neither the
fraction of embryos surviving nor fertilization rate as a function of the
concentration of selenium in maternal fathead minnows was suitable for
estimating EC values. Although there were low survival and fertilization rates at
some higher selenium concentrations, these responses were quite varied and did
not follow a defined concentration-response relationship (Figure D-l).
Of the 9 broods from fish collected at the three exposed sites only one brood
(from East Tollgate Creek) had deformities greater than 10%. The fathead
minnow females that produced the brood with the greatest number of deformities
and highest GSI also had the second highest concentration of whole body
selenium, 46.4 mg/kg dw (Table D-l 1; Figures D-2 and D-3). Approximately
half of the larvae from this brood exhibited some sort of malformation. Similar to
the embryo parameters, EC values were not able to be estimated for any of the 4
malformation parameters.
The authors used probit analysis and TRAP to determine effect levels for each of
the embryonic and larval endpoints (Table D-l2). Although there is an indication
of effect due to selenium exposure in both the embryonic and larval endpoints,
there is too much variation in the responses observed with the embryos and
insufficient response observed with the larvae to derive a reasonable estimate of
effect levels. Therefore, no effect level was determined for this study.
Effect
Concentration: Unable to determine due to high variability or insufficient response.
D-27 Draft Document
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Table D-9. Mean fathead minnow first brood embryo and larval parameters and adult whole-body
(WB) selenium concentrations (dw) for each site (± 1SE); CON = control, SCC = Sand Creek at
Colfax Avenue bridge, TGC = Tollgate Creek, and ETC = East Tollgate Creek.
Parameter
n (number of breeding pairs)
WB Se concentration (mg/kg dw)
Egg fertilization (%)
Embryo mortality (%)
Mean spawn size (# of eggs per spawn)
Total larva evaluated (total # of broods)
Mean brood GSI score
Larval craniofacial defects (%)
Larval skeletal defects (%)
Larval finfold defects (%)
Larval edema (%)
Larval length (mm)
Site
Con
10
2.86±0.18
84.75 ±3.32
22.03 ±3.34
129 ±23
957
4.85 ± 1.22
2.64 ±0.90
4.74 ±0.89
2.19 ±0.78
3.89± 1.01
4.90 ±0.05
SCC
3
9.17 ±0.46
23.99 ±22.45
89.04 ±9.70
318±63
89
8. 88 ±8.88
4. 65 ±4.65
9.30 ±9.30
4.07 ±4.07
5.23 ±5.23
4.97 ±0.12
TGC
3
35.87 ±3.73
63 .42 ±31. 82
46.40 ±26.86
162 ±61
281
14.88 ±4.63
6.26 ±3.63
6.21 ±1.48
5.71 ±3.08
6.26 ±3.63
4.83 ±0.14
ETC
4
44.53 ±2.41
59.6 ±22.26
50.76 ±23.63
317 ± 158
254
21.75 ±9.53
18.48 ± 13.84
19.62 ± 12.11
17.23 ± 14.48
20.32 ± 12.93
4.90 ±0.07
Table D-10. Fathead minnow first brood embryo parameters and adult whole-body (WB)
selenium concentrations (dw) for each site (± 1SE); for site acronyms see Table D-9
Total eggs (total
Maternal WB dead+total Survival
Se Cone dw hatch+not fraction (total
Brood Code Treatment (mg/kg) hatched) dead/total eggs)
Fert. Rate ((Initial Egg
Count - 1st day
mortalities)/Initial Egg
Count)
T-la-1
T-lf-1
T-lf-1
T-2a-l
T-3a-l
T-3b-l
T-3d-l
T-4d-l
T-5d-l
T-6d-l
T-2b-l
T-4a-l
T-6a-l
T-2a-l
T-3a-l
T-4a-l
CON
CON
CON
CON
CON
CON
CON
CON
CON
CON
SCC
SCC
SCC
TGC
TGC
TGC
2.90
3.24
1.94
2.25
2.71
2.64
3.67
3.43
3.33
2.52
9.92
8.35
9.25
32.29
43.33
31.99
19
238
19
135
154
90
76
199
149
183
395
193
340
132
79
262
0.79
0.77
0.63
0.98
0.68
0.90
0.70
0.85
0.73
0.76
0.00
0.03
0.30
0.83
0.00
0.77
0.96
0.88
0.73
0.98
0.72
0.95
0.71
0.91
0.87
0.78
0.00
0.03
0.69
0.91
0.00
1.00
D-28 Draft Document
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T-lf-1
T-3b-l
T-5a-l
ETC
ETC
ETC
39.76
47.47
46.37
141
208
634
0.52
0.88
0.07
0.70
0.92
0.17
Table D-ll. Fathead minnow first brood larval malformations and adult whole-body (WB)
selenium concentrations (dw) for each site (± 1SE); CON = control, SCC = Sand Creek
at Colfax Avenue bridge, TGC = Tollgate Creek, and ETC = East Tollgate Creek.
Brood
Code
T-lf-1
T-2a-l
T-6d-l
T-3b-l
T-3a-l
T-la-1
T-lf-1
T-5d-l
T-4d-l
T-3d-l
T-4a-l
T-6a-l
T-4a-l
T-2a-l
T-lf-1
T-5a-l
T-3b-l
Treatmen
t
CON
CON
CON
CON
CON
CON
CON
CON
CON
CON
SCC
SCC
TGC
TGC
ETC
ETC
ETC
Maternal
WBSe
Cone dw
(mg/kg)
1.94
2.25
2.52
2.64
2.71
2.90
3.24
3.33
3.43
3.67
8.35
9.25
31.99
32.29
39.76
46.37
47.47
Total
Larvae
11
141
117
81
96
14
189
95
164
49
3
86
190
91
65
39
150
Spinal
Incidence
9
3
2
4
1
7
8
4
3
6
0
19
5
8
5
44
11
%larvae
w/o
spinal
deformity
91
97
98
96
99
93
92
96
97
94
100
81
95
92
95
56
89
%larvae
w/o
craniofacial
deformity
100
99
99
98
100
93
98
97
98
92
100
91
97
90
95
54
95
%larvae
w/o
finfold
deformity
100
98
99
99
100
93
98
99
99
94
100
92
97
91
98
54
96
%larvae
w/o edema
100
96
97
98
100
93
94
98
96
90
100
90
97
90
94
54
91
Total
GSI
Score
1
24
16
12
1
10
53
20
28
29
0
71
41
78
20
152
89
Table D-12. Authors calculation and comparison of fathead minnow larval deformity
estimates using probit analysis and TRAP.
Effect
Edema
Finfold
Skeletal
Endpoint
ECio
EC10
EC10
Probit Results
WB [Se]
mg/kg,
dw (±SE)
39.48 ± 16.21
68.55 ±27.26
27.80 ±9.53
TRAP Results
WB [Se] mg/kg,
dw (95% CL)
45.78
(40.95-51.20)
48.31
(39.41 -59.21)
46.08
(41.94-50.62)
Probit Results
Ovary [Se]
mg/kg,
dw (±SE)
52.99 ± 19.99
87.95 ±32.16
38.67 ± 12.32
TRAP Results
Ovary [Se] mg/kg,
dw (95% CL)
61.43
(55.04-68.55)
64.81
(53.01-79.24)
61.82
(56.36-67.80)
D-29 Draft Document
-------
Craniofacial
All
abnormalities
All
abnormalities
except edema
EC10
EC10
EC10
53.86± 18.77
16.98 ±5.38
21.35 ±6.45
47.41
(38.92-57.76)
45.50
(41.10-50.37)
45.69
(41.10-50.79)
70.83 ±22.84
24.23 ± 7.06
30.32 ±8.51
63.56
(52.37-77.16)
61.06
(55.26-67.48)
61.27
(55.23-67.97)
Figure D-l. The fraction survival of embryos (left) and the fraction of embryos successfully
fertilized (right) relative to the maternal whole body selenium concentration.
1.2 •
1 •
I 0.8 •
I 0.6-
o
•a
S 0.4 •
u.
0.2 •
0 •
» . »
* »
«
«
3 0.5 1 1.5 2
Log (maternal whole body [Se] \iglg dw)
1.20 •
1.00 •
| 0.80 •
c
I 0.60-
N
5 0.40 •
u.
0.20 •
0.00 •
t
*-,
• *• .
«
* • •
) 0.5 1 1.5 2
Log (maternal whole body [Se] \iglg dw)
Figure D-2. Percent 2-day post-hatch larvae without edema (A), finfold deformity (B), craniofacial
deformity (C), and spinal deformity (D) relative to maternal whole body selenium
concentration.
D-30 Draft Document
-------
120 •
100 •
1 80 •
o>
•o
| 60-
5? 40 •
20 •
0 •
A
*•'•*.
*
3 0.5 1 1.5 2
Log (maternal whole body [Se] \iglg dw)
120 •
£ 10° '
E
£ 80 •
•o
o 40 •
58 20 •
0 •
t
B
••v.
*
) 0.5 1 1.5 2
Log (maternal whole body [Se] \iglg dw)
c
£100 •
I 80-
•a
1 BU'
0
a 40 •
O
o
1 20 -
5?
•*V. * ••«
* * • »
,
0 0.5 1 1.5 2
Log (maternal whole body [Se] \iglg dw)
I
s
•o
re
c
Q.
V)
^o
120 •
100 •
80 •
60 •
40 •
20 •
0 •
t
D
.**•/• * f.
*
t
) 0.5 1 1.5 2
Log (maternal whole body [Se] \iglg dw)
D-31 Dra//1 Document
-------
Figure D-3. Percent 2-day post-hatch larvae Graduated Severity Index (GSI) relative to maternal
whole body selenium concentration
(D
0
o
U)
8
160 -i
140 -
120 -
100 _
80 -
60 -
40 -
20 _
o
*
1
*
*
*
*
.0
A. A !
X
-*-*- -* I
0.00 10.00 20.00 30.00 40.00 50.00
Maternal whole body[Se] |jg/g dw)
Other Data - Chronic Studies with Invertebrate Species
A limited number of studies have evaluated the effects of selenite on invertebrate species, an important
prey item for fish and birds as summarized by Debruyn and Chapman (2007). The following three tests
with a rotifer, and annelid, and an insect (mayfly) were found suitable for establishing species sensitivity.
Dobbs et al. (1996) exposed Brachionus calyciflorus to selenate in natural creek water for 25 days in a
three-trophic level food chain test system. This is one of two laboratory-based experiments (also see
Bennett et al. 1986) that involved exposing algae to selenium (in this case as sodium selenate) in water,
and subsequently feeding the algae to rotifers which were in turn fed to fish (fathead minnows). In this
particular study, the rotifers and fish were exposed to the same concentrations of sodium selenate in the
water as the algae, but received additional selenium from their diet (i.e., the algae fed to rotifers and the
rotifers fed to fish). The overall exposure lasted for 25 days. Rotifers did not grow well at concentrations
exceeding 108.1 (ig Se/L in water, and the population survived only 6 days at selenium concentrations
equal to or greater than 202.4 (ig Se/L in the water (40 (ig Se/g dw in the algae). Regression analysis of
untransformed growth data (dry weight) determined 4 day post-test initiation resulted in a calculated ECi0
of 37.84 (ig Se/g dw tissue.
D-32 Draft Document
-------
Although not intended to be a definitive toxicity study for this invertebrate, Besser et al. (2006) evaluated
the bioaccumulation and toxicity of selenized yeast to the oligochate, Lumbriculus variegatus, which was
intended to be used for dietary exposure in subsequent studies with the endangered desert pupfish,
Cyprinidon macularius. Oligochaetes fed selenized-yeast yeast diets diluted with nutritional yeast (54 to
210 mg Se/kg) had stable or increasing biomass and accumulated Se concentrations as high as 140 mg/kg
dw. The oligochaetes fed the undiluted selenized-yeast (826 ug/g Se dry wt.) showed reduced biomass.
The effect level is considered >140 mg Se/kg dw.
Conley et al. (2009) exposed mayfly larvae (Centroptilum triangulifef) to dietary selenium contained in
natural periphyton biofilms to eclosion. The periphyton fed to the mayfly larvae were exposed to
dissolved selenite (radiolabeled 75Se) in November 2008 (12.6 and 13.9 (ig/L) and in January 2009 ( 2.4,
2.4, 4.9, 10.3, and 10.7 (ig/L). Periphyton bioconcentrated Se an average of 1113-fold over the different
aqueous Se concentrations (Table D-10). Twenty 4 to 6-day old mayfly larvae were exposed for 4.5 to 6
weeks to each of the periphyton diets until the larvae eclosed to subimagos. The subimagos were allowed
to emerge to the adult imago stage which deposited their egg masses in Petri dishes. Selenium was
measured in postpartum adults along with their dry weights and clutch size.
Table D-13. Selenium Concentrations in Water Exposed to Periphyton, Periphyton and Mayfly
Adults
Treatment
5A
5B
10A
20C
20D
20A
20B
Dissolved [Se] exposed
to periphyton, ug/L
2.4
2.4
4.9
10.3
10.7
12.6
13.9
[Se] in periphyton,
mg/kg dw
2.2
2.0
4.4
8.7
11.3
25.5
17.5
[Se] in mayfly adult,
mg/kg dw
4.2
5.7
9.7
16.2
27.5
56.7
34.8
Selenium increased in concentration from periphyton to the adult mayflies (trophic transfer factor) an
average of 2.2-fold (Table D-13). The authors observed a decrease in fecundity as maternal postpartum Se
concentrations increased. Fecundity was also related to growth of the mayflies. The authors observed a
reduction in fecundity for this mayfly when they were fed diets containing more than 11 mg Se/kg dw.
This threshold is considered the effect value for this study. Using the trophic transfer factor of 2.2, the
D-33 Draft Document
-------
periphyton Se concentration of 11 mg/kg dw translates to an adult mayfly Se concentration of 24.2 mg/kg
dw.
The following invertebrate studies were inconclusive for establishing species sensitivity because of
limitations in the experimental designs, as explained for each.
Malchow et al. (1995) fed fourth instar Chironomus decorus midge larvae a diet of seleniferous algae
under laboratory conditions for 96 hours. For algae cultured with selenite, a larval tissue concentration of
4.05 Og Se/g dry weight resulted in a 46% reduction in growth relative to the controls. At a larval tissue
concentration of 8.6 Og Se/g dry weight, larval growth was reduced by only 39%. Since the study only
reported two exposure concentrations, it is unclear if the tissue effect concentration at 4.05 Og Se/g dry
weight is real or an anomaly. Additional exposure concentrations and subsequent effect levels are needed
to resolve this issue.
Malchow et al. (1995) also fed fourth instar Chrionomus decorus midge larvae a diet of algae cultured
with selenate, and the midge larvae were exposed under laboratory conditions for 96 hours. A dietary
exposure of 2.11 (ig Se/g dry weight significantly reduced larval growth (15% reduction) at tissue
concentrations of 2.55 (ig Se/g dry weight. At a larval tissue concentration of 6.62 (ig Se/g dry weight,
growth was reduced 20% relative to the controls. The 15-20% reduced growth at larval tissue
concentrations 2.55 (ig Se/g dry weight may be statistically significant, but not biologically meaningful.
In addition, exposure to only two selenium concentrations precludes confirmation of a dose-response.
Alaimo et al. (1994) also exposed C. decorus midge larvae to selenite diet, but the selenium source was
from field contaminated widgeongrass (Ruppia maritime?). Ruppia stems and leaves were collected from
four selenium contaminated evaporation ponds located in the San Joaquin Valley of California. Three-day
old larvae were exposed to each of the four treatment diets (Ruppia from each pond) plus a Cerophyll
control for 14 days (egg to pupation), with the moderately hard reconstituted water renewed at day 7 and
every three days thereafter. The growth (weight) of exposed larvae was significantly reduced in all of the
selenium treatments when compared to the controls. The lowest effect level was observed for the
Westlake pond (primarily selenite), where growth was reduced 40 percent relative to the controls at a
larval tissue concentration below the detection level (1.0 ppm dry weight, or 1.0 (ig Se/g dry weight).
These results are suspect because the field collected Ruppia likely contained contaminants other than
selenium, the control organisms were fed a different diet (Cerophyll), and the single concentration
exposure is difficult to defend.
D-34 Draft Document
-------
Other Data - Field Study West Virginia Impoundments
In response to the USEPA (2004) draft whole fish tissue criterion for selenium, the West Virginia
Department of Environmental Protection (2010) initiated a study to assess selenium bioaccumulation
among fishes residing in the State's lakes and streams. A focus of the study was the collection and
evaluation of bluegill, Lepomis macrochirus, larvae (ichthyoplankton) from selected waterbodies since
2007, based on concerns regarding fish population health at locations subjected to elevated selenium
inputs, particularly during the more sensitive developmental life stages of fishes (e.g. yolk-sac larvae).
Also, in 2009, WVDEP began acquiring data about selenium concentrations within fish eggs of various
species within reference and selenium-impacted waters. WVDEP also conducted deformity surveys of
adult fishes in selenium enriched waters as well as at reference locations in 2008-2009.
WVDEP scientists found that larval deformity rates were variable throughout the study duration but were
nonetheless correlated with waterborne selenium exposure. Reference locations produced age-based
larval bluegill subsamples (24-168 hours) with low deformity rates (0 - 1.27%); whereas, locations with
seleniferous inputs exhibited bluegill deformity rates ranging from 0% to 47.56% in developmental stages
up to 312 hours. Maximum deformity rates among staged bluegill subsamples as determined through
these evaluations were 19.28%, representing specimens collected from selenium-enriched waters.
Concentrations of selenium within fish eggs also varied according to study location and ranged from <0.8
mg/kg dry weight among bluegill eggs at the control site to 64.62 mg/kg dry weight among largemouth
bass, Micropterus salmoides, eggs collected from selenium-enriched waters. Searches for more mature,
yet developmentally-deformed fishes revealed increased deformity rates (14%) among largemouth bass
residing in a selenium impacted reservoir as compared to deformity rates among largemouth bass found in
the reference lake (0%). The data on egg selenium concentrations are not adequate for constructing a
concentration-response curve. Nevertheless, the overall deformity rate in the contaminated Upper Mud
River Reservoir was 5% among 10,000 individual fish, average egg selenium concentration 9.8 mg/kg
dw. The overall deformity rate in the reference Plum Orchard Lake was 0.5% among 13,000 individuals,
average egg selenium concentration nondetectable or <0.8 mg/kg dw.
D-35 Draft Document
-------
APPENDIX E: Toxicity of Selenium to Aquatic Plants
E-l Draft Document
-------
Selenite
Data are available on the toxicity of selenite to 13 species of freshwater algae and plants (Table E-l).
Results ranged from an LC50 of 70,000 ug/L for the green alga, Chlorella ellipsoidea (Shabana and El-
Attar 1995) to 522 ug/L for incipient inhibition of the green alga, Scenedesmus quadricauda (Bringmann
and Kuhn 1977a, 1978a,b, 1979, 1980b). Foe and Knight (Manuscript) found that 75 ug/L decreased the
dry weight of Selenastrum capricornutum (Table E-l). Wehr and Brown (1985) reported that 320 ug/L
increased the growth of the alga Chrysochromulina breviturrita. Thus, the sensitivities of freshwater
algae to selenite cover about the same range as the acute and chronic sensitivities of freshwater animals.
The 96-hr EC50 for the saltwater diatom, Skeletonema costatum, is 7,930 ug/L, based on reduction in
chlorophyll a (Table E-l). Growth of Chlorella sp., Platymonas subcordiformis, and Fucus spiralis
increased at selenite concentrations from 2.6 to 10,000 ug/L (Table E-l). Other marine algae exposed to
selenite from 14 to 60 days had no observed effect concentrations (NOAEC) that ranged from 1,076 to
107,606 ug/L. These data suggest that saltwater plants will not be adversely affected by concentrations of
selenite that do not affect saltwater animals.
Selenate
Growth of several species of green algae was affected by concentrations ranging from 100 to 40,000 ug/L
(Table E-l). Blue-green algae appear to be more tolerant to selenate with 1,866 ug/L being the lowest
concentration reported to affect growth (Kiffney and Knight 1990). Kumar (1964) found that a blue-green
alga developed and lost resistance to selenate. The difference in the sensitivities of green and blue-green
algae to selenate might be of ecological significance, particularly in bodies of water susceptible to
nuisance algal blooms. For example, Patrick et al. (1975) reported that a concentration of 1,000 ug/L
caused a natural assemblage of algae to shift to a community dominated by blue-green algae.
The saltwater coccolithophore, Cricosphaera elongata, had reduced growth when exposed to 41,800 ug/L
selenate for 14 days (Boisson et al. 1995). Seven other saltwater algal species investigated by Wong and
Oliveira (1991a) exhibited NOEC growth values that ranged from 1,043 to 104,328 ug/L. At 10,000 ug/L,
selenate is lethal to four species of saltwater phytoplankton and lower concentrations increase or decrease
growth (Table E-l). Wheeler et al. (1982) reported that concentrations as low as 10 ug/L reduced growth
of Porphyridium cruentum (Table E-l).
E-2 Draft Document
-------
Although selenite appears to be more acutely toxic than selenate to most aquatic animals, this does not
seem to be true for aquatic plants. Selenite and selenate are about equally toxic to the freshwater algae
Anabaena cylindrica, Anabaena flos-aquae, Anabaena variabilis, Anacystis nidulans, and Scenedesmus
dimorphus (Kiffney and Knight 1990; Kumar and Prakash 1971; Moede et al. 1980) and the saltwater
algae Agemenellum quadroplicatum, Chaetoceros vixvisibilis and Amphidinium carterae (Wong and
Oliveira 1991a). The two oxidation states equally stimulated growth of Chrysochromulina breviturrita
(Wehr and Brown 1985). On the other hand, selenate is more toxic than selenite to the freshwater
Selenastrum capricornutum (Richter 1982; Ibrahim and Spacie 1990) and the saltwater Chorella sp.,
Platymonas subcordiformis and Nannochloropsis oculata (Wheeler et al. 1982; Wong and Oliveira
1991a). In addition, Fries (1982) found that growth of thalli of the brown macroalga, Fucus spiralis, was
stimulated more by exposure to selenite at 2.605 ug/L than to the same concentration of selenate.
A Final Plant Value, as defined in the Guidelines, cannot be obtained because no test in which the
concentrations of selenite or selenate were measured and the endpoint was biologically relevant has been
conducted with an important aquatic plant species.
E-3 Draft Document
-------
Table E-l. Toxicity of Selenium to Aquatic Plants
Species
Green alga,
Chlorella vulgaris
Green alga,
Chlorella
ellipsoidea
Green alga,
Scenedesmus
dimorphus
Green alga,
Scenedesmus
quadricauda
Green alga,
Scenedesmus
quadricauda
Green alga,
Selenastrum
capricornutum
Green alga,
Selenastrum
capricornutum
Blue-green alga,
Anabaena
constricta
Blue-green alga,
Anabaena
cylindrica
Blue-green alga,
Anabaenaflos-
aquae
Blue-green alga,
Anabaena
variabilis
Blue-green alga,
Anacystis nidulans
Blue-green alga,
Microcystis
aeruginisa
Alga,
Euglena gracilis
Chemical
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
Sodium
selenite
-
Hardness
(mg/L as Duration
CaCO,) (days) Effect
FRESHWATER SPECIES
Selenium (IV)
90-120 Reduced
growth
7 EC50
14 Reduced
growth
8 Incipient
inhibition
8 Incipient
inhibition
4 EC50
6 EC50
7 EC50
14 Reduced
growth
10 Reduced
chlorophyll a
6-18 LC50
10-18 LC50
8 Incipient
inhibition
1 5 Reduced
growth
Concentration
5,480
70,000
24,000
522
2,500
2,900
65,000
67,000
24,000
1,866
15,000b
30,000b
9,400
(9,300)
5,920
Reference
De Jong 1965
Shabana and El-
Attar 1995
Moede et al.
1980
Bringmann and
Kuhnl977a;
1978a,b; 1979;
1980b
Bringmann and
Kuhnl959a
Richter 1982
Ibrahim and
Spacie 1990
Shabana and El-
Attar 1995
Moede et al.
1980
Kiffney and
Knight 1990
Kumar and
Prakash 1971
Kumar and
Prakash 1971
Bringmann and
Kuhnl976;
1978a,b
Bariaud and
Mestre 1984
E-4 Draft Document
-------
Hardness
(mg/L as Duration
Concentration
Species
Duckweed,
Lemna minor
Duckweed,
Lemna minor
Duckweed,
Lemna minor
Green alga,
Ankistrodesmus
falcatus
Green alga,
Scenedesmus
dimorphus
Green alga,
Scenedesmus
obliquus
Green alga,
Selenastrum
capricornutum
Green alga,
Selenastrum
capricornutum
Green alga,
Selenastrum
capricornutum
Blue-green alga,
Anabaena
cylindrica
Blue-green alga,
Anabaenaflos-
aquae
Blue-green alga,
Anacystis nidulans
Blue-green alga,
Anabaena
viriabilis
Blue-green alga,
Microcoleus
vaginatus
Duckweed,
Lemna minor
Chemical
Sodium
selenite
Sodium
selenite
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
-CaCOs) (days)
4
14
14
Selenium (VI)
14
14
14
14
4
6
14
10
6-18
10-18
14
14
Effect
EC50
EC50
(mult, rate)
NOEC
(mult, rate)
Did not
reduce
growth
Reduced
growth
Reduced
growth
Reduced
growth
EC50
EC50
Reduced
growth
Reduced
chlorophyll a
EC50
EC50
Reduced
growth
EC50
(mult, rate)
2,400
3,500
800
10
22,100
100
300
199
<40,000
22,100
1,866
39,000b
17,000b
10,000
11,500
Reference
Wang 1986
Jenner and
Janssen-
Mommen 1993
Jenner and
Janssen-
Mommen 1993
Vocke et al.
1980
Moede et al.
1980
Vocke et al.
1980
Vocke et al.
1980
Richterl982
Ibrahim and
Spacie 1990
Moede et al.
1980
Kiffney and
Knight 1990
Kumar and
Prakash 1971
Kumar and
Prakash 1971
Vocke et al.
1980
Jenner and
Janssen-
Mommen 1993
Duckweed,
Sodium
14 NOEC
E-5 Draft Document
>2,400
Jenner and
-------
Species
Lemna minor
Species
Green alga,
Dunaliella
tertiolecta
Cyanophyceae alga,
Agemenellum
quadruplicatum
Diatom,
Chaetoceros
vixvisibilis
Diatom,
Skeletonema
costatum
Coccolithophore,
Cricosphaera
elongata
Dinoflagellate,
Amphidinium
carterae
Dinoflagellate,
Peridinopsis borgei
Eustigmatophyceae
alga,
Nannochloropsis
oculata
Pyrmnesiophyceae
alga,
Isochrysis galbana
Pyrmnesiophyceae
alga,
Pavlova lutheri
Green alga,
Dunaliella
tertiolecta
Cyanophyceae alga,
Agemenellum
Chemical
selenate
Chemical
Sodium
selenite
Sodium
selenite
Sodium
selenite
Selenious
acid0
Sodium
selenite
Sodium
selenite
Selenium
oxide
Sodium
selenite
Sodium
selenite
Sodiun selenite
Sodium
selenate
Sodium
selenate
Hardness
(mg/L as Duration
CaCO,) (days) Effect
(mult. Rate)
Salinity Duration
(g/kg) (days) Effect
SALTWATER SPECIES
Selenium (IV)
60 NOEC growth
60 NOEC growth
60 NOEC growth
4 EC50 (reduction
in chlorophyll a)
14 Reduced growth
60 NOEC growth
70-75 Maximum
growth
60 NOEC growth
60 NOEC growth
60 NOEC growth
Selenium (VI)
60 NOEC growth
60 NOEC growth
Concentration
(fig/Lf
Concentration
iMgM
1,076
10,761
1,076
7,930
4,570
10,761
0.01-0.05
107,606
1,076
1,076
104,328
10,433
Reference
Janssen-
Mommen 1993
Reference
Wong and
Oliveiral991a
Wong and
Oliveiral991a
Wong and
Oliveiral991a
U.S. EPA 1978
Boisson et al.
1995
Wong and
Oliveiral991a
Lindstrom 1985
Wong and
Oliveiral991a
Wong and
Oliveiral991a
Wong and
Oliveiral991a
Wong and
Oliveiral991a
Wong and
Oliveiral991a
E-6 Draft Document
-------
Salinity Duration
Concentration
Species
quadruplicatum
Diatom,
Chaetoceros
vixvisibilis
Coccolithophore,
Cricosphaera
elongata
Dinoflagellate,
Amphidinium
carterae
Eustigmatophyceae
alga,
Nannochloropsis
oculata
Pyrmnesiophyceae
alga,
Isochrysis galbana
Pyrmnesiophyceae
alga,
Pavlova lutheri
Chemical
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
Sodium
selenate
(a/kg) (days) Effect (fig/Lf Reference
60 NOEC growth 1,043 Wong and
Oliveiral991a
14 Reduced growth 41,800 Boissonetal.
1995
60 NOEC growth 10,433 Wong and
Oliveiral991a
60 NOEC growth 10,433 Wong and
Oliveiral991a
60 NOEC growth 10,433 Wong and
Oliveiral991a
60 NOEC growth 104,328 Wong and
Oliveiral991a
a Concentration of selenium, not the chemical.
b Estimated from published graph.
0 Reported by Barrows et al. (1980) in work performed under the same contract.
E-7 Draft Document
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APPENDIX F: Unused Data
F-l Draft Document
-------
Based on the requirements set forth in the guidelines (Stephen et al. 1985) the following studies are not
acceptable for the following reasons and are classified as unused data. Note the acceptance of chronic
toxicity data included diet and field exposures where selenium was the dominant toxicant.
Studies Were Conducted with Species That Are Not Resident in North America
Ahsanullah and Brand (1985) Gotsis(1982) Ringdal and Julshamn (1985)
Ahsanullah and Palmer Hiraika et al. (1985) Rouleau et al. (1992)
(1980) Juhnke and Ludemann (1978) Sastry and Shukla (1994)
Baker and Davies (1997) Kitamura (1990) Savant and Nilkanth (1991)
Barghigiani et al. (1993) Manoharan and Prabakaran Shultz and Ito (1979)
Chidambaram and Sastry (1994) Srivastava and Tyagi (1985)
(1991a,b) Minganti et al. (1994, 1995) Takayanagi (2001)
Congiuetal. (1989) Niimi and LaHam (1975, Tomasik et al. (1995b)
Cuvin and Furness (1988) 1976) Tian and Liu (1993)
Fowler and Benayoun Regoli (1998) Wrench (1978)
(1976a,b) Regoli and Principato (1995)
Gaikwad (1989) Rhodes et al. (1994)
Deelstra et al. (1989), Forsythe and Klaine (1994), Okasako and Siegel (1980) and Petrucci et al. (1995)
conducted tests with brine shrimp species that are too atypical to be used in derving national criteria.
These Studies or Reviews Contain Relevant Data That Have Been Published Elsewhere
Adams and Johnson (1981) Eisler (1985) McKee and Wolf (1963)
Biddinger and Gloss (1984) Hall and Burton (1982) National Research Council
Bowie et al. (1996) Hodson and Hilton (1983) (1976) Neuhold (1987)
Brandao et al. (1992) Hodson et al. (1984) NCDNR&CD (1986)
Brooks (1984) Jenkins (1980) Peterson and Nebeker (1992)
Burton and Stemmer (1988) Kaiser et al. (1997) Phillips and Russo (1978)
Chapman etal. (1986) Kay (1984) Presser (1994)
Davies (1978) LeBlanc (1984) Roux et al. (1996)
Debruyn and Chapman Lemly (1993c, 1996ab, Swift (2002)
(2007) 1997d) Thompson etal. (1972)
Devillers etal. (1988) Lemly and Smith (1987) Versar(1975)
F-2 Draft Document
-------
Authors Did Not Specify the Oxidation State of Selenium Used in Study
Greenberg and Kopec (1986) Kapu and Schaeffer (1991)
Hutchinson and Stokes Kramer et al. (1989) Rauscher (1988)
(1975) Mahan et al. (1989) Snell et al. (1991b)
Not Useful Because of No Effects Observed at Exposure Concentrations or Insufficient Number of
Treatments
Muscatello and Janz (2009)
Pyle et al. (2005)
Schlenk et al (2003)
Chronic Study with no Dietary Exposure
Hopkins etal. (2002)
Oti (2005)
Rowe (2003)
Teh et al. (2002)
Selenium Was a Component of an Effluent, Fly Ash, Formulation, Mixture, Sediment or Sludge
Apte et al. (1987) Cherry et al. (1987) Eriksson and Forsberg (1992)
Baer et al. (1995) Cieminski and Flake (1995) Eriksson and Pedros-Alio
Baker et al. (1991) Clark et al. (1989) (1990)
Berg et al. (1995) Cooke and Lee (1993) Fairbrother et al. (1994)
Besser et al. (1989) Cossu et al. (1997) Fava et al. (1985a,b)
Biedlingmaier and Schmidt Coyle et al. (1993) Feroci et al. (1997)
(1989) Crane etal. (1992) Finger and Bulak (1988)
Bjoernberg (1989) Crock et al. (1992) Finley(1985)
Bjoernberg et al. (1988) Cushman et al. (1977) Fisher and Wente (1993)
Bleckmann et al. (1995) Davies and Russell (1988) Fjeld and Rognerud (1993)
Boisson et al. (1989) de Peyster et al. (1993) Fletcher et al. (1994)
Bondavalli et al. (1996) Dickman and Rygiel (1996) Follett (1991)
Bowmer et al. (1994) Dierenfeld et al. (1993) Gerhardt (1990)
Brieger et al. (1992) Doebel et al. (2004) Gerhardt et al. (1991)
Burton and Pinkney (1984) Drndarski etal. (1990) Gibbs and Miskiewicz (1995)
Burton et al. (1983, 1987a) Graham et al. (1992)
F-3 Draft Document
-------
Gundersonetal. (1997)
Hall (1988)
Hall etal. (1984, 1987,
1988,1992)
Hamilton et al. (1986, 2000)
Harrison etal. (1990)
Hartwell et al. (1987ab, 1988,
1997)
Hatcher etal. (1992)
Haynes etal. (1997)
Hayward etal. (1996)
Hellouetal. (1996b)
Henebry and Ross (1989)
Henny etal. (1989, 1990,
1995)
Hildebrandetal. (1976)
Hjeltnes and Julshman (1992)
Hockett and Mount (1996)
Hodson(1990)
Hoffman etal. (1988, 1991)
Homziak etal. (1993)
Hopkins etal. (2000)
Hopkins etal. (2004)
Hothem and Welsh (1994a)
Jackson (1988)
Jackson etal. (1990)
Jacquez etal. (1987)
Jay and Muncy (1979)
Jayasekera(1994)
Jayasekera and Rossbach
(1996)
Jenner and Bowmer (1990)
(1992)
Jin etal. (1997)
Jorgensen and Heisinger
(1987)
Karlson and Frankenberger
(1990)
Kemble etal. (1994)
Kennedy (1986)
Kersten etal. (1991)
King and Cromartie (1986)
King etal. (1991, 1994)
Kluseketal. (1993)
Koh and Harper (1988)
Koike etal. (1993)
Krishnajaetal. (1987)
Kruuk and Conroy (1991)
Kuehl and Haebler( 1995)
Kuehl etal. (1994)
Kuss etal. (1995)
Landau etal. (1985)
Livingston et al. (1991)
Lobel etal. (1990)
Luoma and Phillips (1988)
Lundquistetal. (1994)
Lyle (1986)
MacFarlane etal. (1986)
Mann and Fyfe (1988)
Marcogliese et al. (1987)
Marvin and Ho well.
(1997)Maureretal (1999)
McCloskey and Newman
(1995)
McCloskey etal. (1995)
McCrea and Fischer (1986)
McLean etal. (1991)
Mehrle etal. (1987)
Metcalf-Smith (1994)
Micallef and Tyler (1989)
Mikac etal. (1985)
Miles and Tome (1997)
Miller etal. (1996)
Misitano and Schiewe (1990)
Moore (1988)
Munawar and Legner (1993)
Muskettetal. (1985)
Naddy etal. (1995)
Nielsen and Bjerregaard
(1991)
Norman etal. (1992)
Nuutinen & Kukkonen
(1998)
Oberbach and Hartfiel (1987,
1988)
Oberbach etal. (1989)
Ohlendorf etal. (1989, 1990,
1991)
Olson and Welsh (1993)
Peters etal.(1999)
Phillips and Gregory (1980)
Pratt and Bowers (1990)
Presser and Ohlendorf (1987)
Prevot and Sayer-Gobillard
(1986)
Pritchard(1997)
Pyle etal. (2001)
Reash et al. (1988, in press)
Rhodes and Burke (1996)
F-4 Draft Document
-------
Ribeyre et al. (1995) Sorenson and Bauer (1983) Wang et al. (1992, 1995)
Rice et al. (1995) Specht et al. (1984) Welsh (1992)
Riggs and Esch (1987) Steele et al. (1992) Weres et al. (1990)
Riggs et al. (1987) Stemmer et al. (1990) White and Geitner (1996)
Robertson et al. (1991) Summers et al. (1995) Wiemeyer et al. (1986)
Roper et al. (1997) Thomas et al. (1980b) Wildhaber and Schmitt
Rowe et al. (1996) Timothy et al. (2001) (1996)
Russell et al. (1994) Trieff et al. (1995) Williams et al. (1989)
Ryther et al. (1979) Wolfe et al. (1996)
Turgeon and O^iConner
Saiki and Jenings (1992) Wolfenberger (1987)
Saiki and Ogle (1995) (1991) Wong and Chau (1988)
Saleh et al. (1988) Twerdok et al. (1997) Wong et al. (1982)
Seelye et al. (1982) Unsal (1987) Wu et al. (1997)
Sevareid and Ichikawa Van Metre and Gray (1992) Yamaoka et al. (1994)
(1983) Wahl et al. (1994) Zagatto et al. (1987)
Skinner (1985) Wandan and Zabik (1996) Zaidi et al. (1995)
Somerville et al. (1987) Zhang et al. (1996)
Exposed enzymes, excised tissue or tissue extractor
Tripathi and Pandey (1985) and Heinz (1993b) used test organisms that had been previously exposed to
pollutants in food or water.
Albers et al. (1996) Bell et al. (1984, 1985, Byl et al. (1994)
Al-Sabti (1994, 1995) 1986a,b, 1987ab) Chandy and Patel (1985)
Arvy et al. (1995) Berges and Harrison (1995) Chen et al. (1997)
Augier et al. (1993a, b) Blondin et al. (1988) Cheng et al. (1993)
Avery et al. (1996) Boisson et al. (1996) Christensen and Tucker
Baatrup (1989) Bottino et al. (1984) (1976)
Baatrup and Dansher (1987) Braddon (1982) Dabbert and Powell (1993)
Baatrup et al. (1986) Braddon-Galloway and DeQuiroga et al. (1989)
Babich et al. (1986, 1989) Balthrop (1985) Dierickx (1993)
Barrington et al. (1997) Bradford et al. (1994a,b) Dietrich et al. (1987)
Becker et al. (1995a,b) Brandt et al. (1990) Dillio et al. (1986)
F-5 Draft Document
-------
Doyotteetal. (1997)
Drotaretal. (1987)
Dubois and Callard (1993)
Ebringeretal. (1996)
Engberg and Borsting (1994)
Engbergetal. (1993)
Eunetal. (1993)
Foltinova and Gajdosova
(1993)
Foltinova etal. (1994)
Freeman and Sanglang
(1977)
Grubor-Lajsic et al. (1995)
Hait and Sinha( 1987)
Hanson (1997)
Heisinger and Scott (1985)
Heisinger and Wail (1989)
Henderson etal. (1987)
Henny and Bennett (1990)
Hoffman and Heinz (1988,
1998)
Hoffman etal. (1989, 1998)
Hoglund(1991)
Hontelaetal. (1995)
Hsu etal. (1995)
Hsu and Goetz (1992)
Ishikawaetal. (1987)
James etal. (1993)
Jovanovic et al. (1995, 1997)
Kai etal. (1995)
Kedziroski etal. (1996)
Kelly etal. (1987)
Kralj and Stunja( 1994)
Lalitha and Rani (1995)
Lanetal. (1995)
Lemaire etal. (1993)
Livingstone et al. (1992)
Low and Sin (1995, 1996)
Micallef and Tyler (1990)
Montagnese et al. (1993)
Murataetal. (1996)
Nakonieczny (1993)
Neuhierl and Boeck (1996)
Nigro (1994)
Nigro etal (1992)
Norheim and Borch-Iohnsen
(1990)
Norheim etal. (1991)
O\iBrien etal. (1995)
Olson and Christensen (1980)
Overbaugh and Fall (1985)
Palmisano etal. (1995)
Patel etal. (1990)
Patel and Chandy (1987)
Perez Campo etal. (1990)
Perez-Trigo etal. (1995)
Phadnis etal. (1988)
Price and Harrison (1988)
Radyetal. (1992)
Rani and Lalitha (1996)
Regoli etal. (1997)
Schmidt etal. (1985)
Schmittetal. (1993)
Segneretal. (1994)
Sen etal. (1995)
Shigeoka et al. (1990, 1991)
Siwicki etal. (1994)
Srivastava and Srivastava
(1995)
Sun etal. (1995)
Takedaetal. (1992a,b,(1993,
1997)
Treuthardt(1992)
Vazquez etal. (1994)
Veenaetal. (1997)
Wise etal. (1993a,b)
Wong and Oliveira (1991b)
Yokotaetal. (1988)
Test procedures test material or results were not adequately described by Botsford (1997), Botsford et al.
(1997, 1998), Bovee (1978), Gissel-Nielsen and Gissel-Nielsen (1973, 1978), Greenberg and Kopec
(1986), Mauk (2001), and Nassos et al. (1980) or when the test media contained an excessive amount
(>200 ug/L) of EDTA (Riedel and Sanders (1996).
F-6 Draft Document
-------
Some data obtained from tests conducted with just one exposure concentration to evaluate acute or
chronic toxicity were not used (e.g., Bennett 1988; Heinz and Hoffman 1998; Munawar et al. 1987;
Pagano et al. 1986; Wolfenberger 1986).
Kaiser (1980) calculated the toxicities of selenium(IV) and selenium(VI) to Daphnia magna based on
physiochemical parameters. Kumar (1964) did not include a control treatment in the toxicity tests. The
daphnids were probably stressed by crowding in the tests reported be Schultz et al. (1980). Siebers and
Ehlers (1979) exposed too few test organisms as did Owsley (1984) in some tests.
Selenium Concentrations Reported in Wild Aquatic Organisms Were Insufficient to Calculate BAF
Abdel-Moati and Atta (1991)
Adeloju and Young (1994)
Aguirreetal. (1994)
Akesson and Srikumar
(1994)
Aksnesetal. (1983)
Allen and Wilson (1990)
Ambulkar et al. (1995)
Amiardetal. (1991, 1993)
Andersen and Depledge
(1997)
Andreev and Simeonov
(1992)
Angulo(1996)
Arrulaetal. (1996)
Arway (1988)
Ashton(1991)
Augieretal. (1991, 1993,
1995a,b)
Augspurger et al. (1998)
Averyetal. (1996)
Badsha and Goldspink (1988)
Baines and Fisher (2001)
Baldwin and Maher (1997)
Baldwin etal. (1996)
Barghigiani(1993)
Barghigiani etal. (1991)
Baron etal. (1997)
Batley(1987)
Baumann and Gillespie
(1986)
Baumann and May (1984)
Beal (1974)
Beck etal. (1997)
Belandetal. (1993)
Beliaeffetal. (1997)
Bell and Cowey (1989)
Benemariya et al. (1991)
Berry etal. (1997)
Bertram etal. (1986)
Besseretal. (1994, 1993)
Birkner(1978)
Boisson and Romeo (1996)
Bowerman et al. (1994)
Braune eta. (1991)
Brezina and Arnold (1977)
Brugmann and Hennings
(1994)
Brugmann and Lange (1988)
Brumbaugh and Walther
(1991)
Burger(1992, 1994, 1995,
1996, 1997a,b)
Burger and Gochfeld
(1992a,b, 1993, 1995 ab,
1996,1997)
Burger et al. (1992a,b,c,1993,
1994a,b)
Byrne and DeLeon (1986)
Byrne etal. (1985)
Cantillo etal. (1997)
Capar and Yess (1996)
Capelli etal. (1987, 1991)
Cappon (1984)
Cappon and Smith (1981)
(1982a,b)
Cardellicchio(1995)
Carell etal. (1987)
F-7 Draft Document
-------
Carter and Porter (1997)
Caurantetal. (1994, 1996)
Chau and Riley (1965)
Chiang etal. (1994)
Chou and Uthe (1991)
Chvojka(1988)
Chvojkaetal. (1990)
Clifford and Harrison (1988)
Collins (1992)
Combs etal. (1996)
Cossonetal. (1988)
Courtney etal. (1994)
Cruwys etal. (1994)
Crutchfield (2000)
Cumbie and Van Horn
(1978)
Currey etal. (1992)
Custer and Hohman (1994)
Custer and Mitchell (1991,
1993)
Custer etal. (1997)
Dabeka and McKenzie
(1991)
Davoren(1986)
Deaker and Maher (1997)
Demon etal. (1988)
Dietz etal. (1995, 1996)
Dohertyetal. (1993)
Elliott and Scheuhammer
(1997)
Eriksson etal. (1989)
Evans etal. (1993)
Feltonrtal. (1990)
Feltonetal. (1994)
Fitzsimons et al. (1995)
Focardi etal. (1985, 1988)
Fowler (1986)
Fowler etal. (1975, 1985)
France(1987)
Friberg (1988)
Froslie etal. (1985, 1987)
Gabrashanske and Daskalova
(1985)
Gabrashanska and Nedeva
(1994)
Galgan and Frank (1995)
Garcia - Hernandez et al.
(2000)
Giardinaetal. (1997)
Gillespie and Baumann
(1986)
Gochfeld (1997)
Goede (1985, 1991, 1993a,b)
Goede etal. (1989, 1993)
Goede and DeBruin (1984,
1985)
Goede and Wolterbeek
(1993, 1994a,b)
Gras etal. (1992)
Greig and Jones (1976)
Gutenmann et al. (1988)
Gutierrez-Galindo et al.
(1994)
Guven etal. (1992)
Halbrooketal. (1996)
Hall and Fisher (1985)
Hamilton and Waddell
(1994)
Hamilton and Wiedmeyer
(1990)
Hansenetal. (1990)
Hardiman and Pearson
(1995)
Hargrave etal. (1992)
Harrison and Klaverkamp
(1990)
Hasunuma et al. (1993)
Haynes etal. (1995)
Hein etal. (1994)
Heiny and Tate (1997)
Heinz (1993a)
Heinz and Fitzgerald
(1993a,b)
Heit(1985)
Heit and Klusek (1985)
Heitetal. (1980, 1989)
Hellouetal. (1992a,b)
(1996a,b)
Henny and Herron (1989)
Hodge etal. (1996)
Hilton etal. (1982)
Honda etal. (1986)
Hothem and Ohlendorf
(1989)
Hothem and Welsh (1994b)
Hothem and Zador( 1995)
Hothem etal. (1995)
Houptetal. (1988)
Hunter etal. (1995, 1997)
Ibrahim and Farrag (1992)
Ibrahim and Mat (1995)
Ishikawaetal. (1993)
F-8 Draft Document
-------
Itanoetal. (1984, 1985a,b)
Jarmanetal. (1996)
Johns etal. (1988)
Johnson (1987)
Jop etal. (1997)
Jorhem etal. (1994)
Julshamn etal. (1987)
Kaietal. (1986a,b, 1988,
1992a,b, 1996)
Kaiser etal. (1979)
Kalas etal. (1995)
Kidwell etal. (1995)
Koeman etal. (1973)
Kovacs etal. (1984)
Krogh and Scanes(1997)
Krushevska et al. (1996)
Lakshmanan and Stephen
(1994)
Lalithaetal. (1994)
LamLeung et al. (1991)
Lanetal. (1994a,b)
Langlois and Langis (1995)
Larsen and Stuerup (1994)
Larsen etal. (1997)
Lauchli (1993)
Law etal. (1996)
Lee and Fisher (1992a,b,
1993)
Leighton and Wobeser
(1994)
Leland and Scudder (1990)
Lemly (1985a, 1994)
Leonzio etal. (1986, 1989,
1992)
Leskinen etal. (1986)
Li etal. (1996)
Lie etal. (1994)
Liu etal. (1987)
Lizamaetal. (1989)
Lobel etal. (1989, 1991,
1992a,b)
Lonzarich etal. (1992)
Lourdes etal. (1990)
Lowe etal. (1985)
Lucas etal. (1970)
Lytle and Lytle (1982)
Mackey etal. (1996)
Maher(1987)
Maheretal. (1992, 1997)
Mann etal. (1988)
Mason et al. (2000)
Masuzawa et al. (1988)
Matsumoto (1991)
Mavenetal. (1995)
May and McKinney (1981)
Mcdowell etal. (1995)
McKenzie-Parnell et al.
(1988)
Meadoretal. (1993)
Mehrle etal. (1982)
Meltzeretal. (1993)
Metcalfe-Smith et al. (1992,
1996)
Michotetal. (1994)
Mills etal. (1993)
Moharram et al. (1987)
Moller(1996)
Mora and Anderson (1995)
Moreraetal. (1997)
Muiretal. (1988)
Mutanenetal. (1986)
Nadkarni and Primack (1993)
Nakamoto and Hassler
(1992)
Narasaki and Cao (1996)
Navarrete etal. (1990)
Nettletonetal. (1990)
Nicola etal. (1987)
Nielsen and Dietz (1990)
Norheim(1987)
Norheim etal. (1992)
Norrgren etal. (1993)
Norstrom etal. (1986)
O\iConner(1996)
O^Sheaetal. (1984)
Oberetal. (1987)
Oehlenschlager(1997)
Ohlendorf(1986)
Ohlendorf and Harrison
(1986)
Ohlendorf and Marois (1990)
Ohlendorf et al. (1986a,b,
1987, 1988a,b)
Okazaki and Panietz (1981)
Ostapczuketal. (1997)
Pakkalaetal. (1972)
Pal et al. (1997) Palawski et
al. (1991)
Palmer-Locarnini and Presley
(1995)
F-9 Draft Document
-------
Paludan-Miller et al. (1993)
Papadopoulou and Andreotis
(1985)
Park and Presley (1997)
Park etal. (1994)
Paveglio et al. (1994)
Payer and Runkel (1978)
Payer etal. (1976)
Pennington et al. (1982)
Presley etal. (1990)
Quevauviller et al. (1993a,b)
Ramos etal. (1992)
Rao etal. (1996)
Reinfelder and Fisher (1991)
Reinfelder et al. (1993, 1998)
Renzonietal. (1986)
Rigetetal. (1996)
Risenhoover(1989)
Roditi (2000)
Rouxetal. (1994)
Ruelle and Keenlyne (1993)
Sager and Cofield (1984)
Saiki (1986 ab, 1987, 1990)
Saiki and Lowe (1987)
Saiki and May (1988)
Saiki and Palawski (1990)
Saiki etal. (1992, 1993)
Sanders and Gilmour (1994)
Scanes(1997)
Scheuhammer et al. (1988)
Schantzetal. (1997)
Schmitt and Brumbaugh
(1990)
Schramel and Xu( 1991)
Schuleretal. (1990)
Scott and Latshaw (1993)
Secoretal. (1993)
Seelye etal. (1982)
Sharif etal. (1993)
Shen etal. (1997)
Shirasaki etal. (1996)
Shultz and Ito (1979)
Simopoulos (1997)
Skaare etal. (1990, 1994)
Smith and Flegal (1989)
Smith etal. (1992)
Sorensen (1988)
Sorensen and Bauer
(1984a,b) Sorensen and
Bjerregaard(1991)
Sorensen etal. (1982, 1983,
1984)
Southworth et al. (2000)
Sparling and Lowe (1996)
Speyer(1980)
Steimle etal. (1994)
Stoeppleretal. (1988)
Stone etal. (1988)
Stripp etal. (1990)
Sundarrao etal. (1991)
(1992)
Svensson et al. (1992)
Tabakaetal. (1996)
Talbot and Chang (1987)
Tallandini etal. (1996)
Tan and Marshall (1997)
Tang etal. (1997)
Tao etal. (1993)
Teherani(1987)
Teigen etal. (1993)
Thomas etal. (1999)
Tilbury etal. (1997)
Topcuogluetal. (1990)
TranVan and Teherani (1988)
Trocine and Trefry (1996)
Uthe and Bigh (1971)
Vanderstoep et al. (1990)
Varanasi etal. (1993, 1994)
Vitaliano and Zdanowicz
(1992)
Vlieg(1990)
Vlieg etal. (1993)
Vos etal. (1986)
Waddell and May (1995)
Wagemann (1988)
Wagemann and Stewart
(1994)
Wagemann etal. (1988)
(1996) Walsh etal. (1977)
Wang (1996)
Ward and Flick (1990)
Warren etal. (1990)
Weber (1985)
Welsh and Maughan (1994)
Wen etal. (1997)
Wenzel and Gabrielsen
(1995)
Whyte and Boutillier (1991)
Williams etal. (1994)
Wilson etal. (1992, 1997)
F-10 Draft Document
-------
Winger and Andreasen Wu and Huang (1991) Zatta et al. (1985)
(1985) Yamaoka et al. (1996) Zeisler et al. (1988, 1993)
Winger et al. (1984, 1990) Yamazaki et al. (1996) Zhou and Liu (1997)
Woock and Summers (1984) Yoshida and Yasumoto
Wren etal. (1987) (1987)
F-11 Draft Document
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APPENDIX G: Supplementary information on Selenium
Bioaccumulation in Aquatic Animals
G-l Draft Document
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1.0 Effects of Growth Rate on the Accumulation of Selenium in
Fish
EPA analyzed the effect of the growth rate parameter g when estimating selenium bioaccumulation using
the mechanistic bioaccumulation modeling described in Equation 1 of the main text. Because the addition
of tissue associated with growth could have a dilution effect on the chemicals present in tissue, a
parameter representing growth rate is present in the denominator of the Equation 1. Indeed, growth can be
an important factor in the bioaccumulation of very hydrophobic chemicals with low excretion rates such
as polychlorinated biphenyls, (Connolly and Pedersen, 1988). However, the effect of growth may not be
as important for selenium because of its unique biogeochemical characteristics, route of exposure, and
role as a micronutrient.
EPA tested the effect of the growth rate parameter g on estimates of selenium bioaccumulation using
Equation 1 with different food web scenarios. Increasing growth rates from 0 (no growth) to 0.2/day (a
relatively high rate of growth) reduced selenium concentrations in trophic level 2 and 3 organisms by as
much as a factor of 10 to 20. Thus incorporating growth rate in Equation 1 could result in significant
dilution of selenium and lower estimates of selenium bioaccumulation.
Although increasing the value of the growth parameter g in Equation 1 reduces estimates of selenium
bioaccumulation, this simple analysis neglects an important physiological linkage between growth and
food consumption. Organisms must consume enough food to support growth and meet their energy
requirements for respiration, specific dynamic action, waste loss, and reproduction. These physiological
requirements suggest that higher growth rates are associated with greater rates of food consumption.
Because food consumption is the primary route of selenium exposure in aquatic organisms, increased
selenium exposure associated with higher food consumption could counterbalance the dilution of
selenium in tissue associated with higher growth rates.
EPA tested the effects of growth on estimates of selenium bioaccumulation using Equation 1 when
increased food consumption was associated with higher growth rates. EPA modified Equation 1 to
incorporate a simple relationship for bioenergetics (Thomann et al. 1992) and applied the model to
reexamine the sensitivity of selenium bioaccumulation to growth rates in trophic level 2 and 3 organisms.
The results of this analysis showed that increasing growth rates over two orders of magnitude increased
selenium concentrations in trophic level 2 by a factor of 2, and decreased selenium concentrations in
G-2 Draft Document
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trophic level 3 by 10%. When growth rates were increased simultaneously in trophic levels 2 and 3, the
selenium concentrations increased by less than a factor of 2. This analysis suggests that when
bioenergetics is considered, selenium bioaccumulation is generally insensitive to organism growth rates.
EPA believes that uncertainties in the toxicokinetic parameters of selenium far outweigh the effects on
growth rate on selenium bioaccumulation. Thus, the growth rate parameter g was removed from Equation
1 for the purpose of deriving a translation equation that could be used to implement a tissue-based
selenium water quality criterion.
2.0 Analysis of the Relative Contribution of Aqueous and
Dietary Uptake on the Bioaccumulation of Selenium
EPA analyzed the relative contributions of direct aqueous uptake versus ingestion of selenium in
consideration of removing the uptake rate constant ku from Equation 1. Because an important exposure
route for some chemicals is direct contact with water, an uptake rate constant ku is present in the
numerator of Equation 1. However, fish and invertebrate organisms absorb selenium primarily through
the consumption of food rather than from direct aqueous uptake (Forester 2007; Lemly 1985; Luoma et
al. 1992). Thus, removing the uptake rate constant ku could simplify Equation 1 while maintaining the key
determinants of selenium bioaccumulation.
EPA tested the relative contribution of aqueous versus dietary uptake of selenium using a version of
Equation 1 that incorporates both exposure pathways (Thomann et. al. 1992). For trophic level 2,
selenium bioaccumulation was estimated for a range of uptake rates that varied according to the
respiration rate and aqueous transfer efficiency of selenium relative to dissolved oxygen. For trophic level
3, uptake rates were varied within a range of values reported in Besser et al. (1993) and Bertram and
Brooks (1986).
EPA's analysis showed that diet accounted for 34% - 92% of selenium bioaccumulation at trophic level 2,
with a median of 74%. At trophic level 3, diet accounted for 62% - 100% of tissue selenium, with a
median of 95%. Thus, disregarding aqueous uptake of selenium only resulted in a small (~5%) reduction
in estimated selenium bioaccumulation in trophic level 3 organisms. These results are consistent with
previous studies indicating that diet is the primary exposure route of selenium, and suggests that the
uptake rate constant for selenium can be removed from Equation 1 with negligible effect for higher
trophic levels organisms.
G-3 Draft Document
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3.0 Kinetics of Accumulation and Depuration: Averaging Period
3.1 Background
For setting averaging periods for aquatic life criteria, U.S. EPA (1995b) used the concept that the criterion
averaging period should be less than or equal to the "characteristic time" describing the toxic speed of
action. In the context of the water-borne direct toxicity of metals, characteristic time = 1/k, where k is the
first-order kinetic coefficient in a toxico-kinetic model fitted to the relationship between LC50 and
exposure duration.
In the context of selenium bioaccumulation in a single trophic level, k would the first-order depuration
coefficient, and 1/k would equal the time needed to depurate to a concentration of 1/e times the initial
concentration (where e=2.718). For depuration of multiple trophic levels sequentially, the characteristic
time is likewise the time needed for c/c0 reach a value of 1/e, as shown in Figure G-la. The accumulation
curve is the inverted depuration curve, as shown in Figure G-lb.
1
>
•
1
,
\
V
—
*
•^H
\
\
\
V
1/p
1/C
\
^^^•1
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~ 0 — i —
0 20 40
Time,
Fish
inv*
Fish
Invt
s
^N
(
on
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on
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dep
).
clea
). or
urat
n di
i cle
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ing
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an i
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iet
60 80 100
days
_3
re
~i n Q
ativeto Plateai
S 0 <
i Oi C
•5 "
B
u
o 0.2 -
u
01
1
1
*
r /
; /
.
0
/
..* '*
„•
J Fishc
X
teb.
m c<
OIK
>nta
S*
out
111. (
-.».
am
liet
diet
^^— Fisb on accumulating
inverteb.
0 20 40 60 80 100
Time, days
Figures G-l a & b. Depuration and accumulation behavior for invertebrate k=0.1/day and fish
k=0.02/day
In the Figures G-l a & b examples, the characteristic time for invertebrates on an invariant diet is 10 days,
the characteristic time for fish on an invariant diet is 50 days, and the characteristic time for fish on an
invertebrate diet that is itself depurating or accumulating is the approximate sum of the individual
characteristic times, or ~60 days.
G-4 Draft Document
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In contrast to the model depuration rate, k, the model uptake rate (AE, assimilation efficiency, multiplied
by IR, intake rate) does not affect the characteristic response time. Rather it affects the magnitude of the
accumulation plateau. Uptake rate thus affects the TTF value itself but is not relevant to setting an
averaging period.
Because short averaging periods are more environmentally conservative than long averaging periods,
selecting parameter values for fast kinetics is more environmentally conservative. Figure G-l reflects
environmentally conservative choices for k values.
3.2 Approach for Modeling Effects of Time- Variable Se Con centrations
Expression of concentrations. None of the concentrations in this analysis are expressed in ordinary units
of concentration. All concentrations are modeled as values normalized to their allowable benchmark
concentration - that is, concentration = 1 for a particular medium (water, algae-detritus-sediment,
invertebrates, or fish) means that the medium is at its criterion concentration or corresponding benchmark.
It is assumed that the benchmarks correctly align - water held at its benchmark concentration will
ultimately yield Trophic Levels 1, 2, and 3 at their respective benchmark concentrations. The Trophic
Level 3 benchmark is the reproductive EC10 for the 5th percentile taxon: i.e., the fish tissue criterion.
Formulation of the bioaccumulation model for kinetic analysis. Lacking any k values for Trophic Level 1
(algae-detritus-sediment), this analysis makes the worst-case assumption that these respond
instantaneously to water concentrations. Thus, at any time t, after normalization to their respective
benchmarks:
Algae-detritus-sediment: CTL1[t] = C[t]water
The above is not saying that in ordinary units of concentration, the Trophic Level 1 equals water, but only
when normalized to their respective benchmarks. In ordinary (non-normalized) units of concentration,
TL2 would equal TL1 x ER, the enrichment function.
For invertebrates and fish, accumulation at time t equals accumulation at time t-1 plus intake minus
depuration, as follows:
Invertebrates:
CTL2[t] = CTL2[t-l] + AETL2IRTL2 CTL1[t-l] - kTL2 CTL2[t-l]
G-5 Draft Document
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Fish:
CTL3[t] = CTL3[t-l] + AETL3IRTL3 CTL2[t-l] - kTL3 CTL3[t-l]
For invertebrates, values for kTL2 are tabulated in elsewhere in the document. It was assigned here a value
of 0. I/day, considerably higher than those for Lumbriculus, Asian clam, zebra mussel, but a bit lower
than that for copepods, which are very small in size. As previously mentioned, higher k (more rapid
kinetics) is an environmentally conservative assumption in this context.
For fish, the median depuration coefficient measured by Bertram and Brooks (1986) for 6-9 month-old
(early adult) fathead minnows was used, providing a kTL3 value of 0.02/day. Because of the small size of
adults of this species, this represents faster kinetics than would likely be applicable the salmonids and
centrarchids of greatest concern for selenium toxicity. The striped bass k value of Baines et al. (2002) is
inapplicable here because it was measured in the early juvenile life stage, a size that is too small to be
relevant to reproductive impairment stemming from exposure of adult females. The concentration in fish
could be equivalently viewed as either whole body or egg-ovary, relative to their respective benchmarks.
That is, partitioning within body of the fish is assumed not to involve a time delay.
The value of a TTF is given by AE x IR/k. Concentrations in TL1, TL2, and TL3 are normalized to their
benchmarks, meaning that all benchmark concentrations have a value of 1.0. In this normalized context,
the TTFs must also equal 1.0, since upon reaching steady state, TL1 at its benchmark will yield TL2 at its
benchmark, which in turn will yield TL3 at its benchmark. Again, the analysis is not intended to reflect
actual concentrations, merely portray temporal behavior. Since 1 = TTF = AE x IR/k, it follows that AE x
IR = k within this normalized framework. Although only the product AE x IR is relevant, they are
retained as distinct parameters to maintain parallelism with remainder of the criterion document. AE was
assigned a value of 0.5 for fish and invertebrates, and IR = k/AE in the normalized framework.
Time step durations of 0.1-1.0 day were considered. Short time steps increase accuracy by decreasing the
numerical dispersion inherent in expressing C[t] = f(C[t-l]). A time step of 0.5 day was found to yield
sufficient accuracy, as measured by predicted values at the characteristic time for depuration or
accumulation (per Figure G-l).
Prediction of Effects. The effect level associated with the tissue concentration at any time t is calculated
via the log probit concentration-response curve, one of the commonly used sigmoid curves. It assumes
G-6 Draft Document
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that the sensitivities in the underlying population are log-normally distributed such that the concentration
yielding effects on k percentage of the population is given by:
ECk = EC50 exp(o z)
where o is the inverse of the concentration-response curve slope and z is the normal deviate
corresponding to k percent (e.g., for k=10%, z=NORMSINV(0.!)=-!.28155). Among the reproductive
impairment studies presented in Appendix G, an approximate median ratio for EC50/EC10 is 1.5. This
translates to o=0.3164.
Since the fish tissue criterion concentration equals 1.0 in this normalized framework, at any time t, the
fractional level of effect corresponding to any value of CTL3 is given by:
Fractional Effect [t] = NORMSDIST(z[t])
where z[t] is given by:
zftj =LN(CTL3[t]/1.5)/0.3164
Exposure Scenarios. A range of exposure scenarios were evaluated under which the criterion was attained
while maximizing exposure and effects. These begin with absolute worst case scenarios, followed by
more realistic situations. The following represent worst case situations because the 30-day average water
concentration remains continuously at the criterion concentration at all times.
1. Steady concentrations at the criterion: this is worst-case continuous exposure.
2. Uniform 1-day spikes at SOX the water criterion concentration, occurring at uniform 30-day
intervals (i.e., separated by 29 days of zero concentration) such that the 30-day average always
equals the criterion. This is the worst-case intermittent scenario, attaining the criterion through a
time series that continually maximizes the 30-day average exposure at the water criterion
concentration while also imposing the highest variability possible from spikes of 1-day duration.
Because they lack real-world random variability, the above two scenarios are not realistic, but are
used as absolute worst cases for purposes of comparison. The following two scenarios represent
somewhat more realistic possibilities for intermittent releases.
G-7 Draft Document
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3. Normally distributed 1-day spikes having CV=0.12, again occurring at uniform 30-day intervals
(separated by 29 days of zero concentration). Because a normal distribution has no upper bound,
some percentage of 30-day average exceedances must be provided for. For this scenario, 5% of
the 1-day spikes exceeded 30X the 30-day average water criterion concentration.2
4. Log-normally distributed 1-day spikes, with log standard deviation = 0.20, again occurring at
uniform 30-day intervals. As with the previous scenario, 5% of the 1-day spikes exceeded 3 OX
the 30-day average water criterion concentration.
Finally, the following represents a realistic and indeed typical situation for continuous exposure:
5. Log-normally distributed, smoothly variable concentrations, continuous rather than intermittent
exposure, with the 30-day average exceeding the criterion once in three years when counted using
the procedure of EPA (1986). The log standard deviation of 0.5 applied here represents typical
real-world time variability for continuously flowing waters. The log serial correlation coefficient
p = 0.8 represents that typical of smaller streams (Delos 2008).
With respect to maximizing toxic effects while attaining the criterion, Scenarios #1 and #2 are absolute
worst cases. Scenarios #3 and #4 are somewhat more reasonable worst cases. While relatively low
variability of concentrations occurring in intermittent releases might correspond to some real-world
situations, the uniformity of spacing between spikes is unrealistic. That uniformity of spacing allows
maximizing exposure while attaining the criterion a high percentage of the time. Were the spacing not
uniform, exposures concentrations would have to be substantially less in order to continue to attain the
30-day average water criterion concentration. In contrast to the others, Scenario #5 represents typical time
variability in ambient waters.
Scenarios 3, 4, and 5 require randomly generated concentrations (having specified target statistical
characteristics). Multiple runs of long series are therefore needed to assure some reasonable degree of
accuracy. A minimum of 20 runs of random series of 3000 days were used. In Scenario 5, the
concentrations at each half-day time step were generated by the following formula:
2 Concentrations of selenium in contaminated groundwater tend to be rather stable. Dilution by rainfall events may punctuate
these rather stable elevated concentrations with inverted spikes of low concentrations. The 2008-2012 selenium data reported by
FTN Associates, Ltd., in "Evaluation of Selenium Concentrations in Fish Collected in the Receiving Stream for Mueller Copper
Tube Products, Inc., Wynne, Arkansas" dated September 17,2012, showed that the upper half of the concentration distribution
tended to fit a normal distribution having the low variability represented by CV=0.12. In contrast to situation assessed by FTN
Associates, this scenario envisions intermittent spike releases at uniform intervals where the spike concentrations have this low
variability.
G-8 Draft Document
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Cftjwater = Cft-lJwaterA(p') * GMA(l-p') * EXP{a * SQRT(l-p 'A2) *NORMSINV(RAND)
where p' (rho prime) is the desired serial correlation coefficient between half-day time steps: p'=SQRT(p)
[approximation], where p (rho) is the desired serial correlation coefficient between daily values; GM is
the desired geometric mean or median, and o is the desired log standard deviation (Delos 2008). The
above formula allows a time series with the desired statistical characteristics to be generated.
3.2.1 Model Results
3.2.1.1 Steady concentrations at the water criterion concentration.
No graphic is needed to explain this scenario. With water steady at its criterion, algae-detritus-sediment
and invertebrates are likewise steady at their benchmark concentrations, and fish tissue is at its criterion
concentration. For the 5th percentile taxon, the effect would thus be 10% since the concentration is steady
attheEClO.
Figure G2. Scenario 2, uniform 1-day spikes at 3 OX the water criterion concentration, occurring at
uniform 30-day intervals such that the 30-day average always equals the criterion. Read invertebrate and
fish tissue concentrations on left scale, water concentrations on right scale. Time=0 does not represent the
beginning of exposure; prior to Time=0 the same exposure pattern had been going on for a long time
(e.g., 10,000 days).
G-9 Draft Document
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•E
(V
£
fish tissue
inverteb. tissue
water 1-cl spike
water 30-d avg
- 30
2.5
w
ffi
B
o
0.5
10 20 30 40 50 60 70 80
Time after Achieving Dynamic Steady State, days
90
Figure G2. Uniform 1-day spikes at 30X the water criterion concentration, occurring at uniform
30-day intervals such that the 30-day average always equals the criterion.
With their more rapid kinetics, invertebrate tissue concentration swings are much more drastic than fish
tissue concentration swings, but were the spike to continue as a steady exposure 30-fold above the water
benchmark, both invertebrate tissue and fish tissue would ultimately plateau at 30-fold above their
respective benchmarks.
The key point here is that attaining the 30-day average via 1-day spikes spaced 30 days apart generates a
small oscillation in fish tissue concentrations. Averaged over the 30-days, the fish tissue concentrations
exactly attain their criterion. However, the nonlinearity of the log-probit concentration-response curve
means that the reduction in toxicity while below the criterion does not quite balance the increase in
toxicity while above the criterion. The average effect ends up being 10.3% instead of 10.0%.
G-10 Draft Document
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3.2.1.2 Normally distributed 1-day spikes having CV=0.12, uniformly separated by 29 days at zero
concentration.
This scenario simulates the low variability of elevated selenium concentrations sometimes observed in
releases of groundwater selenium, but discharged here in uniformly spaced spikes.
1.2
water 1-d spike
water 30-d avg
500 1000 1500 2000 2500
Time after Achieving Dynamic Steady State, days
3000
Figure G3. A typical random series for Scenario 3, normally distributed 1-day spikes having
CV=0.12, uniformly separated by 29 days at zero concentration. Again read fish tissue concentrations
from left scale, water concentrations from right scale. Note: (a) the time scale is 3000 days (100 sets of 30
days), compared to 90 days in Figure G-2, and (b) the left axis is scaled differently than Figure G-2
because the wide concentrations swings of invertebrate tissue are not presented here. Note also that on 29
days of each 30 day period the water concentration is zero - because of pixel limitations, the spike line
thickness on the graph overstates the spike duration relative to the intervening white space. And note that
Time=0 does not represent the beginning of exposure; prior to Time=0 the intermittent spikes had been at
their median concentration for thousands of days.
In the Figure G-3 example run, 5 of the 100 spikes yield exceedance of the criterion, possibly
representing a detection frequency resolution limit in an ambitious real-world monitoring program. On
G-11 Draft Document
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0.78% of the days, the fish tissue criterion exceeded its criterion, and the aggregate effect (considering all
days, whether exceeding or not) was 3.73%. The effect in this somewhat more realistic scenario is less
than in the previous absolute worst case because the average spike concentration must necessarily be less
here in order to prevent the variable spikes from causing exceedance of the 30-day average water target.
3.2.1.3 Log-normally distributed 1-day spikes, with log standard deviation = 0.20, uniformly separated
by 29 days at zero concentration.
Although still representing low variability when comparing among spike concentrations, the spikes in this
scenario have greater and more typically skewed variability than the previous scenario.
40
- 35
c
_g
'w
v
?-0
0.2
25
fish tissue
water 1-d spike
water30-d avg
u
u
'E
o
j=
u
i/)
20 O
Oi
>
15 •=
10
5
ro
Qj
CC
u
c
o
u
ro
500 1000 1500 2000 2500
Time after Achieving Dynamic Steady State, days
3000
Figure G4. A typical random series for Scenario 4, log-normally distributed 1-day spikes, with log
standard deviation = 0.20, uniformly separated by 29 days at zero concentration. This figure is
structured like Figure G-3, and again exposure had long preceded Time=0.
In the Figure G-4 example, 5 of the 100 the spikes cause exceedance of the criterion, but did not cause
any exceedance of the fish tissue criterion. The aggregate effect was 1.64%. Because the exposure is more
variable than in the previous scenario, the average exposure (and aggregate effect) must necessarily be
less in order to prevent exceedance of the 30-day average water target.
G-12 Draft Document
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3.2.1.4 Log-normally distributed, smoothly variable concentrations, continuous rather than intermittent
exposure
This is the most realistic of the scenarios, corresponding to typical variability observed in streams.
2.5
J£
1_
ro
c
Hi
CO
0
4-»
IP
fish tissue
water daily
water 30-d avg
> 1.5
OJ
K
u
5
ro
0.5
'i I ilplHf™
500 1000 1500 2000 2500
Time after Achieving Median Steady State, days
3000
Figure G-5. A typical example of log-normally distributed, smoothly variable concentrations,
continuous rather than intermittent exposure. The standard deviation of natural logs is 0.5 and the
serial correlation coefficient of logs is 0.8 for daily values, both typical real-world situations. (The
compression of 3000 days into the graph might make it difficult to recognize that the time series is
smoothly varying - it has serial correlation.) Prior to Time=0, exposure had been at the median
concentration for a long period of time.
In the Figure G5 example run, instantaneous water concentrations exceed the 30-day average criterion
7.5% of the time. The 30-day average concentrations exceed the criterion 1.15 times per 3 year period,
counted per the EPA (1986) counting method. Tissue concentrations do not exceed their criterion at any
time, and the aggregate effect is 0.17%.
G-13 Draft Document
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In contrast to the previous scenarios, the elevated concentrations here are random in their spacing and
duration. This additional randomness reduces the average exposure (and aggregate effect) compatible
with attainment of the 30-day average water target.
3.2.2 Summary of Scenario Results
Because Scenarios 3, 4, and 5 involve generation of random concentrations, the above graphs show just
one run (3000 days) for each. Full results for the 20 runs for each scenario are shown below.
Scenario
1. Steady
2. Uniform
spikes
3. Norm.
spikes
4. Lognorm
spikes
5. Smooth
variable
Water:
# 30-day avg.
exceedances /
3-vr1
0.00
0.00
3.58
3.59
0.99
Water:
% of time
exceeding
0.00
3.33
5.00
5.00
7.45
Tissue:
% of time
exceeding
0.00
56.7
0.59
0.13
0.00
Mean
effect
for 5th
%ile
Taxon
10.0
10.3
3.60
1.57
0.18
Comment
Steady at water and tissue
benchmarks
30-d avg water cone, remains
steady at benchmark (Fig. 2)
Spike median=25 . 1 x
benchmark, spike CV=0.12
(e.g., Fig. 3) 2
Spike median=21.6 x
benchmark, spike log stdev=0.2
(e.g., Fig. 4) 3
Median=0.49 x benchmark, log
stdev=0.5, rho(daily)=0.8 (e.g.,
Fig. 5) 4
1. Counting procedure for 30-d avg. exceedances is that of U.S. EPA (1986).
2. Results for Scenario 3 are average of 20 runs of 3000 days, each run with exactly 5% exceedances. Runs not
yielding 5% exceedances were not used. Among the 20 runs with 5% exceedances, the effect CV=0.05
(coefficient of variation).
3 . Results for Scenario 4 are average of 20 runs of 3000 days, each run with exactly 5% exceedances. Runs not
yielding 5% exceedances were not used. Among the 20 runs with 5% exceedances, the effect CV=0. 1 1 .
4. Results for Scenario 5 are average of 20 runs of 3000 days, each run with 0.6-1. 4 exceedances / 3 yr. Runs not
yielding exceedances within these bounds were not used. Among the 20 runs used, the effect CV=0.28.
3.2.3 Conclusion
The kinetics of selenium accumulation and depuration are sufficiently slow that applying a 30-day
averaging period to the water criterion concentration affords protection even under unrealistic worst case
conditions.
G-14 Draft Document
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APPENDIX H: Binary Classification Statistics
H-l Draft Document
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Sensitivity, Specificity, and Related Statistics in Applying Water-
Column Criteria
The ability of any water criterion to predict exceedance of the egg-ovary FCV can be evaluated in terms
of sensitivity, specificity, and related statistical measures of a binary classification test. In this case, each
species-site combination in our data set of aquatic sites can be classified as (a) either above or below the
egg-ovary criterion and (b) either above or below the water criterion concentration. Although such a
binary classification scheme does not consider the degree to which measurements are above or below a
criterion, water quality standards are usually implemented as a binary decision (a water body either attains
or exceeds criteria). Thus, a statistical analysis using binary classification can provide valuable
information about how well the recommended selenium water-column criteria values ensure attainment of
the egg-ovary FCV in a regulatory context.
Separate binary classification tables for the 49 lotic species-site combinations and 83 lentic species-site
combinations were generated using the following format:
Tissue concentration
greater than tissue
criterion
Tissue concentration less
than tissue criterion
Water concentration greater than
water criterion
True Positive (TP)
False Positive (FP)
Water concentration less than
water criterion
False Negative (FN)
True Negative (77V)
Below are Tables 9a and 9b reproduced from the main text showing the frequency of binary
classifications for lentic and lotic waters in the confirmation dataset:
Lentic waters
Water concentration greater than
water criterion
Water concentration less than
water criterion
Tissue concentration
greater than tissue
92
14
Tissue concentration less
than tissue criterion
13
21
H-2 Draft Document
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Lotic waters
Water concentration greater than
water criterion
Water concentration less than
water criterion
Tissue concentration
greater than tissue
248
52
Tissue concentration less
than tissue criterion
206
182
From the counts in the binary classification tables, the sensitivity, specificity, and related statistics
(Fawcett 2006, Lowry 2011) are given as:
• Sensitivity or True Positive Rate (TPR) = TP I (TP + FN)
• Specificity or True Negative Rate (TNR) = TNI (FP + TN)
• Positive Predictive Value (PPV) = TP I (TP + FP))
• Negative Predictive Value (NPV) = 77V / (TN + FN))
Sensitivity-specificity and PPV-NPV look at the problem from different perspectives. Sensitivity-
specificity indicate probabilities given tissue value exceedance or attainment; PPV-NPV indicate
probabilities given water value exceedance or attainment.
These statistics evaluate the performance of the water criterion values as predictors of attainment or
exceedance of the egg-ovary FCV. In this application, sensitivity addresses how well exceedance of the
water criterion detects exceedance of the tissue criterion. It is the ratio of the species-site combinations
exceeding both water and tissue criteria compared to the total number actually exceeding the tissue
criterion. Thus, given actual exceedance as determined by tissue measurement, sensitivity indicates the
probability that the water measurement will correctly indicate such exceedance. Conversely, specificity
addresses how well attainment of the water criterion indicates attainment of the tissue criterion. It is the
ratio of the number of species-site combinations attaining both water and tissue criteria compared to the
total number attaining the tissue criterion. Thus, given actual attainment as determined by tissue
measurements, specificity indicates the probability that the water measurement will correctly indicate
such attainment. For both sensitivity and specificity, higher values (i.e., closer to 1.0) are better,
respectively indicating few false negatives or false positives. However, even if selenium bioaccumulation
(the relationship between tissue and water concentrations) were not variable across sites, such that water
concentrations have limitations for predicting selenium risks (as discussed for example by Chapman et al.
2009, 2010), statistical uncertainty in the tissue and water concentration measurements themselves would
H-3 Draft Document
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generally prevent perfect sensitivity and specificity. Consequently, establishing a reliable criterion
involves a tradeoff between sensitivity and specificity. Lowering the water criterion to increase its
sensitivity generally decreases its specificity (possibly making it over-protective). Raising the water
criterion to increase its specificity generally decreases its sensitivity (possibly making it under-
protective).
The positive predictive value (PPV) addresses how well water criterion exceedance (test positive) predicts
tissue criterion exceedance. That is, given a water criterion exceedance, the PPV indicates the probability
that the tissue criterion is also exceeded. It is the ratio of the number of species-site combinations
exceeding both water and tissue criteria compared to the total number actually exceeding the water
criterion (i.e. the proportion of true positives out of all positive results). Conversely, the negative
predictive value (NPV) addresses how well water criterion attainment (test negative) predicts tissue
criterion attainment. That is, given water criterion attainment, the PPV indicates the probability that the
tissue criterion is also attained. It is the ratio of the number of species-site combinations attaining both
water and tissue criteria compared to the total number attaining the water criterion (i.e. the proportion of
true negatives out of all negative results). Again, PPV and NPV values closer to 1.0 are better; however,
these statistical measures are also subject to the same tradeoffs as described for selectivity and sensitivity.
Thus, the limitations of the statistical parameters presented here should be recognized when evaluating
the performance of the water criterion concentration values, and no single parameter should be relied
upon at the exclusion of the others.
Sensitivity and specificity are inherent to the firmness of relationship between water and tissue
concentrations and to the particular water criterion concentration chosen. PPV and NPV likewise depend
on that, but they also depend on the prevalence of tissue concentrations exceeding the criterion in the
study population. When the binary statistics are taken from a relatively high risk population (such as from
studies dominated by sites having elevated selenium inputs), the following alternative forms of the
equations are useful for calculating PPV and NPV for a lower risk population (such as represented by fish
tissue concentrations observed in the EPA (2012) National Rivers and Streams Assessment, representing
in all U.S. flowing waters) having known prevalence, P, of exceeding the tissue criterion:
• PPV = Sens • P / ((Sens • P) + (1 - Spec)(l - P))
• NPV = Spec • (1 - P) / (Spec • (1 - P) + (1 - Sens) • P)
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Where Sens and Spec are the calculated sensitivity and specificity (from the data on the higher risk
population). Where prevalence is very low, as in the nationwide survey population, PPV is calculated to
be low and NPV high.
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APPENDIX I: Site Specific Criteria
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1.0 Translating the concentration of selenium in tissue to a
concentration in water
The EPA is recommending a selenium criterion to protect aquatic life that is expressed as a concentration
in the eggs or ovaries offish. Although the selenium concentration in eggs or ovaries is the most sensitive
and reliable basis for a criterion, implementation can be challenging because most state and tribal Clean
Water Act programs require the expression of water quality criteria as an ambient concentration in the
water-column. Therefore, the EPA is also recommending two water-column criterion values, one for lotic
or flowing waters, and the other for lentic or still waters.
The EPA derived the water-column criterion values by developing a translation equation based on
selenium bioaccumulation modeling. The EPA worked with the United States Geologial Survey to derive
a translation equation that utilizes a mechanistic model of bioaccumulation previously published in peer-
reviewed scientific literature (Luoma et. al., 1992; Wang et. al., 1996; Luoma and Fisher, 1997; Wang,
2001; Schlekat et al. 2002b; Luoma and Rainbow 2005; Presser and Luoma 2006; Presser and Luoma
2010; Presser 2013). The selenium egg-ovary FCV is translated to water concentration values at a set of
lentic and lotic aquatic systems, and the distribution of site-specific water concentrations from these sites
is used to derive water-column criterion values protective of aquatic life. This appendix describes how
states and tribes may use this methodology to translate the egg-ovary FCV into site-specific water-column
concentrations to more precisely manage selenium in specific aquatic systems. The use of a
Bioaccumulation Factor (BAF) approach is also briefly discussed. States and tribes may also derive site-
specific water concentration values using other scientifically defensible methods.
The relationship between the concentration of selenium in the eggs or ovaries offish and the
concentration of selenium in the water-column can vary from one aquatic system to another. The species
offish, the species and proportion of prey, and a variety of site-specific biogeochemical factors can
substantially affect selenium bioaccumulation and thus determine the allowable concentration of selenium
in surface waters that is protective of aquatic life. Because most state and tribal Clean Water Act
programs require the expression of water quality criteria as ambient concentrations in water,
implementation of the selenium criterion expressed as a concentration in the eggs or ovaries offish
requires the ability to translate the egg-ovary FCV into site-specific water-column concentrations. The
EPA considered two different modeling approaches to implement the selenium egg-ovary FCV:
mechanistic model of bioaccumulation, and a site-specific, field-derived bioaccumulation factor (BAF).
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The mechanistic modeling approach uses scientific knowledge of the physical and chemical processes
underlying bioaccumulation to establish a relationship between the concentrations of a chemical in the
water-column and the concentration of a metal in the tissue of aquatic organisms. The EPA used this
approach to develop a mathematical equation that allows states and tribes to formulate site-specific
models of trophic transfer of selenium through aquatic food webs and translate the egg-ovary FCV into an
equivalent site-specific water concentration.
The BAF approach is an empirical model of bioaccumulation based on the measured concentration of a
chemical found in the tissue of aquatic organisms and the water from where the aquatic organisms reside
(U.S. EPA 2001c). The concentration of the chemical is measured in both fish tissue and the water-
column, and a BAF is calculated by taking the ratio of the two concentrations. The BAF can then be used
to estimate the concentration of the chemical in one media when only the concentration of the chemical in
the other media is known.
Both the mechanistic modeling approach and the BAF approach have advantages and disadvantages that
states and tribes should consider before deciding which approach to use. On the one hand, the mechanistic
modeling approach has the advantage of not requiring extensive fish tissue sampling and analysis by
using knowledge of aquatic system food webs and relatively simple measurements of selenium from the
aquatic system that are easier and less expensive to obtain. However, uncertainty can be introduced if
inappropriate parameters are chosen to model selenium bioaccumulation in the aquatic ecosystem. On the
other hand, the BAF approach is conceptually and computationally simpler because it is completely
empirical, relying only on field measurements with no need for any knowledge of the physical, chemical,
or biological characteristics of the waterbody. However, the BAF approach requires multiple
measurements of selenium in fish tissue that may be unavailable or expensive to obtain.
The appropriate modeling approach to use when translating the selenium egg-ovary FCV to a site-specific
water-column concentration depends on individual circumstances. Although the mechanistic modeling
approach may be a cost-effective method in situations where there is little or no current information about
selenium bioaccumulation, the BAF approach may be desirable in circumstances where substantial
resources have already been invested in fish tissue sampling and analysis. Because the national egg-ovary
selenium criterion is intended to apply to all waters of the United States, and site-specific BAF values or
the data required to derive site-specific BAF values are not available for the vast majority of those waters,
the EPA developed and utilized a translation methodology based on the mechanistic modeling approach.
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Below is a description of how states and tribes may use this methodology to translate the egg-ovary FCV
to a site-specific water-column concentration for site-specific management of selenium.
1.1 Relating the Concentration of Selenium in Tissue and Water using the mechanistic modeling
approach
To relate the concentration of selenium in the eggs or ovaries of fish to the concentration of selenium in
the water-column, the EPA derived the equation:
C.
s~i _ egg-ovary
water rprpj-,Composite 7777,, /">77 _,
Lib ' xhb xC/< (Equation 9)
Where:
CWater = the concentration of selenium in water ((ig/L),
C egg-ovary = the concentration of selenium in the eggs or ovaries offish ((ig/g),
TTFomposUe = the product of the trophic transfer function (TTF) values of the fish species that
is the target of the egg-ovary FCV and the TTF values of all lower trophic
levels in its food web (no units of measurement, see explanation below).
EF = the steady state proportional bioconcentration of dissolved selenium at the base
of the aquatic food web (L/g),
CF = the species-specific proportion of selenium in eggs or ovaries relative to the
average concentration of selenium in all body tissues (no units of
measurement),
The basic principles expressed in Equation 9 are illustrated in the conceptual model shown in Figure 1-1.
Selenium dissolved in surface water enters aquatic food webs by becoming associated with trophic level 1
primary producer organisms (e.g. algae) and other biotic (e.g. detritus) and abiotic (e.g. sediment)
particulate material. An enrichment function (EF) quantifies the bioconcentration of selenium in
particulate material and thus its bioavailability in the aquatic system. The parameter EF in Equation 9 is a
single value that represents the steady state proportional concentration of selenium in particulate material
relative to the concentration of selenium dissolved in water.
Particulate material is consumed by trophic level 2 organisms (usually aquatic invertebrates) resulting in
the accumulation of selenium in the tissues of those organisms. Trophic level 2 invertebrates are
consumed by trophic level 3 fishes resulting in further accumulation of selenium in the tissues offish.
Bioaccumulation of selenium from one trophic level to the next is quantified by a trophic transfer function
(TTF). A TTF is a single value that represents the steady state proportional concentration of selenium in
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the tissue of an organism relative to the concentration of selenium in the food it consumes. Different
species of organisms metabolize selenium in different ways. Thus each species is associated with a
specific TTF value. Because the trophic transfer of selenium through all trophic levels is mathematically
equal to the product of the individual TTF values, all consumer-resource interactions in a particular
aquatic ecosystem are simplified in Equation 9 by representing the product of all the individual TTF
values as the single parameter 777^°"^°^
Fish accumulate selenium in different tissues of the body in differing amounts. Because the selenium
criterion is expressed as a concentration in the eggs and/or ovaries, a conversion factor (CF) quantifies the
relationship between the concentration of selenium in the eggs and/or ovaries and the average
concentration of selenium in the whole body. The parameter CF in Equation 9 is a single value that
represents the steady state proportional concentration of selenium in the eggs and/or ovaries relative to the
average concentration of selenium in all body tissues. Different species offish accumulate selenium in
their eggs and ovaries to different degrees. Thus each species offish is associated with a specific CF
value.
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Egg-Ovary FCV
Species Egg-Ovary to Whole-Body Conversion Factor (CF)
Fish Whole-Body
Concentration
Species Trophic Transfer Function (TTF)
Invertebrate
Concentration
Species Trophic Transfer Function (TTF)
(J'J'pcomposite\
Concentration in
Particulate Material
Enrichment Factor (EF)
Water-Column
Concentration
Figure 1-1. Conceptual model for translating the egg-ovary FCV to a water-column concentration.
Once the parameters that quantify the transfer of selenium through each step in this pathway are
identified, they can be used with Equation 9 to translate the concentration of selenium in eggs and ovaries
to a concentration of selenium in the water-column.
Because each TTF value is species-specific, it is possible to differentiate bioaccumulation in different
aquatic systems by modeling the food web of the target fish species. For example, where the food web
contains more than 3 trophic levels, TTFcomposlte can be represented as the product of all TTF values for
each trophic level given as:
TTF
composite
= TTF x TTF
TL3
TTFTLn
(Equation 10)
where n is the highest trophic level.
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The consumption of more than one species of organism at the same trophic level can also be modeled by
expressing the TTF value at a particular trophic level as the average TTF values of all species at that
trophic level weighted by the proportion of species consumed given as:
TLx
TTF = TTF^ x w;. (Equation 1 1)
i
where:
j,j,piLx _ ^e j-j-Qpiug transfer function of the ith species at a particular trophic level
w; = the proportion of the ith species consumed.
These concepts can be used to formulate a mathematical expression of ffpcomp°site that models selenium
bioaccumulation in a variety of aquatic ecosystems. Figure 1-2 illustrates four hypothetical food web
scenarios and the formulation of ffpcomp°site for each of them. For each scenario, the value of ffpcomp°site^
the CF value associated with the targeted fish species, and the site -specific EF value can be used with
Equation 9 to translate the egg -ovary FCV to a site-specific water concentration value. The general steps
required to derive a site-specific translation of the egg -ovary FCV are described below.
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A) Three trophic levels (simple):
lnPosile _ '/ V 7v ^^ V
* <•
B) Four trohic levels (simle):
Three trophic levels (mix within trohic levels):
TTFcompos"e
77
D) Three trohic levels (mix across trohic levels):
TTFTL3
Wj
W 2
E) Four trophic levels (mix across trohic levels):
TTFcompos"e =
x W
2)]
TTFTL2
Figure 1-2. Example mathematical expressions of TTp°omP°slte representing different food-web scenarios.
TTFcomposUe quantitatively represents the trophic transfer of selenium through all dietary pathways of a
targeted fish species. The mathematical expression of the food-web model is used to calculate a value for
TTFcomp°slte using appropriate species-specific TTF values and the proportions of each species consumed at
each trophic level. See text for further explanation.
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1.2 Steps for deriving a site-specific water concentration value from the egg-ovary FCV
Below are the steps that can be taken to derive a site-specific water concentration value the selenium egg-
ovary FCV by completing the following steps:
1) Identify the appropriate target fish species.
2) Model the food-web of the targeted fish species.
3) Identify appropriate TTF values by either:
a. selecting the appropriate TTF values from a list of EPA-derived values, or
b. deriving TTF values from existing data, or
c. deriving TTF values by conducting additional studies, or
d. extrapolating TTF values from existing values.
4) Determine the appropriate value of FFby either
a. deriving a site-specific EF value from field measurements, or
b. deriving an appropriate EF value from existing data, or
c. extrapolating from EF values of similar waters, or
5) Determine the appropriate CF value by either,
a. selecting the appropriate CF value a list of EPA-derived values, or
b. deriving a CF value from existing data, or
c. deriving a CF value by conducting additional studies, or
d. extrapolating a CF value from existing values.
6) Translate the selenium egg-ovary FCV into a site-specific water concentration value using
Equation 9.
Below are detailed descriptions of each step followed by example calculations using a variety of
hypothetical scenarios.
1.2.1 Identify the appropriate target fish species
1.2.1.1 When fish are present
The EPA's selenium criterion is expressed as a concentration in the eggs and ovaries offish because the
selenium concentration in the eggs and ovaries is the toxicological endpoint of selenium and thus the
most sensitive and consistent indicator of toxicity. Nonetheless, the relationship between selenium
concentration in these tissues and selenium toxicity vary across species due to differences in sensitivity
and bioaccumulation potential. Therefore, states and tribes should choose the fish species that resides in
the aquatic system with the greatest risk of selenium toxicity.
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The species most sensitive to selenium are those in the Salmonidae family. Thus, states and tribes should
target nonanadromous species in the Salmonidae family such as trout when they are present. Members of
the genus Lepomis (in the family Centrarchidae) that include bluegill are also sensitive and should be
targeted when no fish in the Salmonidae family are present. Other members of the Centrarchidae family
(such as bass) should be targeted if no fish of the genus Lepomis are present.
States and tribes should target nonanadromous species (species that do not migrate from salt water to
spawn in fresh water), because selenium exposure and subsequent bioaccumulation occurs over a
relatively long period of time through the consumption of locally contaminated aquatic organisms. If
nonanadromous fish species in the Salmonidae family is absent, states and tribes should target the
resident fish species likely to have the highest exposure and sensitivity to selenium. In aquatic systems
with resident fish species of unknown selenium sensitivity and bioaccumulation potential, factors such as
ecological significance can be factors in choosing which species to target. If the state or tribal monitoring
program uses lethal tissue sampling procedures, threatened or endangered species should not be used for
tissue monitoring.
Targeting fish species that consume organisms known or suspected to bioaccumulate selenium can be an
alternative approach to selecting fish species when species-specific information on selenium sensitivity
and bioaccumulation potential is unavailable. Prey organisms that pose a selenium toxicity risk to
predators usually have physiological characteristics that predispose them to selenium bioaccumulation
and/or are in close proximity to relatively high levels of selenium. For example, high levels of selenium
found in San Francisco Bay white sturgeon were linked to consumption of Potamocorbula amurensis, a
bivalve that was known to rapidly accumulate selenium and was in close proximity to selenium-
contaminated sediments (Stewart et al. 2004). In contrast, striped bass from the same aquatic system had
substantially lower levels of selenium due to their zooplankton-based food web with substantially lower
selenium bioaccumulation characteristics (Schlekat et al. 2004; Stewart et al. 2004).
States and tribes can use data from fisheries or biological surveys or other biological assessments to
determine the fish species that reside in specific surface waters. If such information is not available,
general information (often online) on the fish species that are present in state or tribal surface waters can
be found in:
• State Fish and Game agencies.
• U.S. Fish and Wildlife Service (http://www.fws.gov).
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• U.S. Geological Survey (http://www.usgs.gov).
• NatureServe.org (http://www.natureserve.org).
• Fishbase (http://www.fishbase.org).
• State or local sources of biological information (e.g. Biota Information System of New Mexico at
http://www.bison-m.org).
Figure 1-3 shows a decision tree that states and tribes may use to help select the appropriate fish species
for deriving a site-specific water concentration value from the selenium egg-ovary FCV. EPA
recommends this sequence of choices on the basis of taxonomic hierarchies that begin with taxa having
the highest sensitivity to selenium. The use of taxonomic hierarchies utilizes evolutionary relationships to
infer biological similarities among organisms (Suter 1993).
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Yes
^
Are nonanadromous species of the
Salmonidae family present?
'
Target nonanadromous species in the
Salmonidae family (e.g. trout)
Yes
^
'
Target species in genus Lepomis (e.g.,
bluegill)
Yes
^
'
Target species in family Centrarchidae
(e.g. bass)
Yes
^
'
Target resident species with confirmed or
suspected sensitivity or exposure risk to
selenium.
No
1 r
Are species in the genus Lepomis
present?
No
1 r
Is family Centrarchidae present?
No
1 r
Are resident species with confirmed or
suspected sensitivity or exposure risk to
selenium present?
No/do not know
1 r
Target species with highest ecological
significance.
Figure 1-3. Decision process for selection of the fish species to use when deriving a water concentration
from the selenium egg-ovary FCV.
1.2.1.2 When fish are absent
Some aquatic systems do not contain resident fish. Fish may be absent from a waterbody because of
intermittent or persistent low flows, physical impediments such as waterfalls or impoundments, lack of
adequate habitat for feeding and/or spawning, or intolerable aquatic conditions related to pH, turbidity,
temperature, salinity, total dissolved solids, chemical contaminants, or pathogens. These conditions could
be due to naturally occurring or anthropogenic causes. Some streams may be naturally intermittent or
ephemeral, or they might exhibit low or intermittent flows because of impoundments or water draw-down
for agricultural irrigation, industrial uses, drinking water supply, or other uses.
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When fish are naturally absent from a waterbody, states and tribes should target the most sensitive fish
species inhabiting downstream waters. Although the upper reaches of some aquatic systems may not
support fish communities, the invertebrate organisms that reside there may tolerate high concentrations of
selenium and pose a selenium risk if they are transported downstream. In such cases, states and tribes may
still use the decision tree in Figure J-3 to identify downstream fish species to protect. In addition, states
and tribes may evaluate upstream waters without fish by measuring the selenium concentration in water,
biotic and/or abiotic particulate material, and/or the tissues of invertebrate aquatic organisms that reside
there. Because selenium associated with particulate material and invertebrate organisms can be
transported downstream during intermittent high flows, elevated concentrations of selenium in the tissues
of downstream fish could indicate upstream sources of selenium that require a more detailed evaluation of
upstream conditions. A site-specific selenium criterion protecting a limited aquatic environment may be
appropriate if selenium levels are naturally high and fish were not previously present in the aquatic
system.
1.2.2 Model the food-web of the targeted fish species
After selecting the target fish species, states and tribes should formulate a mathematical expression of the
target species food-web that will be used to calculate the value of 777^°"^°^ AS discussed previously,
TTFcomposUe is the product of the TTF values across trophic levels of the target fish species food-web. The
complexity of the food-web model will depend on the species offish that is targeted, the diversity of prey
species in the aquatic system, and the amount of information that is available. Many of the same
information sources used to identify the targeted fish species in a waterbody might also be used to obtain
information about its food web. The types and proportions of food organisms consumed by the targeted
fish species can be directly determined through studies that examine stomach contents, or from
information gathered through biological assessments. If site-specific, field-derived information is not
available, the food-web characteristics of the target species can be estimated using publicly available
databases such as NatureServe (http://www.natureserve.org). For example, in the HUC watershed
#5040004 in Ohio, the NatureServe database record for fathead minnow indicates under the heading:
"Ecology and Life History - Food Comments," the fathead minnow "feeds opportunistically in soft
bottom mud; eats algae and other plants, insects, small crustaceans, and other invertebrates (Becker 1983,
Sublette et al. 1990)."
Additional sources of information include:
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• FishBase (http://www.fishbase.org). FishBase is a relational database developed at the World
Fish Center in collaboration with the Food and Agriculture Organization of the United Nations
(FAO) and many other partners.
• Carlander, K.D. Handbook of Freshwater Fishery Biology, volumes 1, 2 and 3. Iowa state
University Press, Ames, Iowa. 1969-1997.
1.2.3 Identify appropriate TTF values
The food-web model uses appropriately selected species-specific TTF values (and, if appropriate,
proportions within the same trophic level). States and tribes can determine the appropriate TTF values to
calculate ffpcomP°slte by either using one of the following four procedures, or by using other scientifically
defensible methods.
1.2.3.1 Select the appropriate TTF values from a list of EPA-derived values
Species-specific TTF values represent the steady state proportional concentration of selenium in the tissue
of an organism relative to the concentration of selenium in the food it consumes. TTF values for aquatic
invertebrates and fish are provided in Tables J-l and J-2 (see main text for a complete explanation of how
these values were derived).
Table 1-1. EPA-derived Trophic Transfer Function (TTF) values for freshwater aquati c
invertebrates.
Common name Scientific name
AE
IR
TTF
Crustaceans
amphipod
copepod
crayfish
water flea
Insects
dragonfly
damselfly
mayfly
midge
water boatman
Mollusks
asian clama
zebra mussel
Hyalella azteca
Copepods
Astacidae
Daphnia magna
Anisoptera
Coenagrionidae
Centroptilum triangulifer
Chironomidae
Corixidae
Corbiculafluminea
Dreissena polymorpha
1.22
0.520 0.420 0.155 1.41
1.46
0.406 0.210 0.116 0.74
1.97
2.88
0.390 0.720 0.220 1.28
1.90
1.48
0.550 0.050 0.006 4.58
0.260 0.400 0.026 4.00
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Annelids
blackworm Lumbriculus variegatus 0.165 0.067 0.009
Other
zooplankton Zooplankton -
a Not to be confused with Corbula amurensis
Table 1-2. EPA-derived Trophic Transfer Function (TTF) values for freshwater fish
Common name Scientific name AE IR ke
Cypriniformes
bluehead sucker Catostomus discobolus
common carp Cyprinus carpio
creek chub Semotilus atromaculatus
fathead minnow Pimephales promelas
flannelmouth sucker Catostomus latipinnis
longnose sucker Catostomus catostomus
sand shiner Notropis stramineus
white sucker Catostomus commersonii
Cyprinodontiformes
mosquitofish Gambusia sp. . . .
northern plains killifish Fundulus kansae
western mosquitofish Gambusia affmis
Esociformes
northern pike Esox lucius
Gasterosteiformes
brook stickleback Culaea inconstans
Perciformes
black crappie Pomoxis nigromaculatus
bluegill Lepomis macrochirus
green sunfish Lepomis cyanellus
largemouth bass Micropterus salmoides
striped bass Morons saxatilis 0.375 0.335 0.085
walleye Sander vitreus
yellow perch Percaflavescens
Salmoniformes
brook trout Salvelinus fontinalis
brown trout Salmo trutta
1.29
1.89
TTF
1.04
1.29
1.12
1.57
1.06
0.90
1.83
1.18
0.97
1.27
1.25
1.79
1.69
2.67
1.48
1.27
1.27
1.48
1.82
1.42
0.88
1.44
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cutthroat trout Oncorhynchus clarkii - - - 1.07
mountain whitefish Prosopiumwilliamsoni • • • 1.38
rainbow trout Oncorhynchus mykiss • • • 1.19
westslope cutthroat trout Oncorhynchus clarkii lewisi - - - 1.20
Scorpaeniformes
mottled sculpin Coitus bairdi • • • 1.38
sculpin Coitus sp. ... 1.29
Siluriformes
black bullhead Ameiurus melas • • • 0.91
channel catfish Ictalurus punctatus - - - 0.73
The TTF values from these lists could be used exclusively, or in conjunction with TTF values obtained
from other sources (see below). Note that these tables do not represent an exhaustive list of all TTF values
that may be required to calculate a site-specific water concentration value. If this list does not include a
required TTF value, states and tribes should refer to other approaches to obtain an appropriate value.
1.2.3.2 Deriving TTF values from existing data
If one or more appropriate TTF values cannot be found in Tables J-l and/or J-2, states and tribes could
derive species-specific TTF values using existing data. One approach for deriving species-specific TTF
values is to use the physiological coefficients representing food ingestion rate (IR), selenium efflux rate
(ke), and selenium assimilation efficiency (AE) to calculate a TTF value using Equation 3.
If the TTF value of a particular species in a food web is not available, TTF may be derived in several
different ways. One method is to obtain the physiological coefficients of food ingestion rate (IR),
assimilation efficiency (AE), or efflux rate (ke) and apply those values to Equation 3 given as:
AExIR
^
(Equation 3)
Where:
TTF = species-specific trophic transfer function
AE = species-specific assimilation efficiency (%)
IR = species-specific ingestion rate (g/g-d)
ke = species-specific efflux rate constant (/d)
The physiological coefficients IR, AE and ke may be obtained from published literature or may be derived
from laboratory studies. Another way to derive species-specific TTF values is to empirically assess the
relationship between the selenium concentration in the tissue of organisms and the selenium concentration
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in the food they consume using measurements from field studies. Species-specific TTF values can be
derived from such measurements by calculating ratios, using regression techniques, or other scientifically
defensible methods.
The physiological coefficients AE, IR, and ke are species-specific values. Coefficients AE and ke can only
be derived from laboratory experiments, butIR can be derived from either laboratory or field studies.
After the three physiological coefficients are obtained, a TTF value can be calculated using Equation 3.
Another approach for deriving species-specific TTF values is to use paired selenium measurements of
consumer organisms and their potential resources from field studies that directly measure the trophic
transfer of selenium in those organisms. The TTF for any trophic level can be defined by the equation:
rTLn
TTFTLn = iasHs (Equation 1-1)
Cfood
Where:
TTFTLn = The trophic transfer function of a given trophic level,
Ctissue = The selenium concentration (mg/kg dw) in the tissues of the consumer
organism,
selenium concentration (mg/kg dw) in the consumer organism's food.
At a given site, the empirical relationship between an organism and its food is first confirmed with linear
regression analysis. If the regression is both statistically significant (P < 0.05) and the slope of the
relationship is positive (i.e., selenium concentrations in the consumer increases with increasing selenium
in food), then the data are considered acceptable and the species-specific TTF is determined as the median
ratio of the paired consumer-food selenium concentration data. Both of the above methods were used to
derive the TTF values provided in Tables 1-1 and 1-2.
1.2.3.3 Deriving TTF values by conducting additional studies
States and Tribes may conduct additional studies to collect the data needed to derive TTF values for
specific needs or to revise existing TTF values. TTF values could be derived from new data using the
methodology described above, or other scientifically defensible methods. If available TTF values do not
apply to the species in a waterbody, there are no site-specific data available, and the collection of
necessary data by a state is impractical; then the state could require data collection where appropriate
based on Section 308 of the CWA (or comparable State authority).
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1.2.3.4 Extrapolating TTFvalues from existing values
When one or more TTF values are not available for a species, and the information needed to derive a
species-specific TTF value is not available or impractical to obtain, a TTF value can be extrapolated from
known TTF values. One possible method to extrapolate a TTF value is to sequentially consider higher
taxonomic classifications until one or more of the organisms with a known TTF value matches the taxon
being considered. If the lowest matching taxon is common to more than one of the available TTF values,
the average TTF from the matching table entries could be used. The use of taxonomic hierarchies in this
way utilizes evolutionary relationships to infer biological similarities among organisms (Suter 1993).
EPA used this extrapolation procedure to derive the TTF values of some of the organisms at some
representative aquatic sites that were used to derive the recommended water concentration values for lotic
and lentic waters. For example, the TTF value for Gila robusta, the roundtail chub, was not listed in Table
J-2. However, the roundtail chub is in the family Cyprinidae, which also includes Pimephales promelas,
the fathead minnow. Because Cyprinidae is the lowest taxonomic classification where the fish species
being considered matches a taxon in Table J-2, the TTF value for the fathead minnow was used for the
roundtail chub. In another example, Etheostoma exile, the Iowa darter, is not listed in Table J-2. However,
the Iowa darter is in the order Perciformes, which is common to the mangrove red snapper, striped bass,
largemouth bass, and bluegill. Thus the average of the TTF values from those five taxa in Table J-2 was
used for the Iowa darter.
1.2.4 Determine the appropriate EF value
The selenium enrichment function EF represents the bioavailability of selenium at the base of the aquatic
food web. The parameter EF varies more widely across aquatic systems than any other parameter, and is
influenced by the source and form of selenium, water residence time, and the biogeochemical
characteristics of the waterbody. Because EF can vary greatly across waterbodies, this parameter has the
greatest potential to introduce uncertainty in the translation from an egg-ovary concentration of selenium
to a water concentration and should be considered carefully. States and tribes can determine an
appropriate EF value either by using one of the following four procedures, or by using other scientifically
defensible methods.
1.2.4.1 Deriving a site-specific EF value from field measurements
The parameter EF can be expressed as the ratio of the concentration of selenium in particulate material to
the concentration of selenium dissolved in water. Using the equation
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f
7-T7-T particular /,-, . 7 7 ,
EF = — (Equation 11)
water
where
(2
parttcuiat = Concentration of selenium in participate material ((^g/g)
c
water - Concentration of selenium dissolved in water ((ig/L)
EF = Enrichment Function (L/g)
Deriving a site-specific EF value in this manner is a relatively straightforward and inexpensive procedure.
An EF value for a particular aquatic system can be derived by collecting water samples, separating the
particulate material from the water in each sample, measuring the concentration of selenium in the
separated water and particulate material, computing the ratio of the two measurements from each sample,
and then calculating the mean or median of all the ratios. Alternatively, a state or tribe could derive an EF
value by verifying the statistical relationship between paired particulate and water concentrations through
linear regression, calculating the ratios of paired Cparticuiate to Cwater values, and then taking the median
ratio as the value ofEF. This approach statistically evaluates the data representing the relationship
between these two media. However, a sufficient quantity of data is necessary to calculate statistically
significant fits.
Regardless of the method used to derive the value ofEF from field measurements, field and analytical
methods should be carefully planned and implemented when developing a site-specific, field-derived EF
value. Selenium bioaccumulation occurs more readily in aquatic systems with longer residence times
(such as wetlands, oxbows, and estuaries) and with fine particulate sediments high in organic carbon.
Thus EPA recommends a sampling plan that prioritizes areas with these characteristics. Analytical
methods to measure selenium in particulate material and in water are discussed in Appendix J.
1.2.4.2 Deriving an appropriate EF value from existing data
If suitable and sufficient site-specific measurements of Cparticulate and Cwater are available, states and tribes
could derive an EF value using these data. However, states and tribes should ensure that these data
represent current conditions, are based on scientifically acceptable sampling techniques, and are obtained
using proper quality assurance and quality control protocols to minimize uncertainty.
1.2 A3 Extrapolating from EF values of similar waters
In circumstances where a site-specific, field-derived EF value is not available, an EF value from one or
more aquatic systems with similar hydrological, geochemical, and biological characteristics could be used
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to approximate EF. However, states and tribes should carefully consider the possibility of introducing
significant uncertainty into the calculation when using EF values extrapolated from other waterbodies.
States and tribes should not use EF values derived from large-scale sites that encompass multiple water
bodies or ecosystems, or that do not match the characteristics of the waterbody for which the water
column concentration of selenium is being derived.
1.2.5 Determine the appropriate CF value.
1.2.5.1 Selecting the appropriate CF value from the list of values that were used to derive EPA's
recommended water criteria concentration values.
The parameter CF represents the species-specific proportion of selenium in eggs or ovaries relative to the
average concentration of selenium in all body tissues. EPA derived species-specific CF values for 14
species offish from studies that measured selenium concentrations in both eggs and/or ovaries and in
whole body and/or muscle. These CF values can be found in Appendix B and are reproduced below
(Table 1-3).
Table 1-3. Whole Body Se to Egg-Ovary Se Conversion Factors (CF)
Common name Scientific name CF
Cvpriniformes
bluehead sucker Catostomus discobolus 1.82
Common carp Cyprinus carpio 1.92
flannelmouth sucker Catostomus latipinnis 1.41
razorback sucker Xyrauchen texanus 1.43
roundtail chub Gilarobusta 2.07
White sucker Catostomus commersonii 1.41
Esociformes
northern pike Esox lucius 2.39
Perciformes
bluegill Lepomis macrochirus 2.13
green sunfish Lepomis cyanellus 1.45
smallmouth bass Micropterus dolomieu 1.42
Salmoniformes
brook trout Salvelinus fontinalis 1.38
brown trout Salmo trutta 1.45
cutthroat trout Oncorhynchus clarkii 2.30
Dolly Varden Salvelinus malma 1.61
mountain whitefish Prosopium williamsoni 7.39
1-20 Draft Document
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Common name Scientific name CF
rainbow trout Oncorhynchus mykiss 2.44
The data and methods used to derive CF for these species are described in Appendix B.
1.2.5.2 Deriving a CF value from existing data
The parameter CFis mathematically expressed as:
C
CF =
whol^body (Equation 16)
where
CF = Whole-body to egg-ovary conversion factor (dimensionless ratio).
Cegg-ovary = Selenium concentration in the eggs or ovaries of fish ((ig/g)
Cwhoie-body = Selenium concentration in the whole body offish (mg/kg).
If suitable and sufficient data are available, a state or tribe could derive a species-specific CF value using
the same numerical methods used to calculate the parameter EF. A state or tribe could calculate the ratio
of the two concentrations in each sample tissue as defined in Equation 13, and then calculate the mean or
median of all the ratios. Alternatively, a state or tribe could derive a CF value by verifying the statistical
relationship between paired particulate and water concentrations through linear regression, calculating the
ratios of paired Cegg^,ary to Cwhoie_body, and then taking the median ratio of the paired values as the CF (see
Appendix X). This approach statistically evaluates the data representing the relationship between these
two tissue types. However, a sufficient quantity of data is necessary to calculate statistically significant
fits. Regardless of the method used, care should be taken to ensure that the data used accurately represents
current conditions, were based on scientifically acceptable sampling techniques, and were obtained using
acceptable quality assurance and quality control protocols.
1.2.5.3 Deriving a CF value by conducting additional studies
States and tribes could perform additional studies to obtain the data needed to derive CF values for
specific needs or to revise existing CF values if there is reason to believe doing so may increase the
accuracy of the resulting water concentration. Analytical methods to measure selenium in tissue are
discussed in Appendix J. Where appropriate, additional data could be obtained as part of aNPDES permit
application by invoking authority under CWA section 308 (or comparable State authority) to reasonably
require NPDES-regulated facilities to collect information necessary to develop NPDES permit limits.
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1.2.5.4 Extrapolating the CF value from the list of values that were used to derive EPA's recommended
water criteria concentration values.
Because the pattern of selenium concentration in different body tissues varies between species,
extrapolating a species-specific CF value from one or more surrogate species is not recommended.
However, a CF value that is the average of all known species-specific CF values could be used when the
CF value of the target species is not available, and the data needed to derive a species-specific CF value is
not available or impractical to obtain. Using the average species CF value, however, lowers translation
accuracy and should only be used when other species-specific options are not available.
1.2.6 Translate the selenium egg-ovary FCV into a site-specific water concentration value using
Equation 18.
After determining the appropriate values of CF, TTFcomposlte (derived from the product of the individual
TTF values from each trophic level) and EF, a site-specific water concentration can be derived from the
egg-ovary FCV using Equation 18. Note thatNPDES permitting regulations at 40 CFR § 122.45(c)
requires that a Water Quality-Based Effluent Limit (WQBEL) for metals be expressed as total recoverable
metal, unless an exception is met under 40 CFR § 122.45(c)(l)-(3). Equation 18 assumes selenium
concentrations dissolved in water. While states and tribes may express ambient water quality criteria in
water quality standards as dissolved selenium, an additional step is necessary to convert the dissolved
selenium concentration to a total recoverable selenium concentration for the purpose of NPDES
permitting. Guidance for converting expression of metal concentrations in water from dissolved to total
recoverable can be found in Technical Support Document for Water Quality-based Toxics Control (U.S.
EPA 1991) and The Metals Translator: Guidance for Calculating a Total Recoverable Permit Limit from
a Dissolved Criterion (U.S. EPA 1996).
1.3 Managing uncertainty using the mechanistic modeling approach
Derivation of a water concentration from the egg-ovary FCV using the mechanistic bioaccumulation
modeling approach (Equation 18) is subject to uncertainties from several sources. These include:
• Measurement error when deriving input parameters.
• Unaccounted factors affecting bioaccumulation.
• Inaccurate identification or proportions of trophic level 2 food-web organisms.
• Inaccurate or inappropriate TTF, EF, or CF values.
• Biological variability.
• Other unknown factors.
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Though not required, the effectiveness of effluent limits and waste load alsites of selenium that are based
on water concentration values derived from the egg-ovary FCV should be confirmed whenever practical
using appropriate fish tissue assessment methods. In addition, comparing estimated selenium
concentrations in fish tissue with actual selenium concentrations obtained from small-scale field studies
could also help evaluate the suitability of selected equation parameters.
1.4 Example calculations
Below are six hypothetical examples that demonstrate how to calculate a selenium water concentration
from the egg-ovary FCV using Equation 18. These examples derive water concentration values for a
variety of hypothetical aquatic systems with various fish species and food webs. For these hypothetical
examples, species-specific TTF were taken from Tables 6 and 7 in the main text, and CF values were
taken from Table 8 from the main text. To calculate EF in these examples, the EPA used a hypothetical
water concentration of 5 (ig/L and the hypothetical particulate concentrations of 4.25 (ig/g and 8.75 (ig/g
in lotic and lentic aquatic systems, respectively.
1.4.1 Example 1
Bluegill (Lepomis macrochirus) in a river that consume mostly amphipods:
Current water concentration ((ig/L)
Current particulate concentration (mg/kg)
Trophic transfer function for bluegill (TTFTL3)
Trophic transfer function for amphipods (TTFTL2)
Egg-ovary to whole-body conversion factor for bluegill (CF)
Selenium egg-ovary FCV (mg/kg)
5.00
4.25
1.48
1.22
2.13
15.2
EF =
EF
4.25
Cp articulate
= 0.85 L/g
^egg-ovary
TTFcombined x Ep x Cp
(Equation 18)
x TTF1'
= 1.48 x 1.22
1-23 Draft Document
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= 1.81
'water ~
15.2
1.81 xO.85 X2.13
= 4.64 (ig/L
1.4.2 Example 2
Fathead minnow (Pimephales promelas) in a river that consume mostly copepods:
Current water concentration ((ig/L)
Current particulate concentration (mg/kg)
Trophic transfer function for fathead minnow (TTF113)
Trophic transfer function for copepods (TTFTL2)
Egg -ovary to whole-body conversion factor for fathead minnow (species-specific value
not available, so median CF for family Cyprinidae is used) (CF)
Selenium egg-ovary FCV (mg/kg)
5.00
4.25
1.57
1.41
2.00
15.2
EF =
^particulate
'water
(Equation 12)
EF =
4.25
= 0.85 L/g
e -
ovary
TTFcombined x EF X CF
•ombined rrrr-f~J'L3 .. rrrri-JTL2
x 1 lr
= 1.57x 1.41
= 2.21
15.2
'water ~
2.21x0.85x2.00
= 4.14(ig/L
1-24 Draft Document
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1.4.3 Example 3
Bluegill (Lepomis macrochirus) in a lake that consume mostly aquatic insects:
Current water concentration ((ig/L)
Current particulate concentration (mg/kg)
Trophic transfer function for Bluegill (TTFTL3)
Trophic transfer function for aquatic insects (median of Odonates, Water boatman,
Midges, and Mayflies) (TTFTL2)
Egg-ovary to whole-body conversion factor for Bluegill (CF)
Selenium egg-ovary FCV (mg/kg)
5.0
8.75
1.48
1.69
2.13
15.2
EF =
^particulate
'water
(Equation 12)
EF =
8.75
= 1.75L/g
('egg-ovary
^combined x Ep x Cp
(Equation 18)
:ombined
'water ~
= 1.63
= TTF1L3 x TTF
= 1.48x1.69
= 2.50
15.2
2.50x1.75x2.13
.TL2
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1.4.4 Example 4
Fathead minnow (Pimephales promelas) in a river that consume approximately % copepods and Vs aquatic
insects:
Current water concentration ((ig/L)
Current particulate concentration (mg/kg)
Trophic transfer function for fathead minnow (TTFTL3)
Trophic transfer function for copepods and aquatic insects (TTFTL2)
Copepods =1.41
Average of all aquatic insects = 1.69
ZM X10
rprppTL2 _ j=j
= (1.41 x%) + (1.69x i/3)
= 1.50
Egg -ovary to whole-body conversion factor for fathead minnow (species-specific value
not available, so median CF for family Cyprinidae is used). (CF)
Selenium egg-ovary FCV (mg/kg)
5.0
4.25
1.57
1.50
2.00
15.2
EF =
EF =
- particulate
r
'-'water
4.25
(Equation 12)
5.00
= 0.85 L/g
r
'-'egg-ovary
"water ~ TTf combined x £F X CF (EtlUatiOn
bmed=TTF x TTF
= 1.57 x 1.50
= 2.36
15.2
Cwater = 2.36x0.85x2.00
= 3.79(ig/L
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1.5.5 Example 5
Fathead chub (Platygobio gracilis) in a river with a diet of approximately 80% aquatic insects and 20%
algae:
Current water concentration ((ig/L)
Current particulate concentration (mg/kg)
Trophic transfer function of fathead chub:
Lowest matching taxon is the family Cyprinidae. Therefore, the TTF value of
Cyprinidae is used (TTFTL3)
Trophic transfer function for insects (TTFTL2)
Average of all aquatic insects = 1.69
Egg -ovary to whole-body conversion factor for fathead chub (species-specific value not
available, so median CF for family Cyprinidae is used). (CF)
Selenium egg-ovary FCV (mg/kg)
5.0
4.25
1.43
1.69
2.00
15.2
EF =
EF =
"particulate
r
'-'water
4.25
(Equation 12)
5.00
= 0.85 L/g
TTpcombined =
TjpTL2
Where:
wi = Proportion of fathead chub diet from insects; and
w2 = Proportion of fathead chub diet from algae
TTpcomb = [± 43 x 1. 69 X 0. 8] + [1. 43 X 0. 2]
= 2.22
15.2
water ~ 2.22x0.85x2.00
= 4.03 (ig/L
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1.5.6 Example 6
Largemouth bass (Micropterus salmoides) in a large river that consume mostly Western mosquitofish
(Gambusia qffinis) that consume approximately % insects and % crustaceans
Current water concentration ((ig/L)
Current particulate concentration (mg/kg)
Trophic transfer function of largemouth bass (TTFTL4)
Trophic transfer function of Western mosquitofish (TTFTL3)
Trophic transfer function for insects and crustaceans (TTFTL2)
Median all Insects - 1.69
Median all Crustaceans -
£(TTF^WI)
rprppTL2 _ ; = 1
= (1.69 x3/4) + (1.41 x%)
= 1.62
Egg-ovary to whole-body conversion factor for Largemouth bass (species-specific
value not available, so median for the genus Micropterus used) (CF)
Selenium egg-ovary FCV (mg/kg)
5.0
4.25
1.27
1.25
1.62
1.42
15.2
EF =
EF =
- particulate
r
'-'water
4.25
5.00
= 0.85 L/g
(Equation 12)
- i ir 4 x TTFTL3x TTF
= 1.27 x 1.25x 1.62
= 2.57
15.2
r
water 2 5? x Q 85 x ^ 42
= 4.90 (ig/L
TL2
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1.5.7 Example 7a.
Derivation of a site specific water column criterion for a river impacted by selenium.
Available data for a site indicates that the average egg/ovary tissue concentration of selenium for the
bluegill (Lepomis macrochirus) is 22 mg/kg (dw). This concentration exceeds the USEPA proposed
egg/ovary criterion of 15.2 mg/kg (dw). The allowable selenium water column criterion for this lotic
waterbody is 4.8 ug/L. The following calculation shows how to derive a water column concentration that
would achieve the 15.2 mg/kg (dw) egg/ovary tissue criterion.
Current selenium egg/ovary concentration (bluegill; mg/kg dw)
Selenium egg/ovary criterion (mg/kg, dw)
Selenium Water Column Criterion, Lotic Habitats (ug/L)
Allowable lotic water column concentration (ug/L)
22.0
15.2
4.8
X
1. Set up proportional equation to solve for allowable water column concentration
Lotic Water Column Criterion
Allowable Water concentration (X)
Current egg/ovary FT concentration
Selenium egg/ovary criterion
4.8 us/L
X
22 mg/kg dw
15.2 mg/kg dw
X
X
4.8 x 15.2 = 72.96
22 22
3.32 ug/L = Target Site specific Lotic Water Column Criterion
1.5.7 Example 7b.
Derivation of a site specific water column criterion for a lake impacted by selenium.
Available data for a site indicates that the average egg/ovary tissue concentration of selenium for the
bluegill (Lepomis macrochirus) is 22 mg/kg (dw). This concentration exceeds the USEPA proposed
egg/ovary criterion of 15.2 mg/kg (dw). The allowable selenium water column criterion for this lentic
1-29 Draft Document
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waterbody is 1.3 ug/L. The following calculation shows how to derive a water column concentration that
would achieve the 15.2 mg/kg (dw) egg/ovary tissue criterion.
Current selenium egg/ovary concentration (bluegill; mg/kg dw)
Selenium egg/ovary criterion (mg/kg, dw)
Selenium Water Column Criterion, Lentic Habitats (ug/L)
Allowable lotic water column concentration (ug/L)
22.0
15.2
1.3
X
2. Set up proportional equation to solve for allowable water colun concentration
= Lentic Water Column Criterion = Current egg/ovary FT concentration
Allowable Water concentration (X) Selenium egg/ovary criterion
1.3 ug/L = 22 mg/kg dw
X 15.2 mg/kg dw
X = 1.3 x 15.2 = 19.76
22 22
X = 0.89 ug/L = Target Site specific Lentic Water Column Criterion
2.0 Translating the concentration of selenium in tissue to a
concentration in water using Bioaccumulation Factors (BAF).
2.1 Summary of the BAF approach
A bioaccumulation factor (BAF) is the ratio (in milligrams/kilogram per milligrams/liter, or liters per
kilogram) of the concentration of a chemical in the tissue of an aquatic organism to the concentration of
the chemical dissolved in ambient water at the site of sampling (U.S. EPA 2001c). BAFs are used to
relate chemical concentrations in aquatic organisms to concentrations in the ambient media of aquatic
ecosystems where both the organism and its food are exposed and the ratio does not change substantially
overtime. The BAF is expressed mathematically as:
C
BAF =
r
vater (Equation 1-2)
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where
BAF = bioaccumulation factor derived from site-specific field-collected samples of
tissue and water (L/kg)
Ctissue = concentration of chemical in fish tissue (mg/kg)
Cwater = ambient concentration of chemical in water (mg/L)
Solving for Cwater:
f~< tissue /T^ , ,i
L water ~ TT7T7 (Equation 1-3)
To translate a fish tissue criterion to a water concentration value, states and tribes could develop a site-
specific, field-measured BAF for the waterbody, and then calculate a water concentration criterion using
Equation J-3. Detailed information about how to derive a site-specific, field-measured BAF is provided in
Methodology for Deriving Ambient Water Quality Criteria for the Protection of Human Health (2000)
Technical Support Document Volume 3: Development of Site-specific Bioaccumulation Factors (U.S.
EPA 2009). Although this guidance was developed for deriving human health criteria, the methodological
approach is also applicable to the derivation of aquatic life criteria.
2.2 Managing uncertainty using the BAF approach
Considerable uncertainty can be introduced when using the BAF approach to derive a water concentration
value from a fish tissue criterion concentration. Inaccurate water concentration values can result when
BAFs are derived from water and fish tissue concentration measurements that are obtained from sources
that do not closely represent site characteristics, or from field data collected from large-scale sites that
encompass multiple water bodies or ecosystems. Most of this uncertainty results from differences in the
bioavailability of selenium between the study sites where measurements are made to derive the BAF, and
the site(s) to which the BAF is used to derive needed water concentration values.
Because of uncertainties associated with the BAF approach, EPA does not recommend developing BAFs
from data extrapolated from different sites or across large spatial scales. EPA's Framework for Metals
Risk Assessment (U.S. EPA 2007) outlines key principles about metals and describes how they should be
considered in conducting human health and ecological risk assessments. The current science does not
support the use of a single, generic threshold BAF value as an indicator of metal bioaccumulation. The
use of BAFs are appropriate only for site-specific applications where sufficient measurements have been
taken from the site of interest and there is little or no extrapolation of BAF values across differing
exposure conditions and species.
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The preferred approach for using a BAF to implement the selenium fish tissue criterion is to calculate a
site-specific, field-measured BAF from data gathered at the site of interest, and to apply that BAF to that
site. A site-specific, field-measured BAF is a direct measure of bioaccumulation in an aquatic system
because the data are collected from the aquatic ecosystem itself and thus reflects real-world exposure
through all relevant exposure routes. A site-specific, field-measured BAF also reflects biotic and abiotic
factors that influence the bioavailability, biomagnification, metabolism, and biogeochemical cycling of
selenium that might affect bioaccumulation in the aquatic organism or its food web. Appropriately
developed site-specific, field-measured BAFs are appropriate for all bioaccumulative chemicals,
regardless of the extent of chemical metabolism in biota from a site (U.S. EPA 2000).
Although a site-specific, field-measured BAF is a direct measure of bioaccumulation, its predictive power
depends on a number of important factors being properly addressed in the design of the field sampling
effort. For example, sampling in areas with relatively long water residence times should be a priority
because selenium bioaccumulation occurs more readily in aquatic systems with longer residence times
(such as wetlands, oxbows, and estuaries) and with fine particulate sediments high in organic carbon. In
addition, migratory species should generally not be used because their exposure to selenium could reflect
selenium concentrations in areas other than where the fish were caught. Fish may also need to be sampled
and BAF values recalculated if selenium levels significantly change overtime because BAFs are known
to be affected by the ambient concentration of the metals in the aquatic environment (McGeer et al. 2003;
Borgman et al. 2004; DeForest et al. 2007). States and tribes should refer to Methodology for Deriving
Ambient Water Quality Criteria for the Protection of Human Health (2000) Technical Support Document
Volume (U.S. EPA 2009) for guidance on appropriate methods for developing a site-specific, field-derive
BAF.
The advantage of using the BAF approach is its relative simplicity, especially when the data necessary to
derive the BAF is already available. Furthermore, the BAF approach is completely empirical and does not
require any specific knowledge about the physical, chemical, or biological characteristics of the
waterbody. The relationship between the concentration of selenium in fish tissue and water is directly
determined by direct measurements in these media.
Limitations of the BAF approach should be considered before deciding if this method is appropriate for
translating the selenium FCV to a water concentration value. One disadvantage of the BAF approach is
the considerable time and cost necessary to collect sufficient data to establish the relationship between
tissue and water concentrations. Costs increase as the spatial scale and complexity of the aquatic system
1-32 Draft Document
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increases. Furthermore, the BAF approach does not allow extrapolation across species, space, and large
time scales because the site-specific factors that might influence bioaccumulation are integrated within
the tissue concentration measurements and thus cannot be individually adjusted to extrapolate to other
conditions. Thus, site-specific, field-measured BAFs only provide an accounting of the uptake and
accumulation of selenium for an organism at a specific site and point in time.
As noted previously, NPDES permitting regulations at 40 CFR § 122.45(c) require WQBELs for metals
be expressed as total recoverable metal unless an exception is met under 40 CFR § 122.45(c)(l)-(3).
Guidance for converting expression of metals in water from dissolved to total recoverable can be found in
Technical Support Document for Water Quality-based Toxics Control (U.S. EPA 1991) and TheMetals
Translator: Guidance for Calculating a Total Recoverable Permit Limit from a Dissolved Criterion (U.S.
EPA 1996). Whether or not a water concentration value derived from a site-specific, field-derived BAF
requires conversion from dissolved to total recoverable selenium depends on how the BAF is developed.
Generally, conversion would not be necessary if the BAF is derived from water concentration values that
measure total selenium; however, conversion would be necessary if the BAF was derived from water
concentration values that measured dissolved selenium. Table J-4 compares some of the principle
characteristics of the mechanistic bioaccumulation modeling approach or the BAF approach for
translating the selenium FCV to a water concentration.
3.0 Comparison of Mechanistic Bioaccumulation Modeling and
BAF approaches
Mechanistic bioaccumulation modeling
Bioaccumulation Factor (BAF)
Knowledge of the aquatic system needed
Choice of input parameters at discretion of State or
Tribe
Species-specific
Can be applied at different sites
Fish tissue sampling not required for translation
No information on aquatic system needed
No input parameters to choose
Species-specific
Site-specific
Fish tissue sampling required
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APPENDIX J: Analytical Methods for measuring
Selenium
J-l Draft Document
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The Clean Water Act (CWA) establishes an EPA approval process for certain analytical methods used in
the National Pollutant Discharge Elimination System (NPDES) program and for section 401
certifications. EPA has several approved methods for measuring selenium in water under 40 CFR § 136.
EPA generally requires the use of EPA-approved methods for the NPDES program and for CWA section
401 certifications issued by states and tribes (40 CFR § 136.1). However, since there are no EPA
approved methods for the analysis of selenium in fish tissue, states and tribes may use analytical methods
not approved by EPA to evaluate the attainment of water quality standards or to develop or implement
Total Maximum Daily Loads provided that these methods are scientifically sound (40 CFR 122.21(g)(7)).
Implementation of a water quality standard for selenium may require the ability to detect and measure the
concentration of selenium in effluent, ambient water, tissue, and other media that is below the detection
limit or limit of quantitation that some analytical methods can provide. States and tribes should choose an
analytical method that is sufficiently sensitive to implement its water quality standard for selenium.
Below are descriptions of some of the methods available for measuring selenium concentrations with
sufficient sensitivity to implement EPA's recommended selenium criterion. Complete descriptions of
analytical methods appropriate for analyzing selenium in different media can be found in the National
Environmental Methods Index at http://www.nemi.gov.
General considerations when measuring concentrations of selenium
The oxidation states of selenium dissolved in surface water are usually selenate (+6), selenite (+4), and
organo-selenium (-2). The presence of selenium in different oxidation states complicates some analytical
methods (Presser and Ohlendorf 1987). EPA recommends using standard reference samples to check for
the percentage recovery of each species of selenium (selenate, selenite and organo-selenium) during
initial testing of selenium methodologies for quality control and assurance.
If water samples are not filtered, particulate species such as elemental selenium and particulate organo-
selenium will also be measured. In addition, federal regulations at 40 CFR §122.45(c) generally requires
considering total recoverable metals when establishing effluent limits and reporting requirements.
J-2 Draft Document
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Analytical methods recommended for measuring selenium in water
EPA has several approved analytical methods under 40 CFR § 136 specifically for measuring total
selenium in water. These regulations state that measurements for NPDES permit applications and
permittee reporting should be made using analytical methods approved by EPA. Because EPA has
approved methods for analyzing selenium in water, these methods must be used for NPDES permits (40
CFR§ 122.21(g)(7), 122.41(j), 136.1, 136.3, and 136.6).
A complete list of EPA-approved analytical methods for selenium can be found at:
http://www.epa. gov/waterscience/methods/method/. Three EPA-approved methods that may be
sufficiently sensitive3 for the purposes of implementing a selenium water quality criterion are listed below
(Table J-l).
Table J-l. Suggested EPA-Approved Methods for Selenium in Water
Method
American Public Health Standard
Method 3 1 14 B (2009) or 3 1 14 C
(2009)
EPA Method 200.8, Rev 5.4
(1998)
EPA Method 200.9, Rev.2.2
(1994)
Technique
Hydride generation atomic absorption
spectrometry (HG-AAS)
Inductively coupled plasma mass
spectrometry (ICP-MS)
Stabilized temperature graphite
furnace atomic absorption (STGF-AA)
Method
detection limit
2^ig/L
7.9 Mg/L
0.6 ng/L
American Public Health Standard Method 3114 B
For measuring selenium in water, American Public Health Standard Method 3114 B uses the HG-AAS
technique. Method 3114 B has a method detection limit (MDL) of 2 (ig/L. Samples for dissolved analytes
should be filtered on-site through 0.45-micron capsule filters certified free of trace-element contamination
or other appropriate filtering equipment (Wilde et al. 1999). Dissolved samples should be preserved with
high purity hydrochloric acid or nitric acid to a pH less than 2.
For measuring total selenium, samples should not be filtered. In addition, all selenium in the sample
should be in the form of selenite (+4). Thus, the following pre-treatment steps to convert all selenium in
the sample to selenite are critical when using the HG-AAS method:
3For more information on choosing a sufficiently sensitive method, see the memorandum Analytical Methods for Mercury in
National Pollutant Discharge Elimination System (NPDES) Permits from James A. Hanlon, Director of the Office of Wastewater
Management, dated August 23, 2007, available at http://www.epa.gov/npdes/pubs/mercurymemo analvticalmethods.pdf.
J-3 Draft Document
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1. Boiling with persulfate to oxidize and digest organic material.
2. Boiling with hydrochloric acid to reduce selenate species to selenite.
3. Reduction by sodium borohydride to hydrogen selenide in the quartz tube of the AAS.
Optimal conversion conditions are essential for accurate results because too mild a reduction could lead to
incomplete reduction of selenate and too rigorous a reduction could lead to plating out of elemental
selenium (Cutter 1987, 1983; Presser and Barnes 1984, 1985).
Method 3114 B has the advantage that it is a fully validated method, is commonly used by many
laboratories, is relatively inexpensive, is less susceptible to background interference (Cutter 1987, 1983;
Presser and Barnes 1984, 1985), and has sufficient sensitivity to accurately measure what can be expected
in many lotic aquatic systems. However, this method may not be sufficiently sensitive for some lentic
aquatic systems where relatively lower selenium concentrations may need to be measured. If no selenium
is detected in a lentic system using this method, EPA recommends using a more sensitive analytical
method.
EPA Method200.8
EPA method 200.8 has a MDL of 7.9 (ig/L using the ICP-MS analytical technique. This method has the
advantage that no pre-treatment steps are necessary. However, this method may not be sufficiently
sensitive in many applications of the selenium criterion (Lamothe et al. 1999). If no selenium is detected
using this method, EPA recommends monitoring with a more sensitive method.
EPA Method200.9
Method 200.9 has a MDL of 0.6 (ig/L using the STGF-AA analytical technique. This method has the
advantage that it can detect selenium at very low concentrations. However, graphite furnace techniques
require careful matrix matching.
Of these three EPA approved methods, Method 3114 B using the HG-AAS technique is the most cost-
effective, with sufficient sensitivity and relatively low risk of interference in most cases. EPA Method
200.8 may be used where appropriate, such as when selenium concentrations in effluent are known to be
higher than 7.9 (ig/L. EPA Method 200.9 may be used if a very low MDL is needed.
Some additional methods not approved by EPA that states and tribes might consider are:
J-4 Draft Document
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• Collision/Reaction Cell Inductively Coupled Plasma Mass Spectroscopy (cICP-MS) (Garbarino
et al. 2005) - A relatively new technique that is a sensitive and selective detector for metal
analysis. However, isobaric interference can cause problems for quantitative determination as
well as identification based on the analyte pattern. Collision cells, reaction cells or other
interfaces reducing sample matrix effects that might otherwise interfere in the mass selective
determinative step are allowed in CWA analyses provided the method performance specifications
relevant to ICP-MS measurements are met
• Fluorometric Analysis,- a wet chemistry technique using diaminonapthalene. This method also
achieves acceptable precision and accuracy on standard reference samples (Olson 1969; Olson et
al. 1975; American Public Health Association Standard Method 3500, on-line version).
Methods for measuring different species of selenium dissolved in water are also available. These methods
determine the species of dissolved selenium present in a sample through differential digestion and hydride
generation atomic adsorption spectrophotometry (Cutter 1978, 1983; Presser and Barnes, 1984; 1985;
May et al. 2007). Selenite can be measured in samples with no pre-treatment. Selenate plus selenite can
be measured in samples subjected to boiling with hydrochloric acid. Subtraction of the measured selenite
fraction from the measured combined fraction would yield a measure of the selenate fraction. If a sample
is analyzed to measure total dissolved selenium as described above, then measurements of the combined
fraction can be subtracted to yield measurements of the dissolved organo-selenium fraction.
Analytical methods available for measuring selenium in fish tissue
EPA does not have approved methods under 40 CFR § 136 for measuring selenium in fish tissue.
However, states and tribes are not required to use EPA-approved methods for monitoring and assessment
of criteria attainment or other activities not related to permit applications or reports.
The techniques described above for analyzing selenium in water (HG-AAS, ICP-MS, and STGF-AA) can
be used to measure selenium in fish tissue if the samples are made soluble. Tissue samples are
homogenized and digested prior to analysis using strong acid or dry-ashing digestion as described below.
A review of sample digestion techniques has been published (Ihnat 1992). Standard reference materials,
analytical duplicates, and matrix spike samples are recommended to determine the applicability of a
selected digestion procedure.
J-5 Draft Document
-------
Strong acid digestion
Solid samples can be subjected to strong acid digestion to break down mineral and organic matrices.
Samples are typically dried and homogenized before digestion. Determination of percent moisture may be
part of the drying procedure. Note that some strong acid digestion methods may not be suitable for fish
tissue. Strong acid digestion methods are categorized by the type of material or amount of organic
material present (e.g., solid waste; biological tissue, plants, soil, sediment, rock, coal) and degrees of
tissue solubilization needed (extraction, leachate, or complete destruction). Methods differ in acid mixture
and degree and type of heating (EPA Method 3050B, Revision 2, 1996; EPA Method 200.2, Revision 2.8,
1994; Briggs and Crock, 1986; Taggart, 2002, chapters I, J, and K). High boiling acids (perchloric and
sulfuric) may lead to a loss of selenium if solutions are heated to dryness.
Dry-ashing digestion
Dry-ashing digestion is applicable to biological samples (Brumbaugh and Walther, 1989; May et al.,
2007). Biological samples are normally lyophilized (freeze-dried) and homogenized before digestion.
Determination of percent moisture may be part of the drying procedure. Dried solid samples are:
1. Boiled in nitric acid for solubilization and oxidation
2. Ashed at 500° C with magnesium nitrate to complete oxidation and decompose remaining organic
material
3. Heated with hydrochloric acid to dissolve the ash and reduce selenium to the selenite (+4) state
required for detection by HG-AAS.
Analytical methods available for measuring selenium in particulate material
There are no 40 CFR § 136 methods for analyzing selenium in particulate material. However, states and
tribes are not required to use EPA-approved methods for monitoring and assessment of criteria attainment
or other activities not related to permit applications or reports.
The techniques described above for analyzing selenium in water (HG-AAS, ICP-MS, and STGF-AA) can
be used to measure selenium in particulate material after the sample has been separated from the water
and pre-treated using the same methods used for fish tissue. In order to obtain a particulate material
sample, a water column sample should be filtered to separate the particulate material and bed sediment.
Various techniques for collection of suspended particulate material using filtration are available from the
EPA (e.g. Method 1669) and the U.S. Geological Survey (Moulton et al. 2002; USGS, Britton and
Greeson 1987). These techniques include:
J-6 Draft Document
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EPA Method 1669 (1996) includes filtration through a 0.45 (im capsule filter at the field site.
USGS protocols for collection of phytoplankton and seston in rivers and streams as part of their
National Water Quality Assessment Program for watershed and habitat assessment
(http ://water .usgs .gov/nawqa/protocols .html).
Textbooks such as Limnological Analyses address sampling of lakes using traditional techniques
including phytoplankton nets. (Wetzel and Likens 1991).
Sampling of suspended material from estuaries where particulates are a substantial part of the
ecosystem is described in Doblin et al. (2005) as part of their work on the San Francisco Bay-
Delta Estuary.
Separating suspended sediment using high-speed centrirugation and decantation when the
concentration of particulate material is relatively low (Horowitz et al. 1989).
J-7 Draft Document
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APPENDIX K: Abbreviations
K-l Draft Document
-------
Reference and site abbreviations
Reference
Bi:
Birkner 1978
Bu91:
Butler etal. 1991
Bu93:
Butler etal. 1993
Bu95:
Butler etal. 1995
Site
22
27
23
20
7
22
23
30
3
27
23
4
4
4
4
4
4
SP2
N2
SP2
N2
N2
N2
SP2
SP2
SP2
ME2
ME3
Miller's Lake, Wellington CO
Sweltzer Lake, Delta CO
Twin Butter Reservoir, Laramie WY
East Allen Reservoir, Medicine Bow WY
Galett Lake, Laramie WY
Miller's Lake, Wellington CO
Twin Butter Reservoir, Laramie WY
Larimer Highway 9 Pond, Fort Collins CO
Meeboer Lake, Laramie WY
Sweltzer Lake, Delta CO
Twin Butter Reservoir, Laramie WY
Uncompahgre River at Colona
Uncompahgre River at Colona
Uncompahgre River at Colona
Uncompahgre River at Colona
Uncompahgre River at Colona
Uncompahgre River at Colona
Spring Creek at La Boca
Navajo Reservoir, Piedra R. Arm, near La Boca
Spring Creek at La Boca
Navajo Reservoir, Piedra R. Arm, near La Boca
Navajo Reservoir, Piedra R. Arm, near La Boca
Navajo Reservoir, Piedra R. Arm, near La Boca
Spring Creek at La Boca
Spring Creek at La Boca
Spring Creek at La Boca
McElmo Cr., downstream from Alkali Canyon
McElmo Cr., upstream from Yellow Jacket Canyon
Species
FM
FM
FM
ID
ID
ID
ID
NPK
NPK
NPK
NPK
BhS
BnT
FS
MS
RT
WS
BhS
BT
BT
BB
ChC
cc
FM
SD
WS
BhS
BhS
Fathead minnow
Fathead minnow
Fathead minnow
Iowa darter
Iowa darter
Iowa darter
Iowa darter
Northern plains killfish
Northern plains killfish
Northern plains killfish
Northern plains killfish
Bluehead sucker
Brown trout
Flannelmouth sucker
Mottled sculpin
Rainbow trout
White sucker
Bluehead sucker
Brown trout
Brown trout
Black bullhead
Channel catfish
Common carp
Fathead minnow
Speckled dace
White sucker
Bluehead sucker
Bluehead sucker
K-2 Draft Document
-------
Reference
Bu97:
Butler etal. 1997
Site
NW
SJ1
SJ3
ME3
SJ1
SJ3
ME4
ME3
SJ1
SJ3
HD2
ME1
ME2
ME4
ME3
we
SJ1
HD2
ME2
ME4
ME3
SJ3
ME3
ME4
ME3
SJ1
ME1
ME2
ME3
NW
SJ1
HD2
MUD2
MNP2
Navajo Wash near Towaoc
San Juan R. at Four Corners
San Juan R. at Mexican Hat Utah
McElmo Cr., upstream from Yellow Jacket Canyon
San Juan R. at Four Corners
San Juan R. at Mexican Hat Utah
McElmo Cr., downstream from Yellow Jacket Canyon
McElmo Cr., upstream from Yellow Jacket Canyon
San Juan R. at Four Corners
San Juan R. at Mexican Hat Utah
Hartman Draw near mouth, at Cortez
McElmo Cr. at Hwy. 160, near Cortez
McElmo Cr., downstream from Alkali Canyon
McElmo Cr., downstream from Yellow Jacket Canyon
McElmo Cr., upstream from Yellow Jacket Canyon
Woods Canyon near Yellow Jacket
San Juan R. at Four Corners
Hartman Draw near mouth, at Cortez
McElmo Cr., downstream from Alkali Canyon
McElmo Cr., downstream from Yellow Jacket Canyon
McElmo Cr., upstream from Yellow Jacket Canyon
San Juan R. at Mexican Hat Utah
McElmo Cr., upstream from Yellow Jacket Canyon
McElmo Cr., downstream from Yellow Jacket Canyon
McElmo Cr., upstream from Yellow Jacket Canyon
San Juan R. at Four Corners
McElmo Cr. at Hwy. 160, near Cortez
McElmo Cr., downstream from Alkali Canyon
McElmo Cr., upstream from Yellow Jacket Canyon
Navajo Wash near Towaoc
San Juan R. at Four Corners
Hartman Draw near mouth, at Cortez
Mud Cr. at Hwy. 32, near Cortez
Large pond south of G Road, southern Mancos Valley
Species
BhS
BhS
BhS
BB
ChC
ChC
cc
cc
cc
cc
FM
FM
FM
FM
FM
FM
FS
FS
FS
FS
FS
FS
GnS
RSh
RSh
RSh
SD
SD
SD
SD
SD
Su
BhS
FM
Bluehead sucker
Bluehead sucker
Bluehead sucker
Black bullhead
Channel catfish
Channel catfish
Common carp
Common carp
Common carp
Common carp
Fathead minnow
Fathead minnow
Fathead minnow
Fathead minnow
Fathead minnow
Fathead minnow
Flannelmouth sucker
Flannelmouth sucker
Flannelmouth sucker
Flannelmouth sucker
Flannelmouth sucker
Flannelmouth sucker
Green sunfish
Red sunfish
Red sunfish
Red sunfish
Speckled dace
Speckled dace
Speckled dace
Speckled dace
Speckled dace
Sucker
Bluehead sucker
Fathead minnow
K-3 Draft Document
-------
Reference
Ca:
Casey and
Fo:
Formation 2012
Gr:
Grassoetal. 1995
Site
MUD2
WCP
CHI
MUD2
MNP3
DC
LC
CC-1A
CC-3A
CC-150
CC-350
CC-75
DC
HS
HS-3
LSV-2C
LSV-4
SFTC
CC-1A
CC-3A
CC-150
CC-350
CC-75
DC
HS
HS-3
LSV-2C
LSV-4
SFTC
17
17
Mud Cr. at Hwy. 32, near Cortez
Pond on Woods Canyon at 15 Road
Cahone Canyon at Hwy. 666
Mud Cr. at Hwy. 32, near Cortez
Pond downstream from site MNP2, southern Mancos Valley
Deerlick Creek
Luscar Creek
Crow Creek - 1A
Crow Creek - 3A
Crow Creek- 150
Crow Creek - 350
Crow Creek - 75
Deer Creek
Hoopes Spring
Hoopes Spring - 3
Sage Creek - 2C
Sage Creek - 4
South Fork Tincup Creek
Crow Creek - 1A
Crow Creek - 3A
Crow Creek- 150
Crow Creek - 350
Crow Creek - 75
Deer Creek
Hoopes Spring
Hoopes Spring - 3
Sage Creek - 2C
Sage Creek - 4
South Fork Tincup Creek
Arapahoe Wetlands Pond
Arapahoe Wetlands Pond
Species
FM
FM
GnS
GnS
SB
RT
RT
BnT
BnT
BnT
BnT
BnT
BnT
BnT
BnT
BnT
BnT
BnT
Sc
Sc
Sc
Sc
Sc
Sc
Sc
Sc
Sc
Sc
Sc
FM
WS
Fathead minnow
Fathead minnow
Green sunfish
Green sunfish
Smallmouth bass
Rainbow trout
Rainbow trout
Brown trout
Brown trout
Brown trout
Brown trout
Brown trout
Brown trout
Brown trout
Brown trout
Brown trout
Brown trout
Brown trout
Sculpin
Sculpin
Sculpin
Sculpin
Sculpin
Sculpin
Sculpin
Sculpin
Sculpin
Sculpin
Sculpin
Fathead minnow
White sucker
K-4 Draft Document
-------
Reference
HB:
Hamilton and
Buhl 2004
Le:
Lemly 1985
Sa87:
Saiki and
Lowe 1987
Sa93:
Saiki etal. 1993
Site
LEMC
BA
BE
HR
BA
BE
HR
BA
BE
HR
BA
BE
HR
BA
BE
HR
BA
BE
HR
KP11
KP2
KP8
SLD
VP26
VW
GT4
GTS
SJR2
SJR3
GT4
Lower East Mill Creek
Badin Lake
Belews Lake
High Rock Lake
Badin Lake
Belews Lake
High Rock Lake
Badin Lake
Belews Lake
High Rock Lake
Badin Lake
Belews Lake
High Rock Lake
Badin Lake
Belews Lake
High Rock Lake
Badin Lake
Belews Lake
High Rock Lake
Kesterson Pond 1 1
Kesterson Pond 2
Kesterson Pond 8
San Luis Drain
Volta Pond 26
Volta Wasteway
Salt Slough at San Luis Wildlife Refuge
Mud Slough at San Luis Wildlife Refuge
San Joaquin R. above Hills Ferry Rd.
San Joaquin R. at Durham Ferry Recreation Area
Salt Slough at San Luis Wildlife Refuge
Species
CT
BB
BB
BB
CC
CC
CC
FM
FM
FM
GnS
GnS
GnS
WM
WM
WM
RSh
RSh
RSh
WM
WM
WM
WM
WM
WM
Bg
Bg
Bg
Bg
LMB
Cutthroat trout
Black bullhead
Black bullhead
Black bullhead
Common carp
Common carp
Common carp
Fathead minnow
Fathead minnow
Fathead minnow
Green sunfish
Green sunfish
Green sunfish
Western mosquitofish
Western mosquitofish
Western mosquitofish
Red shiner
Red shiner
Red shiner
Western mosquitofish
Western mosquitofish
Western mosquitofish
Western mosquitofish
Western mosquitofish
Western mosquitofish
Bluegill
Bluegill
Bluegill
Bluegill
Largemouth bass
K-5 Draft Document
-------
Reference
St:
Stephens et al. 1988
Site
GTS
SJR2
SJR3
GT4
GTS
SJR2
SJR3
M4720
M4720
Mud Slough at San Luis Wildlife Refuge
San Joaquin R. above Hills Ferry Rd.
San Joaquin R. at Durham Ferry Recreation Area
Salt Slough at San Luis Wildlife Refuge
Mud Slough at San Luis Wildlife Refuge
San Joaquin R. above Hills Ferry Rd.
San Joaquin R. at Durham Ferry Recreation Area
Marsh 4720
Marsh 4720
Species
LMB
LMB
LMB
WM
WM
WM
WM
BB
CC
Largemouth bass
Largemouth bass
Largemouth bass
Western mosquitofish
Western mosquitofish
Western mosquitofish
Western mosquitofish
Black bullhead
Common carp
Reference and site abbreviations
Reference
Bi:
Birkner 1978
Bu91:
Butler etal. 1991
Site
22
27
23
20
7
22
23
30
3
27
23
4
4
4
4
4
4
Miller's Lake, Wellington CO
Sweltzer Lake, Delta CO
Twin Butter Reservoir, Laramie WY
East Allen Reservoir, Medicine Bow WY
Galett Lake, Laramie WY
Miller's Lake, Wellington CO
Twin Butter Reservoir, Laramie WY
Larimer Highway 9 Pond, Fort Collins CO
Meeboer Lake, Laramie WY
Sweltzer Lake, Delta CO
Twin Butter Reservoir, Laramie WY
Uncompahgre River at Colona
Uncompahgre River at Colona
Uncompahgre River at Colona
Uncompahgre River at Colona
Uncompahgre River at Colona
Uncompahgre River at Colona
Species
FM
FM
FM
ID
ID
ID
ID
NPK
NPK
NPK
NPK
BhS
BnT
FS
MS
RT
WS
Fathead minnow
Fathead minnow
Fathead minnow
Iowa darter
Iowa darter
Iowa darter
Iowa darter
Northern plains killfish
Northern plains killfish
Northern plains killfish
Northern plains killfish
Bluehead sucker
Brown trout
Flannelmouth sucker
Mottled sculpin
Rainbow trout
White sucker
K-6 Draft Document
-------
Reference
Bu93:
Butler etal. 1993
Bu95:
Butler etal. 1995
Site
SP2
N2
SP2
N2
N2
N2
SP2
SP2
SP2
ME2
ME3
NW
SJ1
SJ3
ME3
SJ1
SJ3
ME4
ME3
SJ1
SJ3
HD2
ME1
ME2
ME4
ME3
we
SJ1
HD2
ME2
ME4
ME3
SJ3
Spring Creek at La Boca
Navajo Reservoir, Piedra R. Arm, near La Boca
Spring Creek at La Boca
Navajo Reservoir, Piedra R. Arm, near La Boca
Navajo Reservoir, Piedra R. Arm, near La Boca
Navajo Reservoir, Piedra R. Arm, near La Boca
Spring Creek at La Boca
Spring Creek at La Boca
Spring Creek at La Boca
McElmo Cr., downstream from Alkali Canyon
McElmo Cr., upstream from Yellow Jacket Canyon
Navajo Wash near Towaoc
San Juan R. at Four Corners
San Juan R. at Mexican Hat Utah
McElmo Cr., upstream from Yellow Jacket Canyon
San Juan R. at Four Corners
San Juan R. at Mexican Hat Utah
McElmo Cr., downstream from Yellow Jacket Canyon
McElmo Cr., upstream from Yellow Jacket Canyon
San Juan R. at Four Corners
San Juan R. at Mexican Hat Utah
Hartman Draw near mouth, at Cortez
McElmo Cr. at Hwy. 160, near Cortez
McElmo Cr., downstream from Alkali Canyon
McElmo Cr., downstream from Yellow Jacket Canyon
McElmo Cr., upstream from Yellow Jacket Canyon
Woods Canyon near Yellow Jacket
San Juan R. at Four Corners
Hartman Draw near mouth, at Cortez
McElmo Cr., downstream from Alkali Canyon
McElmo Cr., downstream from Yellow Jacket Canyon
McElmo Cr., upstream from Yellow Jacket Canyon
San Juan R. at Mexican Hat Utah
Species
BhS
BT
BT
BB
ChC
cc
FM
SD
WS
BhS
BhS
BhS
BhS
BhS
BB
ChC
ChC
CC
cc
cc
cc
FM
FM
FM
FM
FM
FM
FS
FS
FS
FS
FS
FS
Bluehead sucker
Brown trout
Brown trout
Black bullhead
Channel catfish
Common carp
Fathead minnow
Speckled dace
White sucker
Bluehead sucker
Bluehead sucker
Bluehead sucker
Bluehead sucker
Bluehead sucker
Black bullhead
Channel catfish
Channel catfish
Common carp
Common carp
Common carp
Common carp
Fathead minnow
Fathead minnow
Fathead minnow
Fathead minnow
Fathead minnow
Fathead minnow
Flannelmouth sucker
Flannelmouth sucker
Flannelmouth sucker
Flannelmouth sucker
Flannelmouth sucker
Flannelmouth sucker
K-7 Draft Document
-------
Reference
Bu97:
Butler etal. 1997
Ca:
Casey and
Fo:
Formation 2012
Site
ME3
ME4
ME3
SJ1
ME1
ME2
ME3
NW
SJ1
HD2
MUD2
MNP2
MUD2
WCP
CHI
MUD2
MNP3
DC
LC
CC-1A
CC-3A
CC-150
CC-350
CC-75
DC
HS
HS-3
LSV-2C
LSV-4
SFTC
CC-1A
CC-3A
McElmo Cr., upstream from Yellow Jacket Canyon
McElmo Cr., downstream from Yellow Jacket Canyon
McElmo Cr., upstream from Yellow Jacket Canyon
San Juan R. at Four Corners
McElmo Cr. at Hwy. 160, near Cortez
McElmo Cr., downstream from Alkali Canyon
McElmo Cr., upstream from Yellow Jacket Canyon
Navajo Wash near Towaoc
San Juan R. at Four Corners
Hartman Draw near mouth, at Cortez
Mud Cr. at Hwy. 32, near Cortez
Large pond south of G Road, southern Mancos Valley
Mud Cr. at Hwy. 32, near Cortez
Pond on Woods Canyon at 15 Road
Cahone Canyon at Hwy. 666
Mud Cr. at Hwy. 32, near Cortez
Pond downstream from site MNP2, southern Mancos Valley
Deerlick Creek
Luscar Creek
Crow Creek - 1A
Crow Creek - 3A
Crow Creek- 150
Crow Creek - 350
Crow Creek - 75
Deer Creek
Hoopes Spring
Hoopes Spring - 3
Sage Creek - 2C
Sage Creek - 4
South Fork Tincup Creek
Crow Creek - 1A
Crow Creek - 3A
Species
GnS
RSh
RSh
RSh
SD
SD
SD
SD
SD
Su
BhS
FM
FM
FM
GnS
GnS
SB
RT
RT
BnT
BnT
BnT
BnT
BnT
BnT
BnT
BnT
BnT
BnT
BnT
Sc
Sc
Green sunfish
Red sunfish
Red sunfish
Red sunfish
Speckled dace
Speckled dace
Speckled dace
Speckled dace
Speckled dace
Sucker
Bluehead sucker
Fathead minnow
Fathead minnow
Fathead minnow
Green sunfish
Green sunfish
Smallmouth bass
Rainbow trout
Rainbow trout
Brown trout
Brown trout
Brown trout
Brown trout
Brown trout
Brown trout
Brown trout
Brown trout
Brown trout
Brown trout
Brown trout
Sculpin
Sculpin
K-8 Draft Document
-------
Reference
Gr:
Grassoetal. 1995
HB:
Hamilton and
Buhl 2004
Le:
Lemly 1985
Site
CC-150
CC-350
CC-75
DC
HS
HS-3
LSV-2C
LSV-4
SFTC
17
17
LEMC
BA
BE
HR
BA
BE
HR
BA
BE
HR
BA
BE
HR
BA
BE
HR
BA
BE
HR
Crow Creek- 150
Crow Creek - 350
Crow Creek - 75
Deer Creek
Hoopes Spring
Hoopes Spring - 3
Sage Creek - 2C
Sage Creek - 4
South Fork Tincup Creek
Arapahoe Wetlands Pond
Arapahoe Wetlands Pond
Lower East Mill Creek
Badin Lake
Belews Lake
High Rock Lake
Badin Lake
Belews Lake
High Rock Lake
Badin Lake
Belews Lake
High Rock Lake
Badin Lake
Belews Lake
High Rock Lake
Badin Lake
Belews Lake
High Rock Lake
Badin Lake
Belews Lake
High Rock Lake
Species
Sc
Sc
Sc
Sc
Sc
Sc
Sc
Sc
Sc
FM
WS
CT
BB
BB
BB
CC
CC
CC
FM
FM
FM
GnS
GnS
GnS
WM
WM
WM
RSh
RSh
RSh
Sculpin
Sculpin
Sculpin
Sculpin
Sculpin
Sculpin
Sculpin
Sculpin
Sculpin
Fathead minnow
White sucker
Cutthroat trout
Black bullhead
Black bullhead
Black bullhead
Common carp
Common carp
Common carp
Fathead minnow
Fathead minnow
Fathead minnow
Green sunfish
Green sunfish
Green sunfish
Western mosquitofish
Western mosquitofish
Western mosquitofish
Red shiner
Red shiner
Red shiner
K-9 Draft Document
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Reference
Sa87:
Saiki and
Lowe 1987
Sa93:
Saiki etal. 1993
St:
Stephens et al. 1988
Site
KP11
KP2
KP8
SLD
VP26
VW
GT4
GTS
SJR2
SJR3
GT4
GTS
SJR2
SJR3
GT4
GTS
SJR2
SJR3
M4720
M4720
Kesterson Pond 1 1
Kesterson Pond 2
Kesterson Pond 8
San Luis Drain
Volta Pond 26
Volta Wasteway
Salt Slough at San Luis Wildlife Refuge
Mud Slough at San Luis Wildlife Refuge
San Joaquin R. above Hills Ferry Rd.
San Joaquin R. at Durham Ferry Recreation Area
Salt Slough at San Luis Wildlife Refuge
Mud Slough at San Luis Wildlife Refuge
San Joaquin R. above Hills Ferry Rd.
San Joaquin R. at Durham Ferry Recreation Area
Salt Slough at San Luis Wildlife Refuge
Mud Slough at San Luis Wildlife Refuge
San Joaquin R. above Hills Ferry Rd.
San Joaquin R. at Durham Ferry Recreation Area
Marsh 4720
Marsh 4720
Species
WM
WM
WM
WM
WM
WM
Bg
Bg
Bg
Bg
LMB
LMB
LMB
LMB
WM
WM
WM
WM
BB
CC
Western mosquitofish
Western mosquitofish
Western mosquitofish
Western mosquitofish
Western mosquitofish
Western mosquitofish
Bluegill
Bluegill
Bluegill
Bluegill
Largemouth bass
Largemouth bass
Largemouth bass
Largemouth bass
Western mosquitofish
Western mosquitofish
Western mosquitofish
Western mosquitofish
Black bullhead
Common carp
K-10 Draft Document
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