jfh ¦¦¦¦¦ypi United States Office of Water EPA 822-P-16-002
¦mLXejL Environmental Mail Code 4304T December 2016
¦¦¦¦ ft Protection Agency
Human Health Recreational Ambient Water
Quality Criteria or Swimming Advisories for
Microcystins and Cylindrospermopsin
Draft
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Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin
Draft
Prepared by:
U.S. Environmental Protection Agency
Office of Water (4304T)
Health and Ecological Criteria Division
Washington, DC
EPA Document Number: 822-P-16-002
Date: December 2016
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NOTICES
This information is distributed solely for the purpose of obtaining scientific views on the
content of this document. It does not represent and should not be construed to represent any final
agency determination or policy. Mention of trade names or commercial products does not
constitute endorsement or recommendation for use.
Human Health Recreational Ambient Water Quality Criteria or ii
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FOREWORD
Section 304(a) of the Clean Water Act 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.
U.S. Environmental Protection Agency (EPA) is publishing these recommended values
under Clean Water Act (CWA) 304(a) for states to consider as the basis for swimming advisories
for notification purposes in recreational waters to protect the public. Alternatively, states may
consider using these same values when adopting new or revised water quality standards (WQS).
If adopted as WQS and approved by EPA under CWA 303(c), the WQS could be used for all
CWA purposes. States may also wish to consider using these values as both swimming advisory
values and WQS. EPA envisions that if states decide to use the values as swimming advisory
values they might do so in a manner similar to their current recreational water advisory
programs.
This draft document has undergone an EPA intra-agency peer review process. Final
review by the Health and Ecological Criteria Division, Office of Science and Technology, Office
of Water, U.S. Environmental Protection Agency has been completed and the document is
approved for publication. These values were derived using the existing peer-reviewed and
published science on the adverse human health effects of these toxins, established criteria
methodologies, and recreation-specific exposure parameters from EPA's Exposure Factors
Handbook.
The term "water quality criteria" is used in two sections of the CWA—§304(a)(l) and
§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 effects on human health or aquatic life. The
criteria recommendations presented in this document are such a scientific assessment. If water
quality criteria associated with specific designated uses are adopted by a state or authorized tribe
as water quality standards under section 303, and approved by EPA, they become applicable
Clean Water Act water quality standards in ambient waters within that state or tribe. Water
quality criteria adopted in state or tribal water quality standards could have the same numerical
values as criteria developed under section 304. Alternatively, states and authorized tribes may
derive numeric criteria based on other scientifically defensible methods but the criteria must be
protective of designated uses. It is not until their adoption as part of state or tribal water quality
standards, and subsequent approval by EPA, that criteria become Clean Water Act applicable
water quality standards. Guidelines to assist in modifying the criteria recommendations presented
in this document are contained in the Water Quality Standards Handbook (U.S. EPA 2012b).
This handbook and additional guidance on the development of WQS and other water-related
programs of this agency have been developed by EPA which along with 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 document provides recommendations 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.
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/signed/
Elizabeth Southerland
Director
Office of Science and Technology
Human Health Recreational Ambient Water Quality Criteria or iv
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ACKNOWLEDGMENTS
The development of this document was made possible through an effort led by John
Ravenscroft, EPA Project Manager, Health and Ecological Criteria Division, Office of Science
and Technology, Office of Water. Additionally, the following Office of Science and Technology
staff provided valuable contributions to the development and review of this document: Tracy
Bone, Dr. Lesley D'Anglada, Ashley Harper, Ana-Maria Murphy-Teixidor, and Lars Wilcut, and
Oakridge Institute for Science and Education (ORISE) fellow Meghann Niesen.
EPA gratefully acknowledges the valuable contributions of EPA Internal Technical
Reviewers who reviewed this document. This recreational Ambient Water Quality Criteria
(AWQC) was provided for review and comment from a formal internal agency review by staff in
the following U.S. EPA Program and Regional Offices:
U.S. EPA Office of Children's Health Protection: Suril Mehta
U.S. EPA Office of General Counsel: David Berol, Lee Schroer
U.S. EPA Office of Policy: Sharon Cooperstein
U.S. EPA Office of Research and Development: Neil Chernoff, Paula Estornell (on
detail), Elizabeth Hilborn, Christopher Impellitteri, Nicole Shao
U.S. EPA Office of Water
Office of Ground Water and Drinking Water: Hannah Holsinger, Mike Muse
Office of Science and Technology: Tracy Bone
Office of Wastewater Management: David Hair, Virginia Kibler
Office of Wetlands, Oceans, and Watersheds: Rosaura Conde, Katharine Dowell
U.S. EPA Regional Offices
Region 1: Toby Stover
Region 4: Joel Hansel
Region 5: Meghan Hemkin
Region 7: Amy Shields
Region 8: Alfred Basile, Tina Laidlaw
Technical support was provided by ICF and its subcontractor Bigelow Laboratory for Ocean
Sciences.
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TABLE OF CONTENTS
NOTICES ii
FOREWORD iii
ACKNOWLEDGMENTS v
TABLE OF CONTENTS vi
LIST OF FIGURES viii
LIST OF TABLES ix
ACRONYMS AM) ABBREVIATIONS xi
1.0 EXECUTIVE SUMMARY 1
2.0 INTRODUCTION AM) BACKGROUND 3
2.1 Clean Water Act 6
2.2 International and State Guidelines 7
3.0 NATURE 01 THE STRESSORS 15
3.1 Cyanobacteria and Cyanobacterial Blooms 15
3.2 Cyanotoxins 21
3.2.1 Chemical and Physical Properties 21
3.2.2 Sources and Occurrence 24
3.2.3 Environmental Fate 30
3.2.4 Toxicokinetics 33
4.0 PROBLEM FORMULATION 34
4.1 Conceptual Model 34
4.2 Analysis Plan 36
4.2.1 Approach for Recreational AWQC Derivation 37
4.2.2 Measures of Effect 37
4.2.3 Measures of Exposure 38
4.2.4 Relative Source Contribution 44
5.0 EFFECTS ASSESSMENT 45
5.1 Hazard Identification 45
5.1.1 Noncancer Health Effects 45
5.1.2 Cancer 49
5.2 Dose-Response Assessment 50
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6.0 SWIMMING ADVISORY AND RECREATIONAL CRITERIA DERIVATION 51
6.1 Microcystins Magnitude 51
6.2 Cylindrospermopsin Magnitude 51
6.3 Recommended Swimming Advisory and Recreational Criteria for Microcystins
and Cylindrospermopsin 52
7.0 EFFECTS CHARACTERIZATION 54
7.1 Cyanobacterial Cells 54
7.1.1 Cyanobacterial Cells Related to Inflammatory Health Effects 54
7.1.2 Cyanobacterial Cells as Indicators for Potential Toxin Production 57
7.2 Enhanced Risk or Susceptibility 59
7.3 Other Studies of Ingestion While Swimming 60
7.4 Distribution of Potential Recreational Health Protective Values by Age 65
7.4.1 Evaluation of Criteria Related Lifestages 65
7.4.2 Evaluation of Younger Children's Exposure Factors 69
7.5 Other Recreational Exposures 71
7.5.1 Other Recreational Exposure Pathways 71
7.5.2 Tribal Considerations 75
7.6 Livestock and Pet Concerns 75
7.6.1 States and Animal HAB Guidelines 77
8.0 REFERENCES 79
APPENDIX A. INTERNATIONAL RECREATIONAL WATER GUIDELINES
FOR CYANOTOXINS AND CYANOBACTERIA A-l
APPENDIX B. STATE RECREATIONAL WATER GUIDELINES FOR
CYANOTOXINS AND CYANOBACTERIA B-l
APPENDIX C. LITERATURE SEARCH DOCUMENTATION C-l
APPENDIX D. REVIEW OF THE STATE OF THE SCIENCE ON
CYANOBACTERIAL CELLS HEALTH EFFECTS D-l
APPENDIX E. INCIDENTAL INGESTION EXPOSURE FACTOR COMBINED
DISTRIBUTION ANALYSIS E-l
APPENDIX F. INFORMATION ON CELLULAR CYANOTOXIN AMOUNTS
AND CONVERSION FACTORS F-l
APPENDIX G. TABLES OF STATE-ISSUED GUIDANCE SPECIFIC TO
AMM AI. CYANOTOXIN POISONING G-l
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LIST OF FIGURES
Figure 2-1. Generalized Distribution of Cyanobacterial HABs in the United States
and Territories 4
Figure 2-2. State-reported HAB Advisories by EPA Region for January 1 to August
12, 2016 5
Figure 2-3. State Guidelines for Cyanotoxins and Cyanobacteria in Recreational
Water by Type and Scope of Guidelines 11
Figure 3-1. Environmental Factors Influencing Cyanobacterial Bloom Potential in Aquatic
Ecosystems, Reproduced from Paerl and Otten (2013b) 17
Figure 3-2. Structure of Microcystin (Kondo et al. 1992) 22
Figure 3-3. Structure of Cylindrospermopsin (de la Cruz et al. 2013) 23
Figure 4-1. Conceptual Model of Exposure Pathways to the Cyanotoxins,
Microcystins and Cylindrospermopsin, and Cyanobacteria in Surface
Waters while Recreating 34
Figure 4-2. Incidental Ingestion Rates Measured for Adults and Children (Dufour et
al. 2006) 41
Figure 4-3. Direct Contact Recreational Exposure Duration by Age Group, Based on
Table 15-119 in U.S. EPA (1997) 41
Figure 4-4. Hybrid Distributions for Incidental Ingestion per Day (L/d) 43
Figure 7-1. Comparison of Recreational Health Protective Values for Microcystins
and Cylindrospermopsin for Children, Adults, and General Population 66
Figure 7-2. Comparison of Alternative Health Protective Recreational Values and
Recreational AWQC for Microcystins and Cylindrospermopsin
Calculated based on Evans et al. (2006) 68
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LIST OF TABLES
Table 2-1. WHO (2003b) Recreational Guidance/Action Levels for Cyanobacteria,
Chlorophyll a, and Microcystin
Table 2-2. International Recreational Water Guideline or Action Levels for
Cyanobacteria and Microcystins
Table 2-3. State Guideline or Action Levels for Microcystin, Cylindrospermopsin,
and Cyanobacterial Cells in Recreational Water 1
Table 3-1. Abbreviations for Microcystins (Yuan et al. 1999)
Table 3-2. Chemical and Physical Properites of Microcystin-LR
Table 3-3. Chemical and Physical Properties of Cylindrospermopsin
Table 3-4. States Surveyed as Part of the 2007 National Lakes Assessment with
Water Body Microcystins Concentrations above 10 [j,g/L (U.S. EPA 2009)
Table 4-1. Summary Statistics of Combined Ingestion Volume and Exposure
Duration Distributions
Table 6-1. Recreational Criteria or Swimming Advisory Recommendations for
Microcystins and Cylindrospermopsin
Table 7-1. Studies of Incidental Ingestion Volumes or Rates While Recreating
Table 7-2. Comparison of Daily Ingestion Rates While Recreating between Dufour et
al. (2006) and Evans et al. (2006)a
Table 7-3. Alternative Recreational Criteria Values for Microcystins and
Cylindrospermopsin Calculated based on Alternative Ingestion Data from
Evans et al. (2006)
Table 7-4. Comparison of Younger Children's Exposure Factors and Incidental
Ingestion Data Sets
Table 7-5. Comparison of Recreational Exposure Ingested Dose to Inhaled Dose of
Microcystin
Table 7-6. Comparison of Recreational Exposure Ingested Dose to Dermal Absorbed
Dose of Microcystins
Table B-l. Summary Counts of State Recreational Water Guidelines for Cyanotoxins
and Cyanobacteria by Type and Scope of Guidelines
Table B-2. Summary Counts of State Recreational Water Guidelines for Cyanotoxins
and Cyanobacteria by Basis of Guidelines
Table B-3. State Recreational Water Quality Guideline for Cyanotoxins and
Cyanobacteria Sorted by Type
Table C-l. Internet URL Domains Searched for Research Question 4
Table C-2. Number of Journal Articles Returned by Three Search Strategies for
Research Question 5
Table D-l. Cyanobacteria Epidemiological Studies Summary
Table E-l. Parameters Used to Fit Distributions
Table E-2. Summary Statistics of Hybrid Distribution
Table F-l. Cell Quotas for Cyanotoxins Available from a Spot Check of the
Literature
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Table F-2. A Brief Summary of Cell Concentration - Cyanotoxin Conversions
Available from a Spot Check of the Literature 4
Table G-l. California Environmental Protection Agency (2012) Action levels for
Selected Pet and Livestock Scenarios 1
Table G-2. California Environmental Protection Agency (2012) Reference Doses and
Acute and Subchronic Action Levels for Canine Exposure to Cyanotoxins
in Drinking Water 1
Table G-3. Oregon Dog-specific Guideline Values for Cyanotoxins in Recreational
Waters (|ig/L) 2
Table G-4. Grayson County Texas Microcystin Guidelines for Dogs 2
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ACRONYMS AND ABBREVIATIONS
AWQC Ambient Water Quality Criteria
BGAS blue-green algae supplements
bw body weight
CalEPA California Environmental Protection Agency
CDC U.S. Centers for Disease Control and Prevention
CI confidence interval
CWA Clean Water Act
CYP450 Cytochrome P450
ELISA Enzyme Linked Immunosorbent Assays
EPA U.S. Environmental Protection Agency
GI gastrointestinal
HAB harmful algal bloom
HESD Health Effects Support Document
HPLC high performance liquid chromatography
IARC International Agency for Research on Cancer
i.p. intraperitoneal
kg kilograms
Koc soil organic carbon-water partition coefficient
Kow octanol-water partition coefficient
L liter
LOAEL lowest-observed-adverse-effect-level
LOD level of detection
LPS lipopolysaccharide
mL milliliters
NLA National Lakes Assessment
NOAEL no-observed-adverse-effect-level
OATp organic acid transporter polypeptide
OPP EPA Offi ce of Pe sti ci de Program s
OR odds ratio
pg picogram
RfD reference dose
ROS reactive oxygen species
RSC relative source contribution
SWIMODEL Swimmers Exposure Assessment Model
TMDL Total Maximum Daily Load
UF uncertainty factor
UFa uncertainty factor for interspecies variability
UFd database uncertainty factor
UFh uncertainty factor for intraspecies variability
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UFl
uncertainty factor for LOAEL to NOAEL extrapolation
Hg
microgram
USGS
U.S. Geological Survey
WHO
World Health Organization
WQBEL
Water Quality-Based Effluent Limits
WQS
water quality standards
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1.0 EXECUTIVE SUMMARY
Cyanobacteria, also commonly referred to as blue-green algae, are photosynthetic
bacteria that are ubiquitous in nature, including surface waters. Environmental conditions that
promote excessive growth of cyanobacteria in surface waters can lead to situations in which
cyanobacterial cell density is high, known as blooms. Environmental factors that play an
important role in the development of cyanobacterial blooms and their production of cyanotoxins
include the levels of nitrogen and phosphorus, the ratio of nitrogen to phosphorus, temperature,
organic matter availability, light attenuation, and pH.
Microcystins can be produced by a variety of cyanobacteria genera including
Microcystis, Anabaena, Nostoc, Oscillatoria, Fischerella, Planktothrix, and Gloeotrichia. Some
of these species can be distributed through the water column, concentrate in the upper layers, or
form surface scums depending on environmental conditions. More than 100 microcystin
congeners exist, which vary based on amino acid composition. The majority of toxicological data
on the effects of microcystins are available for microcystin-LR, which is also a frequently
monitored congener. Microcystins are water-soluble and tend to remain contained within the
cyanobacterial cell, until the cell breaks, and they are released into the water. Microcystins
typically have a half-life of 4 to 14 days in surface waters or may persist longer, depending on
such factors as the degree of natural degradation owing to sunlight, organic matter, and the
presence of bacteria. Microcystins can persist even after a cyanobacterial bloom is no longer
visible.
Cylindrospermopsin can be produced by a variety of cyanobacteria species including
Cylindrospermopsis raciborskii, Aphanizomenon species, Anabaena species, Lyngbya wollei,
and Rhaphidiopsis species. Some of these species tend not to form visible surface scums, and the
highest concentrations of cyanobacterial cells typically occur below the water surface.
Cylindrospermopsin may be retained within the cell or released into the water. The
biodegradation of cylindrospermopsin in natural water bodies is a complex process that can be
influenced by many environmental factors, including toxin concentration, water temperature,
sunlight, and the presence of cell pigments and bacteria. Half-lives of 11 to 15 days and up to
8 weeks have been reported for cylindrospermopsin in surface waters.
This document for microcystins and cylindrospermopsin focuses on the human health
risks associated with recreational exposures in waters containing these cyanotoxins. Exposure to
cyanobacteria and their toxins can also occur through non-recreational pathways such as
consumption of cyanotoxin-contaminated drinking water and food (including fish), and during
bathing or showering. The non-recreational exposures were not quantified in the recreational
exposure scenario described herein. Given that cyanobacterial blooms typically are seasonal
events, recreational exposures are likely to be episodic, and may be short-term in nature.
U.S. Environmental Protection Agency (EPA) is publishing these recommended values
for microcystins and cylindrospermopsin under Clean Water Act (CWA) 304(a) for states to
consider as the basis for swimming advisories for notification purposes in recreational waters to
protect the public. Additionally, states may consider using these same values when adopting new
or revised water quality standards (WQS). If adopted as WQS and approved by EPA under CWA
303(c), the WQS could be used for all CWA purposes. States may also wish to consider using
these values as both swimming advisory values and/or WQS. EPA envisions that if states decide
to use the values as swimming advisory values they would do so in a manner similar to their
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current recreational water advisory programs. The recommended values for use as swimming
advisories and/or WQS leverage the information collected and evaluated in EPA's Drinking
Water Health Advisory for the Cyanobacterial Microcystin Toxins and Drinking Water Health
Advisory for the Cyanobacterial Toxin Cylindrospermopsin (Drinking Water Health Advisories)
for these cyanotoxins.
At this time, available data are insufficient to develop quantitative recreational values for
cyanobacterial cell density related to inflammatory health endpoints. The reported
epidemiological relationships in the literature are not consistent for specific health outcomes
(e.g., dermal symptoms, eye/ear irritation, fever, gastrointestinal (GI) illness, and respiratory
symptoms) or for those health outcomes associated with specific cyanobacterial cell densities.
The uncertainties related to the epidemiological study differences, such as study size, species and
strains of cyanobacteria present, and the cyanobacterial cell densities associated with significant
health effects, do not provide sufficient information to determine a consistent association
between cyanobacterial densities associated with adverse inflammatory health effects.
EPA evaluated the health effects of microcystins and derived a Reference Dose (RfD) in
its 2015 Health Effects Support Document for the Cyanobacterial Toxin Microcystins. Exposure
to higher-levels of microcystins can lead to liver damage and renal failure. The critical study for
the derivation of the microcystins RfD was conducted by Heinze et al. (1999) based on rat
exposure to microcystin-LR in drinking water. The critical effect from this study was liver
damage, including increased liver weight, slight to moderate liver lesions with hemorrhages, and
increased liver enzyme levels. EPA established an RfD for microcystin-LR and used it as a
surrogate for other microcystin congeners.
EPA evaluated the health effects of cylindrospermopsin and derived an RfD in its 2015
Health Effects Support Document for the Cyanobacterial Toxin Cylindrospermopsin. The
kidneys and liver appear to be the primary target organs for cylindrospermopsin toxicity. The
critical study for the derivation of the cylindrospermopsin RfD was conducted by Humpage and
Falconer (2002, 2003) based on drinking water exposure to mice. The critical effect was kidney
damage, including increased kidney weight and decreased urinary protein.
Based on available noncancer health effects information, EPA is recommending values
protective of primary contact recreation for two cyanotoxins as follows:
• For microcystins, the recreational value is 4 micrograms (|ig)/liter (L).
• For cylindrospermopsin, the recreational value is 8 |ig/L.
These values are based on overall exposure to children at the 90th percentile. If used as a
swimming advisory to protect swimmers at a beach, these values are not to be exceeded on any
single day. If used as a water quality criterion for assessment and listing purposes, EPA
recommends that states consider the number of exceedances of no more than 10 percent of days
per recreational season up to one year. These criteria are based on noncancer health effects
because EPA concluded in its Health Effects Support Documents (HESDs) for microcystins and
cylindrospermopsin that there is inadequate information to assess carcinogenic potential of these
cyanotoxins (U.S. EPA 2015c; U.S. EPA 2015d). Should additional information become
available in the future, EPA can review and revise these recommendations, as appropriate.
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2.0 INTRODUCTION AND BACKGROUND
This section provides background information about cyanobacteria and cyanobacterial
blooms, the source of the stressors, microcystins and cylindrospermopsin. It discusses briefly the
occurrence of cyanobacterial blooms and these cyanotoxins in the United States. Section 2.1
describes Clean Water Act provisions relevant to these recreational ambient water quality criteria
or swimming advisories. Section 2.2 summarizes international and state recreational water
guidelines for microcystins, cylindrospermopsin, and cyanobacteria to provide context regarding
how other jurisdictions are addressing the human health concerns.
Cyanobacteria are a group of microorganisms that naturally occur in freshwater and
marine environments and can be found at higher densities in eutrophic or nutrient-enriched water
bodies. Many cyanobacteria are capable of producing toxins, generally referred to as
cyanotoxins, which can impact human health. Under the right conditions of temperature, light,
pH, nutrient availability, etc., cyanobacteria can reproduce rapidly to high densities in water,
forming what are commonly referred to as cyanobacterial harmful algal blooms (HABs). Other
microorganisms can form HABs, but for the purpose of this document, which addresses
cyanotoxins, usage of "HABs" will be in reference to cyanobacterial HABs unless otherwise
specified. A variety of factors can influence both cyanobacteria proliferation and toxin
production, including nutrient (e.g., nitrogen and phosphorus) concentrations, light levels,
temperature, pH, oxidative stressors, and interactions with other biota (viruses, bacteria, and
animal grazers), and others, as well as their combined effects (Paerl & Otten 2013 a; Paerl &
Otten 2013b).
Because they are a natural part of algal communities, cyanobacteria are commonly
observed in freshwater systems. The occurrence of HABs has been documented in surface waters
of all 50 states as well as U.S. territories between 2006 and 2015 as shown in Figure 2-1 (Richlen
2016; WHOI2016). Figure 2-1 also identifies areas where more widespread HAB problems have
occurred, e.g., parts of the Great Lakes.
In 2007, the EPA's National Lakes Assessment (NLA) conducted a national probability-
based survey of the nation's lakes, ponds, and reservoirs (Loftin et al. 2016b; U.S. EPA 2009).
These surveys covered a total of 1,028 lakes, which represented nearly 50,000 lakes larger than
4 hectares (10 acres) in the conterminous United States. This assessment found that
cyanobacteria were detected in almost all lakes (U.S. EPA 2009). Cyanobacteria were the
dominant member of the phytoplankton community in 76 percent of lake samples. Subsequent
analysis indicated that potential microcystin- and cylindrospermopsin- producing species
occurred in 95 and 67 percent of samples, respectively (Loftin et al. 2016b).
Microcystins are the most commonly detected class of cyanotoxin and have been found
in lakes in the contiguous United States (U.S. EPA 2009) and streams in the Southeastern United
States (Loftin et al. 2016a). The NLA 2007 reported that 30 percent of lakes in the conterminous
United States had detectable microcystin. In a separate study, Graham et al. (2010) sampled
cyanobacterial blooms in 23 Midwestern lakes and detected microcystins in all blooms sampled.
The researchers also found that cylindrospermopsin co-occurred with microcystins in 9 percent
of samples (Graham et al. 2010). In an expanded analysis of NLA samples, Loftin et al. (2016b)
identified cylindrospermopsin in 4 percent of samples with a mean concentration of 0.56 |ig/L.
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Figure 2-1. Generalized Distribution of Cyanobacterial HABs in the United States and
Territories11
Graphic adapted from a Woods Hole Oceanographic Institute (WHOI) map of HABs that occurred between 2006
and 2015. It reflects input fromHAB experts with broad experience in HAB events and reports to the U.S.
National Office for Harmful Algal Blooms (Richlen 2016; WHOI 2016). Each state that has experienced one or
more cyanobacterial HABs is indicated with a single green dot. Larger green ovals mark areas where more
widespread cyanobacterial HAB problems occurred.
Exposure to HABs can result in adverse human health outcomes such as gastrointestinal,
dermatologic, respiratory, neurologic, and other symptoms. The Centers for Disease Control and
Prevention (CDC) collected information on outbreaks of illness related to HABs reported via the
National Outbreak Reporting System (NORS) and the Harmful Algal Bloom-Related Illness
Surveillance System (HABISS). During 2009 and 2010 in the United States, 11 waterborne
disease outbreaks associated with HABs were reported to CDC, all occurring in freshwater lakes:
eight of these investigations evaluated the presence of cyanotoxins; eight detected microcystins;
and two detected cylindrospermopsin (Hilborn et al. 2014). The 11 outbreaks associated with
HABs affected at least 61 persons, resulting in 2 hospitalizations; 66 percent of the affected
persons overall were aged 18 or younger, and 35 percent were aged 9 years or younger. Hilborn
et al. (2014) reported that microcystins were present during all eight outbreak investigations in
which cyanotoxin testing was performed. In four of the outbreaks, microcystin concentrations
exceeded 20 (J,g/L. During investigations of these outbreaks, cylindrospermopsin, and anatoxin-a
also were detected. The researchers concluded that the disease outbreak data suggest that the
time to onset of effects might be rapid, that children might be at higher risk for illness, and that
HAB-associated outbreaks occur during the warmer months. Hilborn et al. (2014) noted that
recognizing HAB-associated illness from recreational exposure might be underreported due to
multiple possible exposure routes and the non-specific nature of potential health effects. In
addition, Graham et al. (2009) counted 36 states with anecdotal reports of acute cyanotoxin
poisonings of animals, humans, or both as reported in journal articles and newspaper articles
(Chorus & Bartram 1999; Huisman et al. 2005; Yoo et al. 1995).
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As a result of potential adverse health effects associated with recreational exposure to
HABs, many states have developed guidelines and/or health advisories related to HABs. For the
summer of 2008, Graham et al. (2009) identified at least 13 states that posted recreational health
advisories because the concentrations of cyanobacteria or cyanotoxins were large enough to be
considered a risk to animals and people by the state (Graham et al. 2009). These included
cautions, warnings, public health advisories, and public health warnings, due to the presence of
cyanobacteria, cyanotoxins, or both.
Figure 2-2 shows the number of 2016 freshwater HAB recreational advi sories states
publicly reported in each EPA region between January 1 and August 12, 2016. To develop this
regional summary map, EPA researched and compiled publically available reports posted on
states' websites between these dates. During that time, states reported at least 255 notices for
freshwater HABs with reported microcystins concentrations ranging from not detected (i.e.,
below the limit of detection) to 392 |ig/L. These notices included cautions, warnings, public
health advisories, and public health warnings due to the presence of cyanobacteria, cyanotoxins,
or both. Advisories can last for multiple days. The review was not exhaustive and might not
reflect all of the monitoring, beach, or general health advisories (e.g., some advisories at local or
county-level may not be posted on the state website). Thus, the number of actual HAB advisories
during this time might be higher. In addition, many states have only recently begun to monitor
HABs, thus monitoring may be limited.
Figure 2-2. State-reported HAB Advisories by EPA Region, January 1 to August 12, 2016
21
The presence of detectable concentrations of cyanotoxins in the environment is closely
associated with HAB occurrences. Cyanotoxin concentrations in surface waters can be higher
after the initial bloom fades, so potential exists for human exposures even after the visible signs
of a bloom are gone. Thus, high densities of cyanobacteria and high cyanotoxin concentrations
are capable of affecting the health of humans, domestic animals, and wildlife in contact with
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affected waters. These events are not always independent; animal health effects associated with
harmful cyanobacteria have served as sentinel events to warn of potential human health risks
(Hilborn & Beasley 2015). Cyanotoxin production by cyanobacteria can vary spatially and
temporally, and studies of the impacts of environmental factors on cyanotoxin production are
ongoing.
Nutrients are key environmental drivers that influence the proportion of cyanobacteria in
the phytoplankton community, the cyanobacterial biovolume, cyanotoxin production, and the
impact that cyanobacteria may have on ecosystem function and water quality (Paerl et al. 2011).
Cyanobacteria production and cyanotoxin concentrations are dependent on nutrient levels (Wang
et al. 2002); however, nutrient uptake rates and the utilization of organic and inorganic nutrient
forms of nitrogen and phosphorus vary considerably by cyanobacteria species. In addition to
nutrient concentrations, factors such as the nitrogen:phosphorus ratio and organic matter
availability, as well as other physico-chemical processes, can play a role in determining HAB
composition and cyanotoxin production (Paerl & Huisman 2008; Paerl & Otten 2013b).
2.1 Clean Water Act
Section 304(a) of the Clean Water Act (CWA) requires the Administrator of EPA 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.
EPA is publishing these recommended values under CWA 304(a) for states to consider as
the basis for swimming advisories for notification purposes in recreational waters to protect the
public. Additionally, states may consider using these same values as criteria when adopting new
or revised WQS. If adopted as WQS and approved by EPA under CWA 303(c), the WQS could
be used for CWA purposes. States may also wish to consider using these values as both
swimming advisory values and WQS. EPA envisions that if states decide to use the values as
swimming advisory values they might do so in a manner similar to their current recreational
water advisory programs.
This document recommends values for cyanotoxins that are protective of human health
given a primary contact recreational exposure scenario. The cyanotoxins included in this
document have been demonstrated to occur in nutrient-enriched waters affected by
cyanobacterial HABs.
The term "water quality criteria" is used in two sections of the CWA§304(a)(l) and
§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 effects on human health or aquatic life. The
criteria recommendations presented in this document are such a scientific assessment. If water
quality criteria associated with specific designated uses are adopted by a state or authorized tribe
as water quality standards under section 303, and approved by EPA, they become applicable
CWA water quality standards in ambient waters within that state or tribe. Water quality criteria
adopted in state or tribal water quality standards could have the same numerical values as criteria
developed under section 304. Alternatively, states and authorized tribes may derive numeric
criteria based on other scientifically defensible methods, but the criteria must be protective of
designated uses. It is not until their adoption as part of state or tribal water quality standards, and
subsequent approval by EPA, that criteria become CWA applicable water quality standards.
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Guidelines to assist in modifying the criteria recommendations presented in this document are
contained in the Water Quality Standards Handbook (U.S. EPA 2012b). This handbook and
additional guidance on the development of WQS and other water-related programs of this
Agency have been developed by EPA, which along with 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.
2.2 International and State Guidelines
In 2003, World Health Organization (WHO) derived a series of guideline values for
recreational exposure to cyanobacteria associated with incremental severity and probability of
adverse health effects (WHO 2003b); see Table 2-1. For these guidelines, WHO recommended
values that included the potential health effects from exposure to cyanobacteria because it was
"unclear whether all important cyanotoxins had been identified and that the health outcomes
observed after recreational exposure could be related to cyanobacterial substances other than the
well-known cyanotoxins (WHO 2003b)." They also considered the potential for liver damage by
microcystins. WHO highlighted that there are multiple potential health endpoints related to
recreational exposure to cyanobacteria and their toxins and developed a series of guidelines
associated with incremental severity and probability of health effects at increasing densities of
cyanobacteria and corresponding concentrations of chlorophyll a (if cyanobacteria dominate).
The different levels were an effort to distinguish between irritative or inflammatory-response
symptoms associated with cyanobacterial cells and the more severe hazard of exposure to
elevated concentrations of cyanotoxins, particularly microcystins. The WHO guidelines combine
the potential for both sets of endpoints (i.e., cyanotoxins and cyanobacterial cells) across three
categories of increasing probability of risk. The cell-associated inflammatory responses are
represented by the low probability of adverse health effects category of < 20,000 cells/milliliter
(mL), corresponding to less than 10 |ig/L chlorophyll a if cyanobacteria dominate and estimated
microcystin levels of less than 10 |ig/L. According to the WHO, as the density of cyanobacteria
increase above that level, the probability of inflammatory responses increases, and the potential
for more severe adverse health effects associated with exposure to the cyanotoxins also
increases. The high-risk category identified by WHO related > 100,000 cells/mL (corresponding
to 50 |ig/L of chlorophyll a, if cyanobacteria dominate) and > 20 |ig/L microcystin levels, was
primarily due to the toxic effects of microcystins.
Table 2-1. WHO (2003b) Recreational Guidance/Action Levels for Cyanobacteria,
Chlorophyll a, and Microcystin
Relative Probability of
Aeutc Health Effects
Cyanobacteria
(cells/mL)
Chlorophyll a (jig/L)
Estimated
Microcystin Levels
(jig/L)a
Low
< 20,000
< 10
< 10
Moderate
20,000-100,000
10-50
10-20
High
>100,000-10,000,000
50-5,000
20-2,000
Very High
> 10,000,000
> 5,000
> 2,000
" WHO (2003b) derived the microcystin concentrations from the cyanobacterial cell density levels.
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The WHO guideline value development was supported by results from a review
conducted by Chorus and Bartram (1999). A primary study identified in this review was a
prospective epidemiological study by Pilotto et al. (1997), which evaluated health effects after
recreational exposure to cyanobacteria and reported associations between cyanobacterial cell
densities and health. Pilotto et al. (1997) found a significant association among recreators
exposed to > 5,000 cells/mL for > 1 hour and one or more symptoms and similar significant
associations for symptoms in people exposed to 5,000-20,000 cells/mL. WHO chose a guideline
level of 20,000 cells/mL to represent the upper bound of the low probability of adverse health
effects category (WHO 2003b). The low category includes irritative or inflammatory health
effects associated with exposure to cyanobacterial cells (WHO 2003b). While the association
among recreators exposed to > 5,000 cells/mL for > 1 hour and one or more symptoms reported
in Pilotto et al. (1997) were statistically significant, WHO states that they represented less than
30 percent of the individuals exposed (Chorus & Bartram 1999). Therefore, the level of health
effect and the small number of people affected at 5,000 cells/mL were not considered by WHO
to be a basis to justify action (WHO 2003b).
WHO (2003b) also made the connection between cyanobacterial cell densities and
microcystin concentrations. It assumed microcystin-producing cyanobacteria dominate the
population of cyanobacteria present and that the average microcystin content of Microcystis sp.
cells averages 0.2 pg/cell. Thus, WHO estimated that 20,000 cells/mL could potentially equate to
approximately 2-4 |j,g/L of microcystin. Similarly, using the same assumptions at a
cyanobacterial cell density of 100,000 cells/mL, they estimated approximately 20 |_ig/L. WHO
pointed out that the potential concentration of microcystins could vary based on the composition
of the community of cyanobacteria present. WHO states that, at the same cyanobacterial cell
density, cyanotoxin levels may approximately double if Planktothrix agardhii is the dominant
member of the community. Several states have adopted the estimated microcystins
concentrations as their guideline values rather than the cell density or chlorophyll a values from
the WHO guidelines, as discussed later in this section.
Many countries have adopted the WHO recommendations for recreational waters
including multiple parameters (e.g., cell density, biovolume, and cyanotoxin concentration).
Table 2-2 provides international recreational water guideline or action levels for cyanotoxins or
cyanobacteria for several countries. Table 2-2 lists the lowest concentrations of cyanotoxins or
densities of cyanobacteria that prompt a health protective action. For a more complete list of
guideline or action levels and recommended actions for international jurisdictions, see Appendix
A. EPA did not identify any recreational guideline levels for cylindrospermopsin established by
other international regulatory authorities. Some international authorities have multiple action
levels; for brevity, Table 2-2 that follows presents the guideline reflecting the lowest
concentration of microcystin or density of cyanobacterial cells that recommended or triggered a
health protective action. More details are in Appendix A.
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Table 2-2. International Recreational Water Guideline or Action Levels for Cyanobacteria
and Microcystins
Jurisdiction
Lowest Recreational Water Guideline/Action Level"
Reference
Australia13
microcystins (total): >10 |ig/L
or Microcystis aeruginosa (total): > 500 to < 5,000 cells/mL
or cyanobacteria (total): > 0.4 to < 4 mm3/L (where a known toxin
producer is dominant in the total biovolume)
Australian Government
National Health and Medical
Research Council (2008)
Canada
microcystins (total): > 20 |ig/L (expressed as microcystin-LR)
or cyanobacteria (total): > 100,000 cells/mL
Health Canada (2012)
Cuba
cyanobacteria: > 1 of the species known as potentially toxic
or phytoplankton cells: > 20,000 - to < 100,000 cells/mL,
> 50 percent of cells cyanobacteria
German Federal Environment
Agency (2012)°
Czech Republic
cells: >20,000 cells/mL
German Federal Environment
Agency (2012)°
Denmark
chlorophyll a: >50 |ig/L, dominated by cyanobacteria
or visible surface scum
German Federal Environment
Agency (2012)°
European Union
Appropriate monitoring must be implemented if there is a risk of
proliferation of algae. Member state authorities responsible must
take management measures and provide information immediately
if a proliferation of cyanobacteria (or blue algae) occurs.
European Parliament and the
Council of the European
Union (2006)
Finland
algae (includes cyanobacteria): detected
German Federal Environment
Agency (2012)°
Franceb
microcystins: > 25 |ig/L
or cyanobacteria: > 20,000 to < 100,000 cells/mL (±20 percent)
German Federal
Environment Agency
(2012)°
Germany
Secchi Disk reading > 1 m AND
(microcystins: >10 |ig/L
or chlorophyll a (with dominance by cyanobacteria): > 40 |ig/L
or biovolume: > lmm3/L)
German Federal
Environment Agency
(2012)°
Hungary
microcystins: > 4 to < 10 |ig/L
or cell count: > 20,000 to < 50,000 cells/mL
or chlorophyll a (with dominance by cyanobacteria): > 10 to
< 25 (ig/L
German Federal
Environment Agency
(2012)°
Italyb
microcystins: > 25 |ig/L
or cyanobacterial cell count (combined with identification of genus
and, if possible, species): > 20,000 cells/mL
German Federal
Environment Agency
(2012)°
Netherlands
chlorophyll a: >12.5 to < 75 |ig/L
or biovolume (cyanobacterial cell count): >2.5 to < 15
mm3/L
German Federal
Environment Agency
(2012)°
New Zealand15
microcystins (total): >12 |ig/L
or cyanobacteria (benthic): 20-50 percent coverage of potentially
toxigenic cyanobacteria attached to substrate
or cyanobacteria (total): > 0.5 to < 1.8 mm3/L (biovolume
equivalent of potentially toxic cyanobacteria)
or cyanobacteria (total): > 0.5 to < 10 mm3/L (biovolume equivalent
of the combined total of all cyanobacteria)
Wood et al. (2008)
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Jurisdiction
Lowest Recreational Water Guideline/Action Level"
Reference
Poland
visible blooms
German Federal Environment
Agency (2012)°
Scotland13
chlorophyll a: > 10 |ig/L with dominance of cyanobacteria
or cyanobacteria: > 20,000 cells/mL
Scottish Government Health
and Social Care Directorates
Blue-Green Algae Working
Group (2012)
Spain
cyanobacteria proliferation potential (Low)
German Federal
Environment Agency
(2012)°
Turkey
microcystin-LR: > 25 |ig/L equivalents
or cells: > 20,000 to 100,000 cells/mL
German Federal
Environment Agency
(2012)°
World Health
Organization
(WHO)
cyanobacteria: 20,000 cells/mL
or chlorophyll a: 10 |ig/L (approximately 2-4 |ig microcystin/L.
assuming cyanobacteria dominance)
Chorus and Bartram (1999);
WHO (2003b)
a More details are provided in Appendix A.
b The lowest guideline values for each quantitative parameter (i.e., cyanobacterial cell density, biovolume,
cyanotoxin concentration) are not associated with the same action level. For example, for Australia, the lowest
cyanobacterial cell density and biovolume criteria trigger the green level surveillance mode, and the lowest
cyanotoxin concentration triggered the red level action mode.
0 Following the VIIIth International Conference on Toxic Cyanobacteria, the German Federal Environmental
Agency compiled and published in 2012 regulatory approaches to the assessment and management of cyanotoxin
risks based on contributions by member countries.
Approximately 30 U.S. states have implemented cyanobacterial HAB guidelines for
recreational waterways as of November 2015. As shown in Figure 2-3, five states had
quantitative cyanotoxin guidelines only, and fourteen states had quantitative guidelines for
cyanotoxins, as well as either quantitative or qualitative guidelines for cyanobacterial cell
density. Generally, qualitative guidelines use visual inspection and not quantitative detection
methods. In addition, twelve states had quantitative guidelines for cyanobacterial cell density
only or had qualitative guideline values only.
For brevity, Table 2-3 lists the lowest recreational water guideline or action levels for
microcystins, cylindrospermopsin, or cyanobacteria that trigger or recommend a health
protective action for U.S. states. For a more complete list of state guideline/action levels and
recommended actions see Appendix B. Parameters and values used as the basis for guidelines
varied across states, as does the methodology for developing the values. Similar to international
authorities, many states used a tiered approach, which evaluates multiple parameters including
cyanobacterial cell density, chlorophyll a concentration, cyanotoxin concentration, and visual
appearance. New York, for example, considered all four of these parameters at lower tier
guideline levels, but only considered cyanotoxin concentrations at the highest advisory level.
Other states had only one response guideline level and only consider cyanotoxin concentration
(e.g., California) or had only one response guideline level, but considered cyanobacterial cell
density, cyanotoxin concentration, and visual appearance (e.g., Oregon). In contrast, other states,
like Connecticut, used a tiered approach and did not consider cyanotoxin concentrations at any
tier but rather consider visual inspection and cyanobacterial cell density.
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a Includes states with quantitative cyanotoxin guidelines as well as either quantitative or qualitative cyanobacteria
guidelines.
b Includes states that either have quantitative cyanobacteria guidelines only or qualitative guidelines only.
c EPA found that Texas and North Carolina published guidelines in the past, but the guidelines were no longer on
their websites.
Table 2-3. State Guideline or Action Levels for Microcystin, Cylindrospermopsin, and
Cyanobacterial Cells in Recreational Water
State
Lowest Recreational Water Guideline or Action
Level3
Reference
Arizona
blue-green algae (mean value based on a minimum of
two sample events within one peak season):
20,000 cells/mL
and
chlorophyll a result (mean value based on a minimum
of two sample events within one peak season) in target
range
Arizona Department of Enviromnental
Quality (2008)
California
microcystins: 0.8 |ig/L
Butler et al. (2012)
cylindrospermopsin: 4 ug/L
Colorado
microcystin-LR: > 10 ug/L and < 20 jig/L
Colorado Department of Public Health
& Environment (2016)
cylindrospermopsin: > 7 (.ig/L
Connecticut
visual rank category 2: cyanobacteria present in low
numbers; there are visible small accumulations but
Connecticut Department of Public
Health: Connecticut Energy
Figure 2-3. State Guidelines for Cyanotoxins and Cyanobacteria in Recreational Water by
Type and Scope of Guidelines
Cyanotoxin and cyanobacteria guidelines^0
n Cyanobacteria guidelines onlybc
Cyanotoxin guidelines only
~ No cyanobacteria or cyanotoxin guidelines
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State
Lowest Recreational Water Guideline or Action
Level3
Reference
water is generally clear; OR blue-green algae cells
> 20,000 cells/mL and < 100,000 cells/mL
Environment (2013)
Delaware
thick green, white, or red scum on surface of pond
Delaware Department of Natural
Resources and Environmental Control:
Division of Water (2016)
Florida
cyanobacteria bloom
Florida Department of Environmental
Protection (2016); Florida Department
of Health (2016)
Idaho
Microcystis or Planktothrix: > 40,000 cells/mL
IDEQ (2015)
sum of all potentially toxigenic taxa: > 100,000
cells/mL
Illinois
microcystin-LR: > 10 |ig/L
Illinois Environmental Protection
Agency (2015)
Indiana
blue-green algae: 100,000 cells/mL
Indiana Department of Environmental
Management (2015)
microcystin-LR: 6 |ig/L
cylindrospermopsin: 5 |ig/L
Iowa
microcystin: > 20 |ig/L
Iowa Environmental Council (2015)
Kansas
cyanobacteria: > 80,000 and < 250,000 cells/mL
Kansas Department of Health &
Environment (2015)
microcystin: > 4 and < 20 |ig/L
Kentucky
blue-green algae: > 100,000 cells/mL
Kentucky Department for
Environmental Protection (2014)
microcystins: > 20 |ig/L
Commonwealth of Kentucky:
Department for Environmental
Protection Division of Water (2015)
Maine
Secchi disk reading < 2 meters caused by algae
Maine Department of Environmental
Protection (2013)
Maryland
Microcystis aeruginosa or other potential microcystin
producing blue green algae > 40,000 cells/mL, and
samples contain microcystins: > 10 ppb
Maryland Department of Natural
Resources (2010)
Massachusetts
blue-green algae: > 50,000 cells/mL
Massachusetts Bureau of Environmental
Health (2015)
microcystins: >14 |ig/L
Montana
reservoirs that seem stagnated and harbor large
quantities of algae
State of Montana Newsroom (2015)
Nebraska
microcystin: > 20 |ig/L
Nebraska Department of Environmental
Quality and Nebraska Department of
Health and Human Services: Division of
Public Health (2016)
New Hampshire
cyanobacteria: > 50 percent of total cell counts from
toxigenic cyanobacteria
New Hampshire Department of
Environmental Services (2014)
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State
Lowest Recreational Water Guideline or Action
Level3
Reference
New York
bloom: credible report or digital imagery of a bloom
determined as likely to be potentially toxic
cyanobacteria by DEC or DOH staff
Gorney (2016)
blue green chlorophyll a: > 25-30 |ig/L
potential toxin-producing cyanobacteria taxa: >
50 percent of algae present
microcystin-LR: 4 |ig/L
North Carolina
visible discoloration or surface scum
North Carolina Health and Human
Services: Division of Public Health
(2014)
North Dakota
blue-green algae bloom is present over a significant
portion of the lake AND
microcystin-LR: > 10 |ig/L
North Dakota Department of Health:
Division of Water Quality (2016)
Ohio
microcystin-LR: 6 |ig/L
Kasichetal. (2015)
cylindrospermopsin: 5 |ig/L
Oklahoma
cyanobacteria: 100,000 cell/mL
Oklahoma Legislature (2012)
microcystin: > 20 |ig/L
Oregon
cylindrospermopsin: > 20 |ig/L
Oregon Health Authority (2016)
microcystin: >10 |ig/L
Microcystis: > 40,000 cells/mL
Planktothrix: > 40,000 cells/mL
toxigenic species: > 100,000 cells/mL
visible scum with documentation and testing
Rhode Island
cyanobacteria: > 70,000 cells/mL
Rhode Island Department of
Environmental Management and Rhode
Island Department of Health (2013)
microcystin-LR: > 14 (ig/L
visible cyanobacteria scum or mat
Texas
> 100,000 cell/mL of cyanobacteria cell counts and
> 20|ig/L microcystin
U.S. EPA (2016)
Utah
blue-green algae: 20,000-100,000 cells/mL
Utah Department of Environmental
Quality and Department of Health
(2015)
microcystin: 4-20 |ig/L
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State
Lowest Recreational Water Guideline or Action
Level3
Reference
Vermont
cylindrospermopsin: > 10 |ig/L
Vermont Department of Health (2015)
microcystin-LR (equivalents): > 6 |ig/L
visible known blue-green algae bloom/scum or an
unknown, potentially blue-green algae (i.e., not pollen),
bloom/scum
Virginia
blue-green algal "scum" or "mats" on water surface
Virginia Department of Health (2012)
microcystin: > 6 |ig/L
Microcystis: 5,000 to < 20,000 cells/mL
Washington
bloom is forming or a bloom scum is visible (toxic
algae may be present); cyanotoxin levels do not exceed
thresholds
Hardy and Washington State
Department of Health (2011)
microcystins: 6 |ig/L
cylindrospermopsin: 4.5 |ig/L
West Virginia
blue-green algal blooms observed and monitored
West Virginia Department of Health &
Human Resources (2015)
Wisconsin
cyanobacteria: > 100,000 cells/mL
Wisconsin Department of Natural
Resources (2012)
visible scum layer
Werner and Masnado (2014)
a More details are provided in Appendix B.
Among states that consider the same parameters, there is considerable variation in
guideline levels and associated responses. As shown in Table 2-3, the state recreational
guidelines featuring the lowest microcystins or cylindrospermopsin concentrations that
recommended or triggered a health protective action ranged from 0.8 to 20 [j,g/L and 4 to
20 (J,g/L, respectively. The guidelines reflecting the lowest densities of cyanobacterial cells that
triggered a health protective action ranged from 5,000 to 100,000 cells/mL. Some of the
variation in guideline levels is attributable to variations in exposure parameters, as well as
variations in the basis for guideline values. Ten states base at least one guideline value on either
WHO guidance or a modified version of WHO guidance (e.g., Indiana, Oregon, Utah). Eleven
states, including California, Massachusetts, and Ohio, base at least one guideline value on
jurisdiction-specific risk assessments or monitoring information, or studies or guidelines other
than those from WHO. For more information on individual state guidelines, see Appendix B.
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3.0
NATURE OF I II I STRESSORS
This section discusses cyanobacteria and cyanobacterial blooms that have the potential to
produce microcystins and cylindrospermopsin. It also describes these toxins' chemical and
physical properties, sources and occurrence information in different media, environmental fate,
and toxicokinetics.
3.1 Cyanobacteria and Cyanobacterial Blooms
Cyanobacteria are photosynthetic prokaryotes (Seckbach & Oren 2007) and are
ubiquitous in the environment. The chloroplast, found in photosynthetic eukaryotes like algae
and plants, evolved from an endosymbiotic relationship with cyanobacteria (Kutschera & Niklas
2005). Ecologists historically grouped cyanobacteria, often referred to as "blue-green algae,"
with eukaryotic algae because they contain chlorophyll a and their ability to perform oxygenic
photosynthesis. However, cyanobacteria are prokaryotes (i.e., no discrete membrane-bound
nucleus or membrane-bound subcellular organelles) and are genetically related to other bacteria
in the eubacteria domain. Taxonomically, they are classified in the phylum Cyanobacteria or
Cyanophyceae (Carmichael 2008; O'Neil et al. 2012).
Cyanobacteria can produce bioactive compounds including toxins, which may be
harmful. These biomolecules include hepatotoxic, neurotoxic, and cytotoxic compounds and
compounds that can result in allergic reactions (Carmichael 1994; Jaiswal et al. 2008; Volk &
Mundt 2007). Studies have also shown that exposure to cyanobacterial cells independent of
cyanotoxins can cause health effects; this information is detailed in Appendix D.
Members of Microcystis, Dolichospermum (Anabaena), Nostoc, Oscillatoria,
Fischerella, Planktothrix, and Gloeotrichia can produce microcystins (Carey et al. 2012b; Codd
et al. 2005; Duy et al. 2000; Stewart et al. 2006c). Microcystis sp. have been documented to
occur in blooms on all continents except Antarctica and often dominate phytoplankton
assemblages in the summer (O'Neil et al. 2012). Along the margins of Antarctica, other genera
of cyanobacteria occur in exposed soils, glaciers, ice shelves, frozen lakes, and stream beds,
including Nos/oc, Oscillatoria, Lyngbya, or Synechococcus (Paerl et al. 2016; Vincent 2007).
Microcystis sp. have been documented throughout the United States (Carmichael 2001; Jacoby et
al. 2000).
Several environmental factors, including nutrient load, increased water temperature,
salinity, pH, light intensity, and reduced mixing, provide competitive advantages to Microcystis
relative to other phytoplankton (Jacoby et al. 2000; Marmen et al. 2016). There is evidence that
these environmental factors also affect the relative abundance of microcystin-producing strains
and non-microcystin-producing strains (Marmen et al. 2016). Microcystis thrives in warmer
temperatures, with optimal growth and photosynthesis occurring above 25°C (O'Neil et al.
2012). A Japanese study between May and November 2006 found that the toxin-producing
species, M. aeruginosa, dominated in months with relatively higher water temperatures, while
the non-toxin-producing species, M. wesenbergii, dominated in months with lower water
temperatures (Imai et al. 2009). Elevated nitrogen and phosphorus inputs may both stimulate
Microcystis cell growth and biomass accumulation, and can favor microcystin-producing strains
(Marmen et al. 2016; O'Neil et al. 2012). During the summer of 1994, M. aeruginosa was
observed as the dominant species in a toxic bloom in Washington, associated with elevated
nitrogen inputs resulting in low nitrogen to phosphorus ratios (Jacoby et al. 2000). The genetic
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composition of the bloom can also influence the degree of toxicity associated with an algal
bloom. Lee et al. (2015) found that, although Microcystis sp. was rarely detected in a shallow
lake bloom, most of this population contained the toxin-producing gene. They observed
intracellular microcystins at concentrations two to three orders of magnitude greater than
extracellular microcystins (Lee et al. 2015).
A number of cyanobacterial species including Cylindrospermopsis raciborskii (C.
raciborskii), Aphanizomenon flos-aquae, Aphanizomenon gracile, Aphanizomenon ovalisporum,
Umezakia natans, Anabaena bergii, Anabaena lapponica, Anabaena planctonica, Lyngbya
wollei, Rhaphidiopsis curvata, and Rhaphidiopsis mediterranea can produce cylindrospermopsin
(B-Beres et al. 2015; Kokocinski et al. 2013; McGregor et al. 2011; Moreira et al. 2013).
Cylindrospermopsin-producing cyanobacteria occur in tropical or subtropical regions, but also
have been detected in warmer temperate regions. These species do not tend to form visible
surface scums and the highest concentrations of cyanobacterial cells occurs below the water
surface (Falconer 2005). C. raciborskii occurs in freshwater ponds, rivers, reservoirs and
eutrophic lakes and has been found in Australia, Asia, Europe, Africa and South, Central and
North America (Fuentes et al. 2010). According to a survey conducted in Florida in 1999 from
June to November, the most frequently observed toxigenic cyanobacteria were Microcystis
(43.1 percent), Cylindrospermopsis (39.5 percent), and Anabaena spp. (28.7 percent) (Burns
2008). In Florida, C. raciborskii was found to be the dominant cyanobacteria species in one lake
all year round (Burns 2008). In 2006, C. raciborskii was detected in lakes in southern Louisiana
(Fuentes et al. 2010). Conditions promoting its growth were shallow, warm surface water (over
30°C) and low light intensities. The highest concentrations of C. raciborskii were observed from
June through August with densities ranging from 37,000 cells/mL to more than 160,000
cells/mL. In a study of two lakes directly connected to Lake Michigan, Hong et al. (2006) found
low concentrations only in the late summer, and these were associated with elevated bottom
water temperatures and phosphorus concentrations.
Research indicates that cyanotoxins are associated with physiological functions of
cyanobacterial cell signaling, nutrient uptake, iron scavenging, maintenance of homeostasis, and
protection against oxidative stress and can confer a competitive advantage (Holland & Kinnear
2013). Cylindrospermopsin provides a competitive advantage to cyanobacteria when phosphorus
becomes scarce. Bar-Yosef et al. (2010) observed that, when phosphorus is scarce, the
cyanobacterium Aphanizomenon ovalisporum releases cylindrospermopsin, which causes other
microorganisms to release alkaline phosphatase, a compound which will increase available
phosphorus. Subsequently, Aphanizomenon can gain access to phosphorus made available by
other microorganisms while simultaneously conserving the energy and resources required to
express and excrete alkaline phosphatase (Bar-Yosef et al. 2010). The precise biological function
of microcystin has not been conclusively determined (Zurawell et al. 2005). Studies comparing
wild-types and mutants of a microcystin species, examining the genes involved in microcystin
biosynthesis, and evaluating Microcystis colony size have suggested that microcystins play
important physiological roles in cyanobacteria, including colony formation (Kaplan et al. 2012;
Zurawell et al. 2005). Although cyanotoxins can negatively affect humans and other animals,
research suggests that the primary functions of cyanotoxins are in cyanobacterial physiology and
microbial ecology.
A variety of physical, chemical, and environmental factors affect the growth and
population dynamics of cyanobacteria, including light intensity, temperature, nutrient
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concentrations, biological interactions, and other environmental factors (see summary in Figure
3-1). When the rate of cyanobacterial cell growth exceeds the loss rate for a population,
positively buoyant, floating cyanobacterial cells can form a visibly colored scum on the water
surface, which can contain more than 10,000 cells/mL (Falconer 1998). The floating scum, as in
the case of Microcystis species, can be concentrated by prevailing winds in certain surface water
areas, especially at the shore. In larger freshwater bodies, such as Lake Erie, these areas of high
Microcystis concentrations are readily detected by satellite (Stumpf 2014; Wynne et al. 2010).
Although these blooms can occur naturally, increasing consensus among scientists is that these
blooms have been increasing in recent decades (Carmichael 2008; Flallegraeff 1993; Hudnell
2010).
Figure 3-1. Environmental Factors Influencing Cyanobacterial Bloom Potential in Aquatic
Ecosystems, Reproduced from Paerl and Otten (2013b) with Permission of Springer
Cyanos
Positive
1 High P (High N for some)
¦ Low N (DIN, DON) (only
applies to N2 fixers)
1 Low N:P Ratios
1 Low turbulence
¦ Low water flushing-Long
water residence time
¦ High (adequate) light
1 Warm temperatures
1 High dissolved organic
matter
1 Sufficient Fe (+ trace
metals)
¦ Low grazing rates
© Springer Science+Business
Media New York 2013
(/)
0
•*—>
CO
cr
co
CD
CC
Diversity
Modulating factors
Strong biogeochemical gradients (e.g.
persistent stratification, stable benthos)
Heterogeneous and diverse habitats (e.g.
reefs, seagrasses, marshes, sediments,
aggregates)
Selective grazing
'Toxin" production??
Negative
High DIN/total N (only
applies to N2 fixers)
Low P (DIP)
High N:P ratios
High turbulence & vertical
mixing
High water flushing-Short
water residence time
Low light (for most taxa)
Cool temperatures
Low dissolved organic
matter
Low Fe (+ trace metals)
High grazing rates
Viruses (cyanophages)
Predatory bacteria
Nutrients, particularly nutrient over-enrichment, are key environmental drivers that
influence the proportion of cyanobacteria in the phytoplankton community, the cyanobacterial
biovolume, cyanotoxin production, and the impact that cyanobacteria may have on ecosystem
function and water quality (Beaulieu et al. 2013; Paerl et al. 2011). Cyanobacterial toxin
concentrations are associated with nutrient levels (Wang et al. 2002); however, different
cyanobacteria species use organic and inorganic nutrient forms differently. Loading of nitrogen
and/or phosphorus to water bodies from agricultural, industrial, and urban sources influences the
development of cyanobacterial blooms and are associated with cyanotoxin production (Paerl et
al. 2011). Nitrogen loading can enhance the growth and cyanotoxin levels of Microcystis sp.
blooms and microcystin synthetase gene expression (Gobler et al. 2007; O'Neil et al. 2012).
Gobler et al. (2007) suggested that dominance of toxic Microcystis sp. blooms during summer is
linked to nitrogen loading, which stimulates growth and cyanotoxin synthesis. This may cause
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the inhibition of grazing by mesozooplankton and further accumulation of cyanobacterial cells.
Optimal concentrations of total and dissolved phosphorus (Wang et al. 2002) and soluble
phosphates and nitrates (ILS 2000; O'Neil et al. 2012; Paerl & Scott 2010; Wang et al. 2010)
may also result in the increased production of microcystins. Smith (1983) was the first to
describe a strong relationship between the relative amounts of nitrogen and phosphorus in
surface waters and toxic cyanobacterial blooms. Smith proposed that cyanobacteria should be
superior competitors under conditions of nitrogen-limitation because of their unique capacity for
nitrogen fixation, although many cyanobacteria like Microcystis species that produce toxins do
not fix nitrogen. While the dominance of nitrogen-fixing cyanobacteria at low nitrogen to
phosphorus ratios has been demonstrated in mesocosm- and ecosystem-scale experiments in
prairie and boreal lakes (Schindler et al. 2008), the hypothesis that low nitrogen to phosphorus
ratios favor cyanobacteria formation has been debated and challenged for its inability to reliably
predict cyanobacterial dominance (Downing et al. 2001). Eutrophic systems already subject to
bloom events are prone to further expansion of these blooms due to additional nitrogen inputs,
especially if these nutrients are available from internal sources. As the trophic state increases,
aquatic systems absorb higher concentrations of nitrogen (Paerl & Huisman 2008; Paerl & Otten
2013b). Recent surveys of cyanobacterial and algal productivity in response to nutrient pollution
across geographically diverse eutrophic lakes, reservoirs, estuarine and coastal waters, and in
different experimental enclosures of varying sizes demonstrate that greater stimulation is
routinely observed in response to both nitrogen and phosphorus additions. Further, this evidence
suggests that nutrient co-limitation is widespread (Elser et al. 2007; Lewis et al. 2011; Paerl et al.
2011). These results strongly suggest that reductions in nutrient pollution are needed to stem
eutrophication and cyanobacterial bloom expansion. For example, analysis of observational data
collected at larger spatial scales support the idea that controlling total phosphorus and total
nitrogen could reduce the frequency of high microcystin events by reducing the biomass of
cyanobacteria in the system (Orihel et al. 2012; Scott et al. 2013; Yuan et al. 2014).
The increasing body of laboratory and field data (Carey et al. 2012a; De Senerpont
Domis et al. 2007; Huisman et al. 2005; Jeppesen et al. 2009; Kosten et al. 2012; Reynolds 2006;
Wagner & Adrian 2009; Weyhenmeyer 2001) suggest that an increase in temperature may
influence cyanobacterial dominance in phytoplankton communities. Cyanobacteria may benefit
more from warming than other phytoplankton groups due to their higher optimum growth
temperatures. The optimum temperatures for microcystin production range from 20 to 25°C
(WHO 2003a). The increase in water column stability associated with higher temperatures also
may favor cyanobacteria (Carey et al. 2012a; Wagner & Adrian 2009). Kosten et al. (2012)
demonstrated that during the summer, the percentage of the total phytoplankton biovolume
attributable to cyanobacteria increased steeply with temperature in shallow lakes sampled along a
latitudinal transect ranging from subarctic Europe to southern South America. Furthermore,
warmer temperatures appear to favor the growth of toxigenic strains of Microcystis over non-
toxic ecotypes (Dziallas & Grossart 2011; Paerl & Otten 2013b). Indirectly, warming also may
increase nutrient concentrations by enhancing mineralization (Gudasz et al. 2010; Kosten et al.
2009; Kosten et al. 2010) by temperature- or anoxia-mediated sediment phosphorus release
(Jensen & Andersen 1992; S0ndergaard et al. 2003). Thus, temperature may indirectly increase
cyanobacterial biomass through its effect on nutrient concentrations. Others have suggested that
warmer conditions may raise total phytoplankton biomass through an alteration of top-down
regulation by selective grazing that favors larger size phytoplankton species and cyanobacterial
blooms (Jeppesen et al. 2009; Jeppesen et al. 2010; Teixeira-de Mello et al. 2009). The
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relationship between temperature and cyanobacterial dominance may be explained not only by
temperature effect on the competitive advantage of cyanobacteria, but also factors such as the
percent area covered and the volume of the lake taken up by submerged macrophytes (Carey et
al. 2012a; Kosten et al. 2012). Rising global temperatures and changing precipitation patterns
may stimulate cyanobacterial blooms. Warmer temperatures favor surface bloom-forming
cyanobacterial genera because they are heat-adapted, and their maximal growth rates occur at
relatively high temperatures, often in excess of 25°C (Reynolds 2006; Robarts & Zohary 1987).
At these elevated temperatures, cyanobacteria routinely out-compete eukaryotic algae (Elliott
2010; Paerl et al. 2011). Specifically, as the growth rates of the eukaryotic taxa decline in
response to warming, cyanobacterial growth rates reach their optima. Warmer surface waters,
especially in areas of reduced precipitation, are prone to intense vertical stratification. The
strength of vertical stratification depends on the density difference between the warm surface
layer and the underlying cold water, which is influenced by the amount of precipitation. As
temperatures rise due to climate change, stratification is expected to occur earlier in the spring
and persist longer into the fall (Paerl & Otten 2013b). The increase in water column stability
associated with higher temperatures and climate change may therefore favor cyanobacteria
production and possibly the prevalence of cyanotoxins such as microcystins (Carey et al. 2012a;
Wagner & Adrian 2009).
Sunlight availability and turbidity can have a strong influence on the cyanobacteria
species that predominate, as well as the depth at which they occur (Carey et al. 2012a; Falconer
2005). For example, Microcystis aeruginosa occurs mostly at the surface with higher light
intensities and in shallow lakes. Kosten et al. (2012) surveyed 143 shallow lakes along a
latitudinal gradient (between 5-55°S and 38-68°N) from subarctic Europe to southern South
America. Their analyses found a greater proportion of the total phytoplankton biovolume
attributable to cyanobacteria in lakes with high rates of light absorption. Kosten et al. (2012)
could not establish cause and effect from these field data, but other controlled experiments and
field data have demonstrated that light availability can affect the competitive balance among a
large group of shade-tolerant species of cyanobacteria, mainly Oscillatoriales and other
phytoplankton species (Scheffer et al. 1997; Smith 1986). Overall, results from Kosten et al.
(2012) suggest that higher temperatures interact with nutrient loading and underwater light
conditions in determining the proportion of cyanobacteria in the phytoplankton community in
shallow lakes.
Cyanobacterial blooms have been shown to intensify and persist at pH levels between six
and nine (WHO 2003a). When these blooms are massive or persist for a prolonged period, they
can become harmful. Kosten et al. (2012) noted the impact of pH on cyanobacteria abundance in
lakes along a latitudinal transect from Europe to southern South America. The percentage of
cyanobacteria in the 143 shallow lakes sampled was highly correlated with pH, with an increased
proportion of cyanobacteria at higher pH. Cyanobacteria have a competitive advantage over
other phytoplankton species because they are efficient users of carbon dioxide in water (Caraco
& Miller 1998; Shapiro 1984). This characteristic is especially advantageous for cyanobacteria
under conditions of higher pH when the concentration of carbon dioxide in the water column is
diminished due to photosynthetic activity. Although this could explain the positive correlation
observed between pH and the proportion of cyanobacteria, the high proportion of cyanobacteria
at high pH could be the result of an indirect nutrient effect as described previously (see
discussion in Temperature section). As photosynthesis intensifies, pH increases due to carbon
dioxide uptake by algae, resulting in a shift in the carbonic buffer equilibrium and a higher
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concentration of basic forms of carbonate. Thus, higher water column pH may be correlated with
a higher proportion of cyanobacteria because of higher photosynthetic rates, which can be linked
with high nutrient concentrations (Duy et al. 2000) that stimulate phytoplankton growth and
bloom formation. High iron concentrations (more than 100 |iM) have also been shown to
increase cyanobacterial cell density and chlorophyll content in Microcystis aeruginosa
(Kosakowska et al. 2007).
Cyanobacterial blooms commonly occur from spring to early fall in various regions of
the United States (Wynne & Stumpf 2015). Cyanobacteria take advantage of conditions that can
occur in late summer and early fall such as elevated water temperatures and increased vertical
stratification in lakes and reservoirs (Paerl & Huisman 2008). Some blooms occur later in
summer and early fall. Vertical biomass structure and cyanotoxin production can be influenced
by seasonal changes as well as severe weather conditions (e.g., strong wind or rainfall) and also
by runoff. At times, the hypolimnion (bottom layer of the water column) can have a higher
cyanobacteria biomass and display different population dynamics than the epilimnion (upper
layer of the water column). Conversely, seasonal effects of increasing temperatures and changes
in wind patterns may favorably influence the upper water column cyanobacterial community.
This vertical variability is common and attributed to four causes, each of which may occur at
different times, including: (a) sinking of dead/dying cyanobacterial cells; (b) density
stratification of the water column, especially nutrient concentrations and light, which affects all
aspects of cyanobacteria growth; (c) increased nutrient supply from organic-rich bottom
sediment (even when the water body is not density-stratified), encouraging cyanobacteria growth
at or near the bottom sediment; and (d) species-specific factors such as the tendency to form
surface scums in the case of M aeruginosa or the presence of resting spores in the sediment in
the case of N. spumigena (Drake et al. 2010).
In addition to occurrence in lakes and reservoirs, cyanobacteria and cyanotoxins have
been detected in flowing rivers and streams (Chaney 2016; Commonwealth of Kentucky: Energy
and Environment Cabinet 2015; Florida Department of Environmental Protection 2016; Loftin et
al. 2016a; Otten et al. 2015; Paerl & Otten 2013b; Parker 2016). In some cases, the source of the
cyanobacteria can be traced to an upstream water body such as a lake or reservoir. In 2016, a
bloom in Lake Okeechobee impacted the St. Lucie River and estuary and the Caloosahatchee
River and estuary in Florida (Florida Department of Environmental Protection 2016). Otten et al.
(2015) used microbial source tracking techniques to trace the source of a toxic Microcystis
bloom in the Klamath River in Oregon to a single upstream reservoir. Their results showed that
large quantities of cyanobacterial cells can withstand passage through hydroelectric installations
and transport over 300 kilometers. Cyanobacterial bloom development has been documented
near dams and man-made reservoirs (Chaney 2016; Giannuzzi et al. 2011; Otten et al. 2015;
Sieroslawska et al. 2010). Environmental characteristics including nutrients and flow rate can
affect phytoplankton dynamics (Paerl & Otten 2013b). Zhang et al. (2015) observed that low
flow conditions favored cyanobacteria and higher flow conditions favored green algae.
Cyanobacterial blooms can also occur in rivers and streams without a known lake or
reservoir source in the water column or as part of the benthic community (Commonwealth of
Kentucky: Energy and Environment Cabinet 2015; Loftin et al. 2016a). Loftin et al. (2016a)
suggest that low stream flow, shallow depth, and high water column light penetration in
Piedmont streams favored periphyton occurrence (mixture of algae, cyanobacteria, heterotrophic
bacteria, and detritus). A review by Quiblier et al. (2013) of benthic freshwater cyanobacterial
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ecology found that nutrients, flow regime, wave action, climate, and geology can influence
benthic cyanobacterial community composition and suggest that due to a high desiccation
tolerance, cyanobacteria occurrence in benthic mat communities is also a concern in ephemeral
streams.
In addition, there are microbial interactions that may occur within blooms, such as
competition and adaptation between toxic and nontoxic cyanobacterial strains, as well as impacts
from viruses and zooplankton grazers like Daphnia (large generalist grazers), copepods, and
cladocerans (Ger et al. 2014). Each of these microbial-related factors can cause fluctuations in
bloom development and composition.
In summary, there is a complex interplay of environmental factors that dictates the spatial
and temporal changes in the concentration of cyanobacterial cells and their toxins with respect to
the dominant species as illustrated in Figure 3-1 (Paerl & Otten 2013b). Factors such as the
amount and timing of nutrient supply (i.e., nutrient concentration and nutrient loading), the
relative proportions of nutrients (i.e., nitrogen to phosphorus ratio), dissolved organic matter
availability, temperature, and light attenuation, as well as other physico-chemical processes, can
play a role in shaping cyanobacterial bloom composition and cyanotoxin production (Paerl &
Huisman 2008; Paerl & Otten 2013b). Phytoplankton competition and food web interactions that
occur as blooms develop, persist, and decline can also impact cyanotoxin concentrations in
surface waters. In addition, impacts of climate change, including potential warming of surface
waters and changes in precipitation, could result in changes in ecosystem dynamics that lead to
more frequent formation of cyanobacteria blooms and their associated toxins (Paerl & Huisman
2008; Paerl & Otten 2013b; Paerl et al. 2011).
3.2 Cyanotoxins
Much of the information and the studies summarized in this section for microcystin and
cylindrospermopsin are described in detail in EPA's Health Effects Support Document for the
Cyanobacterial Toxin Microcystins and Health Effects Support Document for the Cyanobacterial
Toxin Cylindrospermopsin (HESDs), and EPA's Drinking Water Health Advisory for the
Cyanobacterial Microcystin Toxins and Drinking Water Health Advisory for the Cyanobacterial
Toxin Cylindrospermopsin (Drinking Water Health Advisories) (U.S. EPA 2015a; U.S. EPA
2015b; U.S. EPA 2015c; U.S. EPA 2015d).
3.2.1 Chemical and Physical Properties
Summary information for chemical and physical properties is provided in this section.
Additional information can be found in the EPA's HESDs for microcystins and
cylindrospermopsin (U.S. EPA 2015c; U.S. EPA 2015d).
Structurally, microcystins are monocyclic heptapeptides that contain seven amino acids
joined end-to-end and then head to tail to form cyclic compounds that are comparatively large;
molecular weights range from approximately 800 to 1,100 g/mole. The cyclic peptides include
more than 100 congeners of microcystins (Niedermeyer 2014). Figure 3-2 provides the structure
of microcystin where X and Y represent variable amino acids. Although substitutions mostly
occur in positions X and Y, other modifications have been reported for all of the amino acids
(Puddick et al. 2015).
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The microcystins are named based on their two variable amino acids (Carmichael et al.
1988). For example, microcystin-LR, the most common congener, contains leucine (L) and
arginine (R) (Carmichael 1992). The letters used to identify the variable amino acids are the
standard single letter abbreviations for the amino acids found in proteins. The variable amino
acids are usually the L-amino acids as found in proteins. For example, microcystin-LR is for the
microcystin with leucine in the X position of Figure 3-2 and arginine in the Y position in Figure
3-2. Table 3-1 lists the most common microcystins congeners.
Figure 3-2. Structure of Microcystin (Kondo et al. 1992)
H C02H
^CH2
h3cv/
h ch ¦ ¦r h- ,cHa h hY
H 3CHa H
O H CO?H
Table 3-1. Abbreviations for Microcystins (Yuan et al. 1999)
Microcystin Congeners
Amino Acid in X
Amino Acid in Y
Microcystin-LR
Leucine
Arginine
Microcystin-RR
Arginine
Arginine
Microcystin-YR
Tyrosine
Arginine
Microcystin-LA
Leucine
Alanine
Microcystin-LY
Leucine
Tyrosine
Microcystin-LF
Leucine
Phenylalanine
Microcystin-LW
Leucine
Tryptophan
The preponderance of toxicological data on the effects of microcystins is restricted to the
microcystin-LR congener. Toxicity data suggest that microcystin-LR is as potent as or more
potent than other studied microcystins and that the most toxic microcystins are those with the
more hydrophobic L-amino acids (-LA, -LR, -YR, and -YM); the least toxic are those with
hydrophilic amino acids, such as microcystin-RR. Data on the -RR, -YR, and -LA congeners,
however, are limited, and toxicity values cannot be derived for them. Values developed from
data specific to microcystin-LR are considered applicable to and appropriate for individual and
mixtures of microcystin congeners.
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Table 3-2 provides chemical and physical properties of microcystin-LR. Microcystins are
water-soluble. In aquatic environments, the cyclic peptides tend to remain contained within the
cyanobacterial cell and are released in substantial amounts only upon cyanobacterial cell lysis.
Table 3-2. Chemical and Physical Properites of Microcystin-LR
Property
Microcystin-LR
Chemical Abstracts Registry (CAS) Number
101043-37-2
Chemical Formula
C49H74N10O12
Molecular Weight
995.17 g/mole
Color/Physical State
Solid
Boiling Point
N/A
Melting Point
N/A
Density
1.29 g/cm3
Vapor Pressure at 25 °C
N/A
Henry's Law Constant
N/A
Log Ko„
2.16
Koc
N/A
Solubility in Water
Highly
Other Solvents
Ethanol and methanol
Sources: Chemical Book (2012); TOXLINE (2012); Ward and Codd (1999) for log Kow.
Cylindrospermopsin is a tricyclic alkaloid with the following molecular formula
C15H21N5O7S (Ohtani et al. 1992) and a molecular weight of 415.43 g/mole. It is zwitterionic
(i.e., a dipolar ion with localized positive and negative charges) (Ohtani et al. 1992). The
chemical structure of cylindrospermopsin is presented in Figure 3-3. Additional congeners
and analogs have been identified; see U.S. EPA (2015b; 2015c) for more information.
Figure 3-3. Structure of Cylindrospermopsin (de la Cruz et al. 2013)
H OH
(R)
(R)
NH
HN
NH
¦NH
©
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The physical and chemical properties of cylindrospermopsin are presented in Table 3-3.
Cylindrospermopsin is highly soluble in water (Chiswell et al. 1999; Moore et al. 1998).
Cylindrospermopsin is isolated for commercial use mostly from C. raciborskii. Many of the
physicochemical properties of cylindrospermopsin in the environment such as vapor pressure
and boiling and melting points are unknown.
Table 3-3. Chemical and Physical Properties of Cylindrospermopsin
Property
Cylindrospermopsin
Chemical Abstracts Service (CAS) Registry
Number
143545-90-8
Chemical Formula
C15H21N5O7S
Molecular Weight
415.43 g/mole
Color/Physical State
white powder
Boiling Point
N/A
Melting Point
N/A
Density
2.03 g/cm3
Vapor Pressure at 25°C
N/A
Henry's Law Constant
N/A
Kow
N/A
Koc
N/A
Solubility in Water
Highly
Other Solvents
Dimethyl sulfoxide (DMSO) and methanol
Sources: Chemical Book (2012); TOXLINE (2012).
3.2.2 Sources and Occurrence
Cyanobacterial density in a bloom and cyanotoxin concentration are not always closely
related. Cyanotoxin concentrations depend on the dominance and diversity of species and strains
within the bloom along with environmental and ecosystem influences on bloom dynamics
(Chorus et al. 2000; Hitzfeld et al. 2000; WHO 1999). Cyanotoxin production by cyanobacteria
is highly variable and strongly influenced by the environmental conditions. It can vary among
strains and clones of a single species (Carmichael 1994; Utkilen & Gj0lme 1992) and within and
between blooms (Codd & Bell 1985). Growth phase also can influence cyanotoxin production
(Jaiswal et al. 2008). Although studies of the impact of environmental factors on cyanobacteria
bloom are ongoing, a variety of factors can influence cyanotoxin production, including nutrient
(nitrogen, phosphorus) concentrations, light levels, temperature, pH, oxidative stressors, and
interactions with other biota (viruses, bacteria, and animal grazers), and the combined effects of
these factors (Paerl & Otten 2013a; Paerl & Otten 2013b). Factors discussed previously that
influence cyanobacterial growth can also influence cyanotoxin production, however, growth and
toxin production do not necessarily coincide. Recent research by Francy et al. (2016) on
modeling the relationship of environmental variables and cyanotoxin levels has shown that
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certain environmental factors may be useful to estimating microcystin concentrations above a
threshold level.
The proportion of intracellular versus extracellular cyanotoxin can also vary.
Extracellular microcystins (either dissolved in water or bound to other materials) typically make
up less than 30 percent of the total microcystin concentration in source water (Graham et al.
2010). Most of the microcystins are intracellular and released into the water when the
cyanobacterial cells rupture or die. Cylindrospermopsin may be retained within the
cyanobacterial cell or released. The ratio of intracellular to extracellular cyanotoxin can change
depending on the growth phase with as much as 50 percent of cylindrospermopsin produced by
C. raciborskii released extracellularly (Griffiths & Saker 2003).
3.2.2.1 Surface Water
Microcystins
Microcystins are the most common cyanotoxins found worldwide and have been reported
in surface waters in most of the states in the United States (Funari & Testai 2008). Dry-weight
concentrations of microcystins in surface freshwater cyanobacterial blooms or surface freshwater
samples reported worldwide between 1985 and 1996 ranged from 1 to 7,300 jag/g. Water
concentrations of extracellular plus intracellular microcystins ranged from 0.04 to 25,000 (J,g/L.
The concentration of extracellular microcystins ranged from 0.02 to a high of 1,800 [j,g/L
reported following treatment of a large cyanobacteria bloom with algaecide (WHO 1999), and
the U.S. Geological Survey (USGS) reported a concentration of 150,000 [j,g/L total microcystins
in a lake in Kansas (Graham et al. 2012).
Microcystins have been detected in most states, and over the years, many studies have
been done to determine their occurrence in surface water. The remainder of this section provides
examples of microcystin occurrence observations throughout the United States.
According to a survey conducted in Florida in 1999 between the months of June and
November, the most frequently observed cyanobacteria werq Microcystis (43.1 percent),
Cylindrospermopsis (39.5 percent), and Anabaena spp. (28.7 percent) (Burns 2008). Of 167
surface water samples taken from 75 waterbodies, microcystin was the most commonly found
cyanotoxin in water samples collected, occurring in 87 water samples.
In 2002, the Monitoring and Event Response to Harmful Algal Blooms in the Lower
Great Lakes project evaluated the occurrence and distribution of cyanotoxins in the lower Great
Lakes region (Boyer 2007). Analysis for total microcystins was performed using Protein
Phosphatase Inhibition Assay. Microcystins were detected in at least 65 percent of the samples,
mostly in Lake Erie, Lake Ontario, and Lake Champlain. The National Oceanic and Atmospheric
Administration Center of Excellence for Great Lakes and Human Health continues to monitor
the Great Lakes and regularly samples cyanobacterial blooms for microcystin in response to
bloom events.
A 2004 study of the Great Lakes found high levels of cyanobacteria during the month of
August (Makarewicz et al. 2006). Microcystin-LR was analyzed by protein phosphatase
inhibition assay (limit of detection of 0.003 (J,g/L) and was detected at levels of 0.084 |ig/L in the
nearshore and 0.076 [j.g/L in the bays and rivers. This study reported higher levels of
microcystin-LR (1.6 to 10.7 (J,g/L) in smaller lakes in the Lake Ontario watershed.
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In 2006, USGS conducted a study of 23 lakes in the Midwestern United States in which
cyanobacterial blooms were sampled to determine the co-occurrence of cyanotoxins in
cyanobacterial blooms (Graham et al. 2010). This study reported that microcystins were detected
in 91 percent of the lakes sampled with 17 percent of microcystin-positive samples exceeding
20 |ig/L. Mixtures of all the microcystin congeners measured (-LA, -LF, -LR, -LW, -LY, -RR,
and -YR) were common, and all the congeners were present in association with the blooms.
Microcystin-LR and -RR were the dominant congeners detected with mean concentrations of
104 and 910 [j.g/L respectively.
In 2007, the NLA conducted the first national probability-based survey of the condition
of the nations' lakes, ponds, and reservoirs (U.S. EPA 2009). This baseline study provided
estimates of the condition of natural and man-made freshwater lakes, ponds, and reservoirs
greater than 4 hectares (10 acres) and at least one meter deep. The NLA measured microcystins
using enzyme linked immunosorbent assays (ELISA) with a detection limit of 0.1 [j.g/L as well
as cyanobacterial cell counts and chlorophyll a concentrations, which were indicators of the
presence of cyanotoxins. Samples were collected in open water at mid-lake. Due to the design of
the survey, no samples were taken nearshore or in other areas where scums were present. These
surveys covered a total of 1,028 lakes, which represented nearly 50,000 lakes in the
conterminous United States. This assessment found that cyanobacteria were detected in almost
all lakes (U.S. EPA 2009). Cyanobacteria were the dominant member of the phytoplankton
community in 76 percent of lake samples. Subsequent analysis indicated that potential
microcystin-producing species occurred in 95 percent of samples (Loftin et al. 2016b).
Microcystins are the most commonly detected class of cyanotoxin and have been found
in lakes in the contiguous United States (U.S. EPA 2009) and streams in the Southeastern United
States (Loftin et al. 2016b). Microcystins were present in 30 percent of the lakes sampled
nationally by the NLA, with sample concentrations that ranged from the limit of detection
(0.1 (J,g/L) to 225 [j,g/L (U.S. EPA 2009). Microcystins were detected in in 32 percent of lake
water samples with a mean concentration of 3.0 |ig/L (based on detections only) and microcystin
concentrations above the WHO thresholds of concern of 10 and 20 [j,g/L were present in
1.1 percent of samples nationally (Loftin et al. 2016b). States with lakes reporting microcystins
levels above > 10 [j.g/L are shown in Table 3-4. NLA data show two states (North Dakota and
Nebraska) had 9 percent of samples above 10 (J,g/L. Other states including Iowa, Texas, South
Dakota, and Utah also had samples that exceeded 10 (J,g/L. Several NLA samples in North
Dakota, Nebraska, and Ohio exceeded 20 [j,g/L (192 and 225 [j,g/L respectively). EPA completed
a second survey of lakes in 2012, however, those data have not yet been released.
USGS did a study in the Upper Klamath Lake in Oregon in 2007 and detected total
microcystin concentrations between 1 [j,g/L and 17 [j,g/L (VanderKooi et al. 2010). USGS also
monitored Lake Houston in Texas from 2006 to 2008, and found microcystin in 16 percent of
samples and at concentrations less than or equal to 0.2 [j,g/L (Beussink & Graham 2011). In
2011, USGS conducted a study on the upstream reservoirs of the Kansas River to characterize
the transport of cyanobacteria and associated compounds (Graham et al. 2012). Concentrations
of total microcystin were low in the majority of the tributaries with the exception of Milford
Lake, which had higher total microcystin concentrations, some exceeding the Kansas
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Table 3-4. States Surveyed as Part of the 2007 National Lakes Assessment with Water Body
Microcystins Concentrations above 10 jig/L (U.S. EPA 2009)
State
Number
of Sites
Sampled
Percentage of Samples with
Detection of Microcystins >
lOjig/L
Maximum Detection of
Microcystins
North Dakota
38
9.1 percent
192 |ig/L
Nebraska
42
9.1 percent
225 |ig/L
South Dakota
40
4.9 percent
33 |ig/L
Ohio
21
4.5 percent
78 |ig/L*
Iowa
20
4.5 percent
38 |ig/L*
Utah
26
3.6 percent
15 |ig/L*
Texas
51
1.8 percent
28 ng/L*
* Single Sample
recreational guideline level of 20 (J,g/L. Upstream from Milford Lake, a cyanobacterial bloom
was observed with a total microcystin concentration of 150,000 (J,g/L. When sampled a week
later, total microcystin concentrations were less than 1 (J,g/L. The study authors indicated that this
may be due to dispersion of microcystins through the water column or to other areas, or by
degradation of microcystins via abiotic and biological processes. Samples taken during the same
time from outflow waters contained total microcystin concentrations of 6.2 (J,g/L.
In 2005, Washington State Department of Ecology developed the Ecology Freshwater
Algae Program to focus on the monitoring and management of cyanobacteria in Washington
lakes, ponds, and streams (WSDE 2012). The data collected have been summarized in a series of
reports for the Washington State Legislature (Hamel 2009; Hamel 2012). Microcystin levels
ranged from the detection limit (0.05 (J,g/L) to 4,620 [j,g/L in 2008, to 18,700 [j,g/L in 2009, to
853 (J,g/L in 2010, and to 26,400 (J,g/L in 2011.
A survey conducted during the spring and summer of 1999 and 2000 in more than
50 lakes in New Hampshire found measureable microcystin concentrations in all samples (Haney
& Ikawa 2000). Microcystins were analyzed by ELISA and were found in all of the lakes
sampled with a mean concentration of 0.1 (J,g/L. In 2005 and 2006, a study conducted in New
York, including Lake Ontario, found variability in microcystin-LR concentrations within the
Lake Ontario ecosystem (Makarewicz et al. 2009).
Since 2007, Ohio EPA (2012) has been monitoring inland lakes for cyanotoxins. Of the
19 lakes in Ohio sampled during the NLA, 36 percent had detectable levels of microcystins. In
2010, Ohio EPA sampled Grand Lake, St. Marys for anatoxin-a, cylindrospermopsin,
microcystins, and saxitoxin. Microcystin levels ranged from below the detection limit
(< 0.15 (J,g/L) to more than 2,000 (J,g/L. Follow-up samples taken in 2011 for microcystins
indicated concentrations exceeded 50 [j,g/L in August. During the same month, sampling in Lake
Erie found microcystins levels exceeding 100 (J,g/L.
In 2008, NOAA began monitoring for cyanobacterial blooms in Lake Erie using high
temporal resolution satellite imagery. Between 2008 and 2010, Microcystis cyanobacterial
blooms were associated with water temperatures above 18°C (Wynne et al. 2013). Using the
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Great Lakes Coastal Forecast System, forecasts of bloom transport are created to estimate the
trajectory of the bloom, and these are distributed as bulletins to local managers, health
departments, researchers, and other stakeholders. To evaluate bloom toxicity, the Great Lakes
Environmental Research Laboratory collected samples at 6 to 8 stations each week for 24 weeks,
measuring cyanotoxin concentrations as well as chlorophyll biomass and an additional
18 parameters (e.g., nutrients) to improve future forecasts of these blooms. Microcystins can be
separated into particulate (cell-bound) and dissolved (extracellular) phases, which can be
measured by testing concentrations in the filter and filtrate fractions of the sampled water
(Graham & Jones 2007; Zastepa et al. 2014). In 2014, particulate microcystin concentrations
ranged from below detection to 36.7 (J,g/L. Samples taken in 2015 and 2016 showed particulate
microcystin concentration ranges from below detection to 9.19 [j,g/L and from below detection to
21.26 |ig/L, respectively. Particulate microcystin concentrations peaked in August 2014 at all
sites, with the Maumee Bay site yielding the highest concentration of the entire three-year
sampling period. Dissolved microcystin concentrations were also collected at each site in 2014
from September until the end of the sampling period in November, as well as during the field
sampling seasons in 2015 and 2016. During the final months of sampling in 2014 (October to
November), dissolved microcystin concentrations were detected with peak concentrations of
0.8 [j,g/L (mean: 0.28 +/- 0.2 (J,g/L) whereas particulate microcystin concentrations were below
detection limits on many dates, indicating that a majority of the microcystin (mean: 72 percent
+/- 37 percent) were in the dissolved form, as the bloom declined in intensity. Measured
dissolved microcystin concentrations in the following two years ranged from levels below
detection to peaks of 0.69 \igFL in September 2015 and 1.76 [j,g/L in July 2016 (NOAA 2014).
Note that the health-protective value for microcystins recommended in this document should be
compared with the total microcystins detected and not delineated between intracellular or
extracellular microcystin. Cells containing microcystin can be swallowed while recreating and
contribute to the overall exposure to the toxin.
Two notable cyanobacterial blooms occurred in Florida and Utah in 2016, resulting in
microcystin detections. From July 14 to September 14, an extensive cyanobacterial bloom
covering 100 square miles occurred in Utah Lake, Jordan River, and nearby canals and included
the cyanobacterial genera Geitlerinema, Oscillatoria, and Pseudanabaena (Utah Department of
Environmental Quality 2016). Microcystin concentrations ranged from < 0.5-176 |ig/L. The
Utah Department of Environmental Quality reported over 500 human exposures with 30 percent
of these cases reporting symptoms such as gastrointestinal distress, headache, and eye and skin
irritation. In addition, 27 animal exposures were reported (Utah Department of Environmental
Quality 2016)^
In 2016, a 239-square mile cyanobacterial bloom in Lake Okeechobee, Florida, and
downstream waterways resulted in a state of emergency in four counties on the Gulf and Atlantic
coasts of Florida (Chaney 2016; Parker 2016). From May 4 to August 4, the Florida Department
of Environmental Protection took approximately 200 water samples from the St. Lucie River and
estuary, Caloosahatchee River and estuary, Lake Okeechobee, Indian River Lagoon, and other
nearshore marine locations (Florida Department of Environmental Protection 2016). Microcystin
concentrations ranged from below the detection limit to 414.3 (J,g/L. Among the species
identified were Microcystis aeruginosa, Scrippsiella trochoidea, Planktolyngbya limnetica,
Dolichospermum circinalis, and Plectonema wollei (Florida Department of Environmental
Protection 2016). Lake Okeechobee, located north of the Everglades, is the largest freshwater
lake in Florida. It is subject to agricultural runoff from adjacent cattle farms and sugar cane
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fields, which contributed to the formation of this massive cyanobacterial bloom (Parker 2016).
Water may be pumped out of the lake to the coast through the St. Lucie River and the
Caloosahatchee River to prevent the lake level from rising too high after periods of heavy rain,
(Parker 2016). As a result of the microcystin levels and visible cyanobacterial scum from water
discharged from the lake that flowed downstream to coastal areas, beaches along the Atlantic
were closed, and a state of emergency was declared in the counties of Martin, St. Lucie, Palm
Beach, and Lee (Chaney 2016; Florida Department of Environmental Protection 2016).
Cylindrospermopsin
As noted above, EPA's NLA conducted the first national probability-based survey of
lakes (U.S. EPA 2009) and published results in 2007. USGS subsequently analyzed the stored
samples collected and detected cylindrospermopsin in 4 percent of samples, with a mean
concentration 0.56 |ig/L and a range from the limit of detection, 0.01 |ig/L, to a maximum of
4.4 |ig/L (Loftin et al. 2016b). Potential cylindrospermopsin-producing species occurred in
67 percent of samples (Loftin et al. 2016b). In general, fewer surface water occurrence data are
available for cylindrospermopsin compared to microcystin. This is likely because during blooms,
testing for microcystin is much more common than testing for cylindrospermopsin.
USGS also detected cylindrospermopsin in 9 percent of blooms sampled during a 2006
USGS survey of 23 lakes in the Midwestern United States (Graham et al. 2010). The low
concentrations of cylindrospermopsin detected (0.12 to 0.14 (J,g/L) in the study occurred in
bloom communities dominated by Aphanizomenon or Anabaena and Microcystis.
Cylindrospermopsin has been detected in lakes throughout multiple states. In a 1999
study, cylindrospermopsin was detected in 40 percent of 167 water samples taken from 87 water
bodies in Florida during the months of June and November (Burns 2008). However, the actual
cylindrospermopsin concentrations were not reported. In 2005, the U.S. Army Corps of
Engineers detected cylindrospermopsin at a maximum concentration of 1.6 [j,g/L in lake water
samples from Oklahoma (Lynch & Clyde 2009). In Grand Lake in St. Marys, Ohio,
cylindrospermopsin concentrations as high as 9 [j,g/L were reported in 2010 (Ohio EPA 2012).
3.2.2.2 Ambient Air
According to Wood and Dietrich (2011), waterborne cyanotoxins can be aerosolized
through a bubble-bursting process, in which the cyanobacteria and cyanotoxins are ejected and
carried into the air by the resulting droplets from the bubble bursting. Microcystin that is free or
bound to particles can be deposited into the deepest bronchiolar or alveolar cavities; the
cyanobacterial cells can be likely deposited in the upper respiratory tract (Wood & Dietrich
2011).
Four studies provide air concentration data indicating that recreational surface waters
with cyanotoxin-producing cyanobacterial blooms can result in aerosolized cyanotoxins. Backer
et al. (2008) used personal air samplers in a 3-day study of recreational activities in a lake with a
cyanobacterial bloom, either carried by the study participant or placed on the participant's boat.
The microcystin concentrations in air ranged from below the limit of detection (0.0037 ng/m3) to
0.456 ng/m3. Backer et al. (2010) also detected microcystins in ambient air for one day, at one
lake, and only from the shoreline sampler. The average air concentration was 0.052 ng/m3. They
also collected 44 personal air samples, which ranged from the limit of detection (0.1 ng/m3) to
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0.4 ng/m3. The authors noted that the daily mean concentrations of microcystin in personal air
samples did not correlate with the concentrations of Microcystis cells, dissolved microcystin, or
total microcystin in the sampled lake water.
Wood and Dietrich (2011) studied Lake Rotorua (New Zealand) when it was
experiencing a dense bloom of microcystin-producing Microcystis species. They measured a
maximum microcystin concentration in the water of (2,140 |ig/L) and air concentrations from
0.0003 to 0.0018 ng/m3.
Cheng et al. (2007) used high volume and personal air samplers to measure microcystins
in the air in at a lake with a cyanobacterial bloom. They measured low concentrations of
microcystin in the water (approximately 1 (J,g/L) and air concentrations ranging from below the
detection limit (0.02 ng/m3) to 0.08 ng/m3.
3.2.2.3 Other Sources of Microcystins and Cylindrospermopsin
Extracts from Arthrospira (Spirulina spp.) and Aphanizomenon flos-aquae have been
used as dietary blue-green algae supplements (BGAS) (Funari & Testai 2008). These
supplements are reported to have beneficial health effects including supporting weight loss, and
increasing alertness, energy and mood elevation for people suffering from depression (Jensen et
al. 2001). A study suggested that BGAS can be contaminated with microcystins ranging from
1 [j,g/g up to 35 [j,g/g (Dietrich & Hoeger 2005). Heussner et al. (2012) analyzed 18 commercially
available BGAS for the presence of cyanotoxins. All products containing Aphanizomenon flos-
aquae tested positive for microcystins at levels < 1 (J-g microcystin-LR equivalents/g dry weight.
Cylindrospermopsin was not found in any of the supplements.
3.2.3 Environmental Fate
Different physical and chemical processes are involved in the persistence, breakdown,
and movement of microcystins and cylindrospermopsin in aquatic systems as described below.
3.2.3.1 Mobility
Cyanotoxins can move within water systems or they can be transported between systems.
Mechanisms concentrating cyanobacterial cells can also act to concentrate their cyanotoxins,
leading to negative human health impacts including impacts on surface waters and direct contact
and aerosol exposure (Blaha et al. 2009; Carmichael 2001; Cheung et al. 2013; Codd et al. 2005).
Microcystins may adsorb onto naturally suspended solids and dried crusts of
cyanobacteria. They can precipitate out of the water column and reside in sediments for months
(Falconer 1998; Han et al. 2012). Ground water is generally not expected to be at risk of
cyanotoxin contamination, however, ground water under the direct influence of surface water
can be vulnerable. A study conducted by the USGS and the University of Central Florida
determined that microcystin and cylindrospermopsin did not sorb in sandy aquifers and were
transported along with ground water (O'Reilly et al. 2011). The authors suggested that the
removal of microcystin was due to biodegradation.
In sediments, cylindrospermopsin exhibits some adsorption to organic carbon, with little
adsorption observed on sandy and silt sediments (Klitzke et al. 2011). The low adsorption of
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cylindrospermopsin reduces its residence time in sediments, thus reducing the opportunity for
microbial degradation.
3.2.3.2 Persistence
Microcystins
Microcystins are relatively stable and resistant to chemical hydrolysis or oxidation at or
near neutral pH. Elevated or low pH or temperatures above 30°C may cause slow hydrolysis.
Microcystins have been observed to persist for 21 days to 2-3 months in solution and up to
6 months in dry scum (Funari & Testai 2008; Rapala et al. 2006). Environmental conditions such
as temperature, pH, presence of light, salinity, and presence of certain aquatic bacteria, can
influence the rate of microcystin degradation (Schmidt et al. 2014). Microcystins can persist
even after a cyanobacterial bloom is no longer visible (Lahti et al. 1997b; Zastepa et al. 2014). In
a study by Zastepa (2014), dissolved microcystin-LA was present at a concentration of 20 [j,g/L
or greater for 9.5 weeks even though the Microcystis bloom was not visible after 5 weeks.
In the presence of full sunlight, microcystins undergo photochemical breakdown, but this
varies by microcystin congener (Chorus et al. 2000; WHO 1999). Zastepa et al. (2014) suggest
that microcystin-LA degrades at a slower rate than microcystin-LR, -RR, and -YR congeners.
The presence of water-soluble cyanobacterial cell pigments, in particular phycobiliproteins,
enhances this breakdown. Breakdown can occur in as few as 2 weeks to longer than 6 weeks,
depending on the concentration of pigment and the intensity of the light (Tsuji et al. 1994; Tsuji
et al. 1995). Several other factors, including photosensitizer concentration, pH, wavelength of
light (Schmidt et al. 2014), and whether microcystins are dissolved or present in particulate
matter (Lahti et al. 1997b) can affect the rate of transformation or photodegradation. According
to Tsuji et al. (1994) and Tsuji et al. (1995), microcystin-LR was photodegraded with a half-life
of about 5 days in the presence of 5 mg/L of extractable cyanobacterial pigment. Humic
substances can also act as photosensitizers and can increase the rate of microcystin breakdown in
sunlight. Others have found that high concentrations of humic acids can slow the rate of
microcystin transformation by sunlight (Schmidt et al. 2014). In deeper or turbid water, the
breakdown rate is slower. Welker and Steinberg (2000) estimated the maximum rate of
microcystin-LR degradation in the presence of humic substance photosensitizers. Extrapolating
results from their small experimental tubes to a water column of 1 meter, Schmidt et al. (2014)
estimated the half-life of microcystin-LR to be 90 to 120 days per meter of water depth in
surface waters. The researchers also demonstrated that the wavelength of light can also affect
degradation rates; complete microcystin degradation has been observed within 1 hour when
exposed to 254-nm light and within 5 days using 365-nm light. According to Lahti et al. (1997b),
microcystin-LR follows first-order decay kinetics, with a decimal reduction time of 30 days for
dissolved microcystins compared with 15 days for microcystins found in particulate matter.
Zastepa et al. (2014) also found that dissolved microcystin-LA persists longer than microcystin-
LA in particulates, with in situ half-lives of 15.8 days and 6.5 days, respectively.
Microcystins are susceptible to degradation by aquatic bacteria found naturally in surface
waters (Jones et al. 1994). Bacteria isolates of Arthrobacter, Brevibacterium, Rhodococcus,
Paucibacter, and various strains of the genus Sphingomonas (Pseudomonas) have been reported
to be capable of degrading microcystin-LR (de la Cruz et al. 2011; Han et al. 2012). These
degradative bacteria have also been found in sewage effluent (Lam et al. 1995), lake water
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(Cousins et al. 1996; Jones et al. 1994; Lahti et al. 1997b), and lake sediment (Lahti et al. 1997a;
Rapala et al. 1994; U.S. EPA 2015a). Lam et al. (1995) reported that the biotransformation of
microcystin-LR followed a first-order decay with a half-life of 0.2 to 3.6 days. In a study done by
Jones et al. (1994) with microcystin-LR in different natural surface waters, microcystin-LR
persisted for 3 days to 3 weeks; however, more than 95 percent loss occurred within 3 to 4 days.
A study by Christoffersen et al. (2002) measured half-lives in the laboratory and in the field of
approximately 1 day, driven largely by bacterial aerobic metabolism. These researchers found
that approximately 90 percent of the initial amount of microcystin disappeared from the water
phase within 5 days, irrespective of the starting concentration. Other researchers (Edwards et al.
2008) have reported half-lives of 4 to 14 days, with longer half-lives associated with a flowing
stream and shorter half-lives associated with lakes. Microcystin-LR degradation by Sphingopyxis
species has been observed with an optimal degradation rate at a pH between 6.5 and 8.5
(Schmidt et al. 2014). Several studies have demonstrated bacterial degradation of microcystin-
LR, but other congeners, such as microcystin-LF or -LA, are not significantly degraded (Zastepa
2014; Zastepa et al. 2014). Although microcystin-degrading bacteria might be present, initial
degradation could be slow as the bacteria need time to become active (Hyenstrand et al. 2003),
and microcystins can accumulate in the water column if these bacteria are not present at the time
of a toxic bloom (Schmidt et al. 2014).
Where rivers discharge to the ocean, freshwater cyanobacteria, cyanotoxins, or both can enter the
marine environment and this may impact aquatic life in marine environments (Andersen et al.
1993; Miller et al. 2010). Miller et al. (2010) confirmed the transfer of freshwater microcystins
to the marine environment. The researchers found that after introducing Microcystis
cyanobacteria to a saline environment, cyanobacteria can survive for 48 hours before lysing and
releasing microcystins. Microcystins concentrations decreased to 29 to 56 percent of the initial
concentration after 1 hour in the saline environment, but continued to be detected in the seawater
for at least 21 days, based on a detection limit of 0.02 [^g/L (Miller et al. 2010). Gibble and
Kudela (2014) made additional observations of microcystins at the interface of freshwater and
seawater, in the Monterey Bay area, California. In the first year of a 3-year study, microcystin
was detected in 15 of 21 fresh-water, estuarine, and marine locations. In the two subsequent
years, monitoring focused on four major watersheds that feed into Monterey Bay. The authors
observed high concentrations of microcystin in both autumn and spring seasons and concluded
that microcystins are likely present throughout the year and transfer to the coastal environment,
with the potential to be a persistent issue in the Monterey Bay area. The authors also correlated
anthropogenic nutrient loadings with microcystin.
Cylindrospermopsin
Cylindrospermopsin is relatively stable in the dark and at temperatures from 4°C to 50°C
for up to 5 weeks (ILS 2000). Cylindrospermopsin is also resistant to changes in pH and remains
stable for up to 8 weeks at pH 4, 7, and 10. In the absence of cyanobacterial cell pigments,
cylindrospermopsin tends to be relatively stable in sunlight, with a half-life of 11 to 15 days in
surface waters (Funari & Testai 2008).
Like microcystin, degradation of cylindrospermopsin increases in the presence of cell
pigments such as chlorophyll a and phycocyanin, a blue photosynthetic pigment found in
cyanobacteria. When exposed to both sunlight and cell pigments, cylindrospermopsin breaks
down rapidly, more than 90 percent within 2 to 3 days (Chiswell et al. 1999).
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Cylindrospermopsin has been shown to be decomposed by bacteria in laboratory studies;
the biodegradation is influenced by the cyanotoxin concentration, temperature and pH. Mohamed
and Alamri (2012) reported that cylindrospermopsin was degraded by Bacillus bacteria and
degradation occurred in 6 days at the highest toxin concentration (300 |ig/L) and in 7 or 8 days at
lower concentrations (10 and 100 (J,g/L, respectively). The biodegradation rate was also reported
to depend on temperature and pH, with the highest rates occurring in warm waters (25 and 30°C)
and neutral to slightly alkaline conditions (pH 7 and 8). Klitzke and Fastner (2012) confirmed the
observations of Mohamed and Alamri (2012), noting that a decrease in temperature from 20 to
10°C slowed down degradation by a factor of 10. They also found that degradation slowed
significantly under anaerobic conditions, with half-lives of 2.4 days under aerobic conditions and
23.6 days under anaerobic conditions.
3.2.4 Toxicokinetics
Limited data are available regarding the toxicokinetics of microcystins in environmental
exposure conditions (U.S. EPA 2015d). Available intestinal data indicate that the organic acid
transporter polypeptide (OATp) family transporters facilitate the absorption of microcystins from
the intestinal tract into liver, brain, and other tissues, as well as their export out of organs and
tissues (Cheng et al. 2005; Fischer et al. 2005; Svoboda et al. 2011). However, bile acids and
other physiologically-relevant substrates compete with microcystins for transporter uptake by the
liver (Thompson & Pace 1992); reduction or elimination of liver toxicity has been observed
during in vivo or in vitro exposures when microcystin uptake by OATp transporters is limited or
inhibited (Hermansky et al. 1990a; Hermansky et al. 1990b; Runnegar et al. 1995; Runnegar &
Falconer 1982; Runnegar et al. 1981). Both in vivo and in vitro studies have shown biliary
excretion of microcystins (Falconer et al. 1986; Pace et al. 1991; Robinson et al. 1991), possibly
via conjugation with cysteine and glutathione (Kondo et al. 1996). Additional details of
microcystin toxicokinetics can be found in U.S. EPA's Drinking Water Health Advisory and
HESD for microcystins (U.S. EPA 2015a; U.S. EPA 2015d).
Limited toxicokinetic data for cylindrospermopsin are available, and are derived from
mice intraperitoneal studies and in vivo studies that do not necessarily reflect environmental
exposure conditions (U.S. EPA 2015c). Cylindrospermopsin is absorbed from the GI tract
(Humpage & Falconer 2003; Shaw et al. 2001; Shaw et al. 2000) and is distributed primarily to
the liver but also to the kidneys and spleen (Norris et al. 2001). The metabolism and toxicity of
cylindrospermopsin is mediated by hepatic cytochrome P450 (CYP450) enzymes, and the
periacinar region of the liver appears to be the main target of toxicity where cylindrospermopsin
and its metabolites bind to proteins (Norris et al. 2001; Runnegar et al. 1995; Shaw et al. 2001;
Shaw et al. 2000). Elimination of cylindrospermopsin was continuous over a monitoring period
of 24 hours, with a large mean total recovery primarily from urine, and to a smaller extent, feces,
after 24 hours (Norris et al. 2001). Additional details of cylindrospermopsin toxicokinetics can
be found in EPA's Drinking Water Health Advisory and HESD for cylindrospermopsin (U.S.
EPA 2015b; U.S. EPA 2015c).
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4.0 PROBLEM FORMULATION
4.1 Conceptual Model
This conceptual model provides useful information to characterize and communicate the
potential health risks related to exposure to microcystins and cylindrospermopsin in recreational
waters. The sources of cyanotoxins in these waters, the recreational route of exposure for
biological receptors of concern, and the potential assessment endpoints (e.g., effects such as
kidney and liver toxicity) are depicted in the conceptual diagram below (Figure 4-1).
Figure 4-1. Conceptual Model of Exposure Pathways to the Cyanotoxins, Microcystins and
Cylindrospermopsin, and Cyanobacteria in Surface Waters while Recreating
STRESSORS
SOURCES
EXPOSURE ROUTES
RECEPTORS
ENDPOINTS
Inhalation
Oral
Dermal
Liver
damage
Children
Microcystins
Cylindrospermopsin
Cyanobacterial cells
Cancer
Kidney
damage
Reproductive
effects
Developmental
effects
Incidental
inhalation while
recreating
Dermal contact
while recreating
Lakes, ponds, and rivers
(freshwater, inland)
Incidental
ingestion while
recreating
Estuaries, bays, lagoons and
oceans (marine, coastal)
Inflammatory response
effects, e.g.,
gastrointestinal distress,
skin irritation
Data insufficient or
incomplete
Data sufficient for
quantitative use
Legend
Conceptual Model Diagram for Exposure via Recreational Exposures
The conceptual model is intended to explore potential links of exposure to a contaminant
or stressor with the adverse effects and toxicological endpoints important for management goals,
including the development of recreational ambient water quality criteria. Boxes that are shaded
indicate pathways that EPA considered quantitatively in estimating the advisory level, whereas
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the white boxes did not have sufficient data for EPA evaluate quantitatively. The solid lines are
for the cyanotoxins and the dotted lines are for the cyanobacterial cells.
Factors Considered in the Conceptual Model for Microcystins and Cylindrospermopsin
Stressors. The stressors are microcystins and cylindrospermopsin concentrations in water. These
toxins can be produced by cyanobacteria occurring in freshwater. Once produced, the toxins
have the potential to affect downstream waters, including coastal areas. The values
recommended in this document could be applied to coastal waters affected by toxins produced by
upstream freshwater cyanobacteria. Cyanobacterial cells as direct stressors in recreational
surface waters are discussed in Appendix D.
Sources. Cyanobacteria occur naturally in surface waters, such as lakes, ponds, rivers, estuaries,
bays, lagoons, and oceans in or surrounding the United States. Some genera of the cyanobacteria,
including Microcystis, Cylindrospermopsis, Anabaena, Planktothrix, and Nostoc, can produce
the cyanotoxins microcystins and cylindrospermopsin. Once these toxins are produced, they can
be stable in the environment for weeks (Funari & Testai 2008; Zastepa et al. 2014).
Routes of exposure. Exposure to cyanotoxins from recreational water sources can occur via oral
exposure (incidental ingestion while recreating); dermal exposure (contact of exposed parts of
the body with water containing cyanotoxins during recreational activities such as swimming,
wading, surfing); and inhalation exposure to contaminated aerosols (while recreating). The route
of exposure considered quantitatively is oral exposure to microcystin and cylindrospermopsin via
incidental ingestion while swimming. Dermal exposure happens during swimming; however,
significant dermal absorption of microcystins and cylindrospermopsin is not expected due to the
large size and charged nature of these molecules (Butler et al. 2012; U.S. EPA 2004; U.S. EPA
2007). EPA estimated that ingestion from inhalation is likely negligible compared to incidental
ingestion while recreating (see section 7.5.1.1). Routes of exposure other than ingestion of
drinking water are taken into account by the application of a relative source contribution value
(U.S. EPA 2000a). Routes of exposure other than incidental oral ingestion while swimming are
discussed further in the Effects Characterization, section 7.5.1.
Receptors. Anyone who recreates in a water body where cyanotoxins are present could be
exposed to cyanotoxins through ingestion, dermal contact, and inhalation of aerosols while
recreating in contaminated surface waters. Childhood is considered a vulnerable lifestage due to
children's potential increased exposure while recreating. Recreating children can be at greater
risk from exposure to microcystins or cylindrospermopsin because they have smaller body mass
compared to adults, they spend more time in contact with the water compared to adults, and they
incidentally ingest more water than adults while recreating. Thus, EPA is specifically
considering the recreational exposures children experience in this assessment. EPA evaluates and
discusses differences between lifestages in section 7.4 of the Effects Characterization. While
there are many examples in the literature and reports of animal poisonings and death from
exposure to cyanotoxins, values protective of animals such as dogs and livestock are not
generated in this document. However, section 7.6 discusses some animal specific issues,
including a summary of guidelines several states have developed for animals.
Endpoints. Available microcystin toxicity data indicate that the primary target organ for
microcystins is the liver as described in EPA's Health Effects Support Document for the
Cyanobacterial Toxin Microcystins (U.S. EPA 2015d). Available cylindrospermopsin toxicity
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data are described in EPA's Health Effects Support Document for the Cyanobacterial Toxin
Cylindrospermopsin (U.S. EPA 2015c). For cylindrospermopsin, EPA selected kidney effects as
the endpoint on which to base the measure of effect. Clinical, epidemiological, and outbreak
study results (see Appendix D) suggest a link between an increase in adverse inflammatory
symptoms among recreators and elevated cyanobacterial cell densities. However, there is
considerable uncertainty and variability associated with the epidemiological results, which did
not identify consistent effects at similar cyanobacterial densities. Specifically, significant
associations occur across a wide range of cell densities; associations vary with different specific
health endpoints or combined symptom categories; and differences in cyanobacterial community
composition are largely uncharacterized. These endpoints are not considered quantitatively in
this assessment, but potential health effects are described in the Effects Characterization section
7.1 along with a discussion of the uncertainties related to the data for cyanobacterial cells.
4.2 Analysis Plan
EPA's 2000 Methodology for Deriving Ambient Water Quality Criteria for the
Protection of Human Health (2000 Human Health Methodology) outlines EPA's process for
deriving Ambient Water Quality Criteria (AWQC) and guides the development of these
recreational criteria and swimming advisories (U.S. EPA 2000a).
The 2000 Human Health Methodology includes identifying the population subgroup that
should be protected, evaluation of cancer and non-cancer endpoints, measures of effect,
measures of exposure, relative source contribution (RSC), and evaluation of bioaccumulation. In
this analysis plan, EPA: (1) describes the RfD previously derived for microcystin and
cylindrospermopsin (measure of effect); (2) describes the calculation for the recreational criteria;
(3) discusses incidental ingestion exposure in terms of volume ingested, duration of exposure,
and body weight (measure of exposure) described in EPA's Exposure Factors Handbook and;
(4) discusses the RSC. These criteria focus on human exposure as a result of primary contact
recreation activities such as swimming where immersion and incidental ingestion of ambient
water are likely.
EPA's Health Effects Support Document for the Cyanobacterial Toxin Microcystins and
Health Effects Support Document for the Cyanobacterial Toxin Cylindrospermopsin (U.S. EPA
2015c; U.S. EPA 2015d) provide the health effects basis for the development of the Drinking
Water Health Advisories for microcystins and cylindrospermopsin (U.S. EPA 2015a; U.S. EPA
2015b), including the science-based decisions providing the basis for estimating the point of
departure. To develop the HESDs for microcystins and cylindrospermopsin, EPA assembled
available information on toxicokinetics, acute, short-term, subchronic and chronic toxicity
along with developmental and reproductive toxicity, neurotoxicity, immunotoxicity,
genotoxicity and cancer in humans and animals. For detailed descriptions of the literature
search strategies, see the HESDs for microcystins and cylindrospermopsin (U.S. EPA 2015c;
U.S. EPA 2015d). This document was subject to rigorous internal and external peer review
before it was finalized in 2015. The information evaluated for these documents also supports
the development of the recreational criteria and swimming advisories for microcystins and
cylindrospermopsin, which evaluate exposure via recreational water ingestion. EPA conducted
supplemental literature searches in September 2015 to capture new references, including effects
related to recreational exposure to cells. For detailed information search terms, see Appendix C.
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4.2.1 Approach for Recreational AWQC Derivation
The Recreational AWQC for microcystins and cylindrospermopsin are calculated as
described in the 2000 Human Health Methodology and presented in the equation below:
RfD x RSC x BW
Recreational AWQC (|_ig/L) =
IR
Where:
RfD = Reference dose (|j,g/kilograms [kg] body weight [bw]/day [d])
RSC = Relative source contribution (RSC is discussed in section 4.2.5).
BW = Mean body weight (kg)
IR = Ingestion rate (L/d) at approximately the 90th percentile (discussed in
section 4.2.3)
4.2.1.1 Magnitude, Duration and Frequency
EPA recommends that recreational criteria consist of a magnitude, duration, and
frequency. Magnitude is the numeric expression of the maximum amount of the contaminant that
may be present in a waterbody that supports the designated use. Duration is the period of time
over which the magnitude is calculated. Frequency of excursion describes the number of times
the pollutant may be present above the magnitude over the specified time period (duration). A
criterion is derived such that the combination of magnitude, duration, and frequency protect the
designated use (e.g., primary contact recreation). For microcystins and cylindrospermopsin, the
magnitude of the criteria is based on the data used to derive the toxicity (in this case the RfDs for
microcystins and cylindrospermopsin) values developed to support the Drinking Water Health
Advisories (U.S. EPA 2015a,b). The duration and frequency components of the criteria are
consistent with the approach discussed in previous recreational criteria, including the application
of the recommended magnitudes using different durations for beach management and waterbody
assessment.
4.2.2 Measures of Effect
A reference dose or RfD is an estimate (with uncertainties spanning perhaps an order of
magnitude) of the daily exposure to the human population (including sensitive subgroups) that is
likely to be without an appreciable risk of deleterious effects during a lifetime. EPA's HESDs for
microcystins and cylindrospermopsin (U.S. EPA 2015c,d), provide the health effects basis for
development of the reference dose or RfD, including the science-based decisions (i.e., selection
of the critical study and endpoints) providing the basis for estimating the point of departure and
application of uncertainty factors. EPA uses the RfD values for oral exposure previously peer
reviewed and documented in the HESDs for microcystins and cylindrospermopsin (U.S. EPA
2015c; U.S. EPA 2015d) in derivation of the recreational criteria and swimming advisories.
Dermal exposure happens during swimming; however, significant dermal absorption of
microcystins and cylindrospermopsin is not expected due to the large size and charged nature of
these molecules (Butler et al. 2012; U.S. EPA 2004; U.S. EPA 2007). Because available data are
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not sufficient, EPA is not quantifying effects resulting from dermal exposure to cyanotoxins. See
section 7.5.1.2 for a characterization of dermal exposure to these cyanotoxins.
Inhalation exposure occurs during swimming; however, data are not sufficient to quantify
health effects resulting from inhalation exposure to cyanotoxins at this time. See section 7.5.1.1
for a characterization of potential effects from inhalation exposure.
Dermal exposure to cyanobacterial cells can also result in adverse health effects, such
skin rashes, eye irritation, and ear irritation. Because adequate data are not available, EPA is not
quantifying effects resulting from exposure to cells at this time. Available epidemiological study
results do not provide consistent associations between cell densities and the inflammatory health
endpoints. Some of the studies have been limited in size, which could affect the ability to detect
an association if one exists. Differences in the cyanobacterial communities present at the study
sites may have affected the detection of associations. Characterization of confounders, such as
the presence of pathogens, was not consistent among the studies. See section 7.1.1 for a
characterization of potential effects from recreational exposure to cyanobacterial cells.
4.2.3 Measures of Exposure
The exposure parameters selected for use in calculating recreational criteria and
swimming advisories for microcystins and cylindrospermopsin include an ingestion rate (volume
of surface water incidentally ingested per day) and body weight (kg). Both body weight and
incidental ingestion while recreating are parameters that vary with age. The key study and other
data supporting these exposure factors are described in the sections that follow.
All recreational exposure studies that included both children and adults found that age
could influence incidental ingestion exposure while recreating. More specifically, children tend
to ingest more water and spend more time in the water compared to adults (Dufour et al. 2006;
Schets et al. 2011; U.S. EPA 1997). EPA's Exposure Factors Handbook (U.S. EPA 2011)
provides recommended values for body weights and incidental ingestion volumes and rates for
children and adults on an event basis. The Handbook recommends using the 97th percentile
ingestion rate for children and the maximum reported value for adults because the dataset is
limited (U.S. EPA 2011).
EPA's Exposure Factors Handbook (2011) edition and (1997) edition provided values
for time spent swimming per month and time spent in a pool/spa per day, respectively. EPA
(2011) compiled mean and 95th percentile swimming durations for different children's age
groups in minutes/month (e.g., mean swimming duration value for children 1 to < 2 years was
105 minutes/month, for children 3 to < 6 years was 137 minutes/month, and for children 6 to
11 years was 151 minutes/month). EPA needed a duration parameter expressed as time exposed
per day to calculate a daily ingestion rate. Converting the monthly values to daily durations (e.g.,
dividing the monthly value by 30 days per month) resulted in very short daily exposures that do
not seem reasonable given the other duration estimates available. Therefore, EPA used the
duration of recreational event per day reported in U.S. EPA (1997) of 2.7 hours per day for
children 5 to 11 years old.
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4.2.3.1 Incidental Ingestion
Primary Contact Exposure Scenario
EPA selected incidental ingestion during primary contact activities such as swimming for
the criteria derivation because data suggest that incidental ingestion can be considered the
highest potential exposure pathway for cyanotoxins while recreating. In a combined analysis of
2,705 individuals recreating in the Chicago Area Waterway System and 662 individuals
recreating at a public outdoor swimming pool, Dorevitch et al. (2011) studied the volume of
water ingested during a range of recreational activities. Study subjects took part in one of the
following activities: canoeing, fishing, kayaking, motor boating, rowing, wading/splashing, head
immersion (i.e., immersed one's head three times over a 10-minute interval), or swimming. At
the end of their exposure, participants self-reported whether they ingested water, and how much,
during their recreational experience. The results indicate that the odds of ingesting a teaspoon or
more of water are significantly higher among swimmers than among those who just immersed
their head in a swimming pool or those who participated in the other, more limited contact
activities on surface waters. More specifically, rowing, motor boating, fishing, wading/splashing,
and non-capsizing kayaking and canoeing were found to be low-ingestion activities, resulting in
95th percentile ingestion volumes between 0.01 and 0.012 L/hr. The study authors considered
those who capsized during canoeing or kayaking a "middle ingestion category," with a 95th
percentile ingestion volume of about 0.017 to 0.02 L/hr. Swimmers were the highest ingestion
category, with a 95th percentile ingestion volume of approximately 0.035 L/hr. Evaluations of
inhalation (see section 7.5.1.1) and dermal (see section 7.5.1.2) exposures suggest that those two
routes are minor compared to the oral exposure route. Thus, EPA determined that using a
swimmer scenario for exposure as the basis for the criteria is protective of these other aquatic
activities.
Incidental Ingestion per Day
To calculate the recreational incidental ingestion rate in units of volume per day, EPA
combined a distribution from EPA's (2011) Exposure Factors Handbook on incidental ingestion
volumes (volume per event normalized to volume per hour) and a distribution of exposure
durations (hours per day) from EPA's (1997) Exposure Factors Handbook. The recommended
97th percentile incidental ingestion volume for children combined with the mean exposure
duration represented the 90th percentile of this combined distribution to represent incidental
ingestion per day. These data are discussed in the following sections.
Ingestion Volume Studies
EPA's Exposure Factors Handbook (2011) cites Dufour et al. (2006) as the basis for its
default recreational ingestion values. Dufour et al. (2006) measured the incidental ingestion of
water while participants were swimming in a pool and found that children under the age of
18 years ingested higher volumes of water while swimming than adults and that males ingested
more than females. This small-scale pilot study (n = 53) used cyanuric acid as an indicator of
amount of pool water ingested while swimming in an outdoor pool. Participants were instructed
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to stay in the pool and actively swim for at least 45 minutes. Pool-water samples were collected
before the start of swimming activities, and participants' urine was collected for 24 hours after
the swimming event ended; pool-water and urine samples were analyzed for cyanuric acid. The
combined study population had a mean incidental ingestion volume of 0.019 L per swimming
event. Because sample size for the Dufour et al. (2006) study was small (i.e., 41 children and 12
adults), the authors reported results for children under the age of 18 years and adults. In addition,
children younger than 6 years were not included in the study design. One study by Schets et al.
(2011) reported surveyed parents' estimates of incidental ingestion for children younger than
6 years old. The Schets et al. (2011) reported ingestion values for children ages 0 to < 15 years
were similar to the Dufour et al. (2006) findings; see section 7.3 for more detail.
The values presented in EPA's Exposure Factors Handbook (2011) adjusted the Dufour
et al. (2006) data from a per event (e.g., 45 minutes) basis to an hourly ingestion rate. The
distribution of Dufour et al. (2006) measured incidental ingestion rates are graphically presented
in Figure 4-2. Based on these data, the Exposure Factors Handbook recommended assessments
use a 97th percentile (0.12 L/hr) for children and a maximum value (0.071 L/hr) for adults as
"upper percentile" values due to the limited sample size of the Dufour et al. (2006) study.
Several other studies (Dufour et al. 2006; Schets et al. 2011; Suppes et al. 2014; U.S. EPA
2000a) characterizing incidental ingestion while swimming are available and described in the
effects characterization section (section 7.3). These other studies reported similar results to
Dufour et al. (2006).
Duration of Recreational Exposure
Duration of recreational exposure quantifies the length of time people might be exposed
to cyanotoxins during their primary contact recreational use of surface waters contaminated with
cyanotoxins. Duration of recreational exposure is needed to convert recreational ingestion rates
in units of volume per hour to an amount incidentally ingested per day, which is the exposure
parameter needed for the recreational AWQC derivation.
EPA's Exposure Factors Handbook (1997) lists, for different age groups, time spent per
24 hours in an outdoor spa or pool, which is interpreted for purposes of this calculation as time
spent in direct contact with water, for example, swimming. The data are based on analysis of the
National Human Activity Pattern Survey by Tsang and Klepeis (1996). Figure 4-3 compares the
recreational duration data for different age groups and shows that recreators ages 5 to 11 years
tend to spend more time in the water than other child age groups and adults, although the 90th
percentile values are similar. A duration was not provided for children younger than 1 year. EPA
evaluated both the mean duration for the various age groups and available exposure parameters
for children younger than 6 years old; see section 7.4.
Other data show a similar trend of longer recreational durations for children. Schets et al.
(2011) investigated swimming durations in freshwater, marine water, and pools. They surveyed
8,000 adults, 1,924 of whom also provided estimates for their eldest child (<15 years of age)
and found that children spend, on average, 25 minutes longer swimming in freshwaters
compared to adults. The mean duration of swimming events for children ages 0-14 years in
freshwater and marine water were 79 minutes (1.3 hours, 95 percent CI: 12-270 minutes) and 65
minutes (1.1 hours, 95 percent CI: 8-240 minutes), respectively. Adult averages were all less
than 1 hour.
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Figure 4-2. Incidental Ingestion Rates Measured for Adults and Children (Dufour et al.
2006)
a>
¦c
-a
ai
0.220
0.200
0.180
0.160
0.140
0.120
0.100
0.080
0.060
0.040
0.020
0.000
III
Adult Study Subjects
Child Study Subjects (6 to < 18 years)
Figure 4-3. Direct Contact Recreational Exposure Duration by Age Group, Based on Table
15-119 in U.S. EPA (1997)
6.0
1
1
i
—
J
1-4 years 5-11 years 12-17 years 18-64 years General Population
~ Mean GD Median ~ 90th percentile
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Additional duration estimates of children's pool swimming have been identified by
EPA's Office of Pesticide Programs for use in its Swimmers Exposure Assessment Model
(SWIMODEL) for estimating chemical exposures during pool swimming, including direct
contact for competitive swimmers (U.S. EPA 2003). EPA's SWIMODEL considers short-term
exposure (using a high-end estimate of exposure time per event in order to represent a maximum,
one-time exposure) and intermediate/long-term exposure (using a shorter event duration to
represent an average of maximum and minimum exposures overtime). Among competitive
children swimmers, the longest short-term exposure duration used by the SWIMODEL is
2 hours/day for children ages 11-15 years (U.S. EPA 2003). Competitive swimming practice
durations, however, are less relevant for recreational scenarios in lakes and rivers than for
exposure in a pool. EPA's Exposure Factors Handbook (1997) lists the mean exposure duration
for children 5 to 11 years as 2.7 hours/day, which is longer than the maximum value for children
11 to 15 years used in the SWIMODEL.
Determination of Incidental Ingestion per Day
The incidental ingestion volume per day EPA used to calculate the recreational criteria or
swimming advisory is the product of the 97th percentile children's incidental ingestion rate
(0.12 L/hr) and mean exposure duration (2.74 hr/day) for children ages 5 to 11 years. EPA
evaluated the effect these multiple parameters had on the level of protection by analyzing the
combined distributions of ingestion volume per hour and duration of recreational exposure. EPA
compiled the published statistical parameters (i.e., mean, standard deviation, and minimum and
maximum data values) and evaluated the resulting distributions for both parameters compared to
a normal, log-normal, and gamma function. For both parameters, the log-normal or gamma
functions better described the distributions. Log-normal and gamma functions are strictly
positive distributions and reflect the apparent skewness in the data. Describing a distribution with
a normal function can result in negative values that are not representative. In the analysis, both
distributions were limited to their respective minimum and maximum data values. Table 4-1
shows the statistics of the combined distributions: (a) a lognormal distribution for both
parameters, (b) a lognormal distribution for the ingestion rate and a gamma distribution for the
exposure duration, and (c) a gamma distribution for both parameters. The combinations of the
two distributions assuming log-normal, gamma, or both, are shown in Figure 4-4 as hybrid
distributions. For all three combined distributions using combinations of log-normal and/or
gamma functions, the incidental ingestion rate per day (0.33 L/d) represents approximately the
90th percentile of the hybrid distributions (range 92nd to 94th percentile). Additional details
including the methodology for this analysis are provided in Appendix E.
Table 4-1. Summary Statistics of Combined Ingestion Volume and Exposure Duration
Distributions
Combined
Distribution
Ingestion
Volume (L/hr)
Distribution
Exposure
Duration
(hr/d)
Distribution
Summary Statistics for Ingestion Rate (L/d)
Minimum
Median
Mean
Maximum
Percentile
Associated with
0.33 L/d
a
Log-normal
Log-normal
0.0023
0.0873
0.12
1.47
94
b
Log-normal
Gamma
0.0018
0.0888
0.13
1.46
93
c
Gamma
Gamma
0.0000
0.0871
0.13
1.40
92
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Figure 4-4. Hybrid Distributions for Incidental Ingestion per Day (L/d)
a) Ingestion volume, tog-normal
Exposure duration: log-normal
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Q
el
O
fM
O
b) Ingestion volume: log-normal
Exposure duration: gamma
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in
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o
CM
o
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o
c) Ingestion volume: gamma
Exposure duration: gamma
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o
S H
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o 1
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o
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o
00 01 0.2 0 3 0.4 0.5 06 0.7 08 0.9 1.0
Ingestion rate (Uday)
4.2.3.2 Body Weight
Table 8-1 in EPA's Exposure Factors Handbook (U.S. EPA 2011) reported
recommended and other body weight statistics based on the National Health and Nutrition
Examination Survey. A range of age groups is included. Mean body weight for children aged
6 to < 11 years was 31.8 kg. EPA selected this body weight because it reflected the age group
with higher ingestion volumes (U.S. EPA 2011; Evans et al. 2006) and exposure duration
(U.S. EPA 1997). A discussion of younger children's exposure factors can be found in section
7.4.2.
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4.2.4 Relative Source Contribution
EPA's 2000 Human Health Methodology (2000a) outlines EPA's process for deriving
AWQC and guides the development of these recreational criteria. The 2000 Human Health
Methodology recommends the application of a RSC in the AWQC derivation to ensure that an
individual's total exposure from all routes of exposure to a contaminant does not exceed the RfD.
EPA considered the 2000 Human Health Methodology's Exposure Decision Tree Approach to
determine the RSC used in deriving the recreational values for microcystins and
cylindrospermopsin (Figure 4-1 in the 2000 Human Health Methodology document).
The RSC component of the AWQC calculation allows a percentage of the RfDs exposure
to be attributed to the consumption of ambient water and fish and shellfish from inland and
nearshore waters when there are other potential exposure sources. The RSC describes the portion
of the RfD available for AWQC-related sources (USEPA 2000a); the remainder of the RfD is
allocated to other sources of the pollutant. The rationale for this approach is that for pollutants
exhibiting threshold effects, the objective of the AWQC is to ensure that an individual's total
exposure from all sources does not exceed that threshold level. Exposures outside the RSC
include, but are not limited to, exposure to a particular pollutant from fish and shellfish
consumption, non-fish food consumption (e.g., fruits, vegetables, grains, meats, poultry, dietary
supplements), dermal exposure, and respiratory exposure.
Cyanotoxins are produced by cyanobacteria. As discussed previously, certain
environmental factors can lead to rapid growth of cyanobacteria in ambient water. Because
environmental factors are not always favorable for cyanobacterial growth, blooms and the
production of cyanotoxins are episodic in nature; therefore, determination of background levels
is not relevant for cyanotoxins in determining the RSC.
EPA determined that an RSC of 80 percent, as recommended in EPA's 2000 Human
Health Methodology, is appropriate for microcystins and cylindrospermopsin. The use of this
RSC means that 80 percent of recreators' exposure to cyanotoxins is from incidental ingestion of
ambient water during recreational activities. The application of an RSC 80 percent takes into
account the uncertainty associated with effects from dermal and inhalation exposures, exposure
to contaminated fish and shellfish or drinking water (i.e., 20 percent is set aside for these other
exposure routes). An RSC of 80 percent represents the ceiling for setting an RSC, and provides a
margin of safety for individuals, given currently available data on exposure to different sources
and via other routes.
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5.0 EFFECTS ASSESSMENT
The health effects studies summarized below for microcystin and cylindrospermopsin are
described in detail in EPA's HESDs and Drinking Water Health Advisories for these two
cyanotoxins (U.S. EPA 2015a,b,c,d).
5.1 Hazard Identification
5.1.1 Noncancer Health Effects
5.1.1.1 Animal Toxicity Studies
Microcystins
Studies in laboratory animals demonstrate liver, kidney, and reproductive effects
following short-term and subchronic oral exposures to microcystin-LR. Studies evaluating the
chronic toxicity of microcystins have not shown clinical signs of toxicity and are limited by
study design and by the lack of quantitative data. Observed effects in animals exposed orally or
via intraperitoneal (i.p.) to microcystin-LR include liver, reproductive, developmental, kidney,
and GI effects.
The preponderance of animal toxicity data on the noncancer effects of microcystins is
restricted to the microcystin-LR congener. Studies evaluating the chronic toxicity of
microcystins have not shown clinical signs of toxicity and are limited by study design and by the
lack of quantitative data. Available data on the RR, YR, and LA congeners did not provide dose-
response information sufficient for quantification. EPA is using data on effects of microcystin-
LR to represent other microcystin congeners (U.S. EPA 2015d).
For details see the Health Effects Support Document for the Cyanobacterial Toxin
Microcystins (U.S. EPA 2015d).
Cylindrospermopsin
Based on oral and i.p. studies in mice treated with purified cylindrospermopsin or
extracts of C. raciborskii cells, the liver and kidneys appear to be the primary target organs for
cylindrospermopsin toxicity.
No oral reproductive or developmental studies are available for cylindrospermopsin.
Developmental toxicity studies following i.p. administration of cylindrospermopsin provide
some evidence for maternal toxicity and decreased postnatal pup survival and body weight
(Chernoff et al. 2011; Rogers et al. 2007).
For details, see EPA's Health Effects Support Document for the Cyanobacterial Toxin
Cylindrospermopsin (U.S. EPA 2015c).
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5.1.1.2 Human Studies
Microcystins
Limited human studies examining microcystin effects on humans are available; however,
no dose-response data are available from ambient exposures to microcystins. The scant human
data on the oral toxicity of microcystin-LR are limited by the potential co-exposure to other
pathogens, cyanotoxins, and microorganisms; by the lack of quantitative information; and by the
failure to control for confounding factors. Available human studies evidence is supportive of the
liver as a target organ for toxicity (Carmichael 2001; Falconer et al. 1983; Giannuzzi et al. 2011;
Hilborn et al. 2013; Jochimsen et al. 1998; Li et al. 201 lb).
More detailed information on the human health effects of microcystins based on
epidemiological studies related to drinking water outbreaks, clinical studies, and cases studies are
discussed in the Health Effects Support Document for the Cyanobacterial Toxin Microcystins
(U.S. EPA 2015d). Of the epidemiological studies EPA identified, three studies evaluated human
health effects associated with recreational exposures to cyanobacteria and microcystins. These
studies are also summarized in the microcystins HESD and are summarized below.
• Backer et al. (2008) conducted an epidemiological study in a small lake in the United
States and compared microcystin concentrations in blood and reported symptoms in
people recreating in a lake with a M. aeruginosa bloom to those of people recreating in a
nearby bloom-free lake. Low levels of total microcystins (detection limit = 0.08 ng/m3)
were detected in air samples collected above a lake bloom. Phytoplankton counts ranged
from 175,000 to 688,000 cells per mL with > 95 percent of those cells being
cyanobacteria. Cell densities of potentially toxigenic cyanobacteria ranged from
approximately 54,000 to 144,000 cells/mL. Although a visible bloom was present and
contained cyanobacterial species capable of producing microcystin, microcystin
concentrations in water during the study ranged from 2 to 5 |_ig/L. Recreational users of
the lake at the time of the bloom had no detectable microcystin in their blood and did not
report an increase in GI, dermal, respiratory, or neurological symptoms after spending
time on the lake. Adenoviruses (level of detection [LOD] = 1,250 gene copy equivalents)
and enteroviruses (LOD = 200 plaque forming units/10 L) were not detected in any water
sample. This study was limited in the number of participants and included a limited
number of exposure days in the analysis. Given a small number of recreators exposed to
low levels of microcystin over the course of 3 study days, the lack of significant
associations is not surprising.
• In a similar study conducted by this same author at three lakes in California, microcystin
concentrations from personal air samples ranged from the limit of detection (0.1 ng/m3)
to 0.4 ng/m3, and extracellular microcystin concentrations in water ranged from < 2 [j.g/L
to > 10 [j,g/L (Backer et al. 2010). No statistically significant differences were noted in
the frequency of reported GI, dermal, or respiratory symptoms between participants
immediately after they engaged in direct- or indirect-contact recreational activities in the
lake with a cyanobacterial bloom and those in a lake without a cyanobacterial bloom. The
study design characterized the potential inhalation of aerosolized microcystin among
people who recreated at the lakes and included blood assays for microcystin among the
study participants. Adenoviruses or enteroviruses were not detected at the study
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locations. This study contained a limited number of participants over the course of
3 exposure days.
• Levesque et al. (2014) conducted a prospective study of residents living in proximity to
three lakes in Canada affected by cyanobacteria to investigate the relationship between
recreational exposure, specifying full contact and limited contact with lake water and the
incidence of GI, dermal, respiratory, and other (e.g., ear pain, muscle pain) symptoms.
Full contact included swimming, waterskiing, windsurfing, use of watercraft involving
launching, accidental falls, and similar activities, and limited contact included fishing,
use of watercraft not involving launching, and other activities. No associations were
observed between any symptoms and recreational exposures to microcystins. The
maximum microcystin concentrations for which recreational-related GI symptoms were
reported was 7.65 |_ig/L. The authors did observe a relationship between cyanobacterial
cell counts and gastrointestinal illness with a significant association above
20,000 cells/mL.
• Additional outbreak and case reports document health effects following exposure to
cyanotoxins. In a case report, acute intoxication with microcystin-producing
cyanobacteria blooms in recreational water was reported in Argentina in 2007 (Giannuzzi
et al. 2011). A single person was immersed in a Microcystis bloom containing 33,680 and
35,740 cells/mL. A level of 48.6 [j,g/L of microcystin-LR concentrations was detected in
water samples associated with the bloom. After 4 hours of exposure, the patient exhibited
fever, nausea, and abdominal pain, and 3 days later, presented dyspnea and respiratory
distress and was diagnosed with an atypical pneumonia. One week after the exposure, the
patient developed a hepatotoxicosis with a significant increase of alanine
aminotransferase, aspartate aminotransferase, and y-glutamyltransferase. The patient
completely recovered within 20 days.
• Dziuban et al. (2006) and Hilborn et al. (2014) reported nine outbreaks associated with
recreational exposure to HABs in which microcystins were detected, one in 2004 and
eight in 2009 and 2010. In the one outbreak in which microcystin was measured at
20.8 |ig/L and other cyanotoxins were either not detected or measured, 9 cases reported
symptoms, which included abdominal cramps (3 cases), diarrhea (3), nausea (3) vomiting
(2), fever (2), headache (2), rash (8), eye irritation (1), earache (1), neurologic symptoms
(2), tingling (2), confusion (1), and respiratory symptoms (1) (Hilborn et al. 2014).
Cyanobacterial cells were present. The results reported from the outbreaks should not be
interpreted as cause and effect. Rather, the stressors and health endpoints discussed can
be considered a co-occurrence due to the nature of the data collated in the outbreak
reports.
Cylindrospermopsin
No epidemiological studies were found for recreational exposure to cylindrospermopsin.
Hilborn et al. (2014) reported two outbreaks associated with recreational exposure to
HABs in which cylindrospermopsin was detected between 2009 and 2010. Cyanobacteria,
microcystins, and other cyanotoxins, however, also were detected in these two outbreaks. As
mentioned above, the results reported from the outbreaks should not be interpreted as cause and
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effect, only that two or more parameters were demonstrated to co-occur spatially and/or
temporally.
5.1.1.3 Noncancer Mode of Action
Microcystins
Mechanistic studies have shown the importance of membrane transporters for systemic
uptake and tissue distribution of microcystin by all exposure routes (Feurstein et al. 2010;
Fischer et al. 2005). The importance of the membrane transporters to tissue access is
demonstrated when a reduction in, or lack of, liver damage happens following OATp inhibition
(Hermansky et al. 1990a; Hermansky et al. 1990b; Thompson & Pace 1992).
The uptake of microcystins causes protein phosphatase inhibition and a loss of
coordination between kinase phosphorylation and phosphatase dephosphorylation, which results
in the destabilization of the cytoskeleton. This event initiates altered cell function followed by
cellular apoptosis and necrosis (Barford et al. 1998). Both cellular kinases and phosphatases keep
the balance between phosphorylation and dephosphorylation of key cellular proteins controlling
metabolic processes, gene regulation, cell cycle control, transport and secretory processes,
organization of the cytoskeleton, and cell adhesion. Each of the microcystin congeners evaluated
(LR, LA, and LL) interacts with catalytic subunits of protein phosphatases PP1 and PP2A,
inhibiting their functions (Craig et al. 1996).
As a consequence of the microcystin-induced changes in cytoskeleton, increases in
apoptosis and reactive oxygen species (ROS) occur. In both in vitro and in vivo studies, cellular
pro-apoptotic Bax and Bid proteins increased while anti-apoptotic Bel-2 decreased (Fu et al.
2005; Huang et al. 2011; Li et al. 201 la; Takumi et al. 2010; Weng et al. 2007; Xing et al. 2008).
Mitochondrial membrane potential and permeability transition pore changes (Ding & Nam Ong
2003; Zhou et al. 2012) lead to membrane loss of cytochrome c, a biomarker for apoptotic
events. Wei et al. (2008) identified a time-dependent increase in ROS production and lipid
peroxidation in mice after exposure to microcystin-LR. After receiving a 55 (J,g/kg of body
weight i.p. injection of microcystin-LR, the levels of hepatic ROS increased rapidly within
0.5 hours and continued to accumulate for up to 12 hours in a time-dependent manner.
Cylindrospermopsin
Despite the number of studies that have been published, the mechanisms for liver and
kidney toxicity by cylindrospermopsin are not completely characterized.
The occurrence of toxicity in the liver suggests a protein-synthesis inhibition mechanism
of action for cylindrospermopsin. In vitro and in vivo studies have been conducted to
demonstrate the ability of cylindrospermopsin to inhibit hepatic protein synthesis, which could
impact mouse urinary protein production leading to decreased urinary excretion of these proteins
(Froscio et al. 2009; Froscio et al. 2008; Terao et al. 1994). Available evidence indicates that
protein synthesis inhibition is not decreased by broad-spectrum CYP450 inhibitors, but they do
reduce cytotoxicity (Bazin et al. 2010; Froscio et al. 2003). Hepatotoxicity appears to be
CYP450-dependent, which indicates a possible involvement of oxidized and/or fragmented
metabolites and mechanisms other than protein synthesis inhibition (Froscio et al. 2003;
Humpage et al. 2005; Norris et al. 2002; Norris et al. 2001).
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In the Reisner et al. (2004) report, microscopic examination of blood samples showed the
presence of red blood cells with spiked surfaces rather than their normal biconcave-disc shape.
The authors attributed the acanthocyte formation to an increase in the cholesterol to phospholipid
ratio of the red blood cell membrane. Phospholipids constitute the matrix material of cell
membranes. The authors hypothesized that this change was the consequence of decreased
activity of plasma lecithin cholesterol acyl transferase, an enzyme associated with high-density
lipoproteins and the esterification of plasma cholesterol. Effects on the cholesterol content of the
red blood cell membrane can occur with inhibition of the enzyme increasing membrane fluidity
and mean corpuscular volume. Removal of the abnormal blood cells by the spleen increases both
spleen weight and serum bilirubin as well as stimulates hematopoiesis. Additional research is
needed to examine the lecithin cholesterol acyl transferase enzyme inhibition hypothesis in order
to confirm whether it accounts for the effects on the red blood cell as a result of
cylindrospermopsin exposure.
Kidney necrosis and a decreased renal failure index at the high cylindrospermopsin doses
provide support for the effects on the kidney. Numerous signs of renal damage including
proteinuria, glycosuria, and hematuria were observed after a hepatoenteritis-like outbreak in
Palm Island, Australia in 1979 (Byth 1980). The outbreak was attributed to consumption of
drinking water with a bloom of C. raciborskii, a cyanobacteria that can produce
cylindrospermopsin. These effects are associated with impaired kidney function (Byth 1980);
however, no mode of action information for kidney effects was observed in the available animal
studies of cylindrospermopsin. Since all the studies were conducted in mice, a species that
excretes low molecular weight proteins in urine, there is a need to conduct a study of
cylindrospermopsin in a laboratory species that does not excrete protein in the urine in order to
determine whether there are comparable effects on kidney weight, protein excretion, and renal
cellular damage.
5.1.2 Cancer
5.1.2.1 Weight of Evidence Classification
While there is evidence of an association between liver and colorectal cancers in humans
and microcystins exposure and some evidence that microcystin-LR is a tumor promoter in
mechanistic studies, there is inadequate information to assess carcinogenic potential of
microcystins in humans (U.S. EPA 2005). The human studies are limited by lack of exposure
information and the uncertainty regarding whether or not these studies adequately controlled for
confounding factors such as hepatitis B infection. No chronic cancer bioassays for microcystins
in animals are available. U.S. EPA (2005) states that the descriptor of "inadequate information to
assess carcinogenic potentiaF is appropriate when available data are judged inadequate for
applying one of the other descriptors or for situations where there is little or no pertinent
information or conflicting information. The guidelines also state that (p. 2-52) "Descriptors can
be selected for an agent that has not been tested in a cancer bioassay if sufficient other
information, e.g., toxicokinetic and mode of action information, is available to make a strong,
convincing, and logical case through scientific inference." In the case of microcystins, the data
suggest that microcystin-LR may be a tumor promoter but not an initiator. Without stronger
epidemiological data and a chronic bioassay of purified microcystin-LR, the data do not support
classifying microcystin-LR as a carcinogen. The International Agency for Research on Cancer
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(IARC) classified microcystin-LR as a Group 2B (possibly carcinogenic to humans) based on the
conclusion that there was strong evidence supporting a plausible tumor promoter mechanism for
these liver toxins (IARC 2010).
No chronic cancer bioassays of cylindrospermopsin were located in the literature.
Limited data from an in vivo study showed no indication that the cyanobacterial extract
containing cylindrospermopsin-initiated tumors in mice (Falconer & Humpage 2001).
5.2 Dose-Response Assessment
The RfD value for microcystin for this recreational AWQC is from EPA's Health Effects
Support Document for the Cyanobacterial Toxin Microcystins (U.S. EPA 2015d), where
additional details are available. EPA identified a study by Heinze (1999) as the critical study in
which male hybrid rats were administered microcystin-LR in drinking water at doses of 0
(n = 10), 50 (n = 10) or 150 (n = 10) (J,g/kg body weight for 28 days (Heinze 1999). The RfD of
0.05 |ig/kg/d derived for microcystins was based on observed liver effects that included
increased liver weight, slight to moderate liver necrosis lesions (with or without hemorrhages at
the low dose and increased severity at the high dose), and changes in serum enzymes indicative
of liver damage.
The RfD value for cylindrospermopsin for this recreational AWQC is from EPA's Health
Effects Support Document for the Cyanobacterial Toxin Cylindrospermopsin (U.S. EPA 2015 c),
where additional details are available. EPA identified a study by Humpage and Falconer (2002;
2003) as the critical study in which male Swiss albino mice were administered purified
cylindrospermopsin in water via gavage at doses of 0, 30, 60, 120, or 240 j_ig/kg/d for 11 weeks.
The RfD of 0.1 |ig/kg/d derived for cylindrospermopsin was based on increases in relative
kidney weights along with indicators of reduced renal function effects at higher doses and
decreased urinary protein.
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6.0 SWIMMING ADVISORY AND RECREATIONAL CRITERIA DERIVATION
This section summarizes the inputs and shows the calculation for the recreational criteria
and swimming advisory for microcystins and cylindrospermopsin.
6.1 Microcystins Magnitude
The magnitude of the swimming advisory and recreational criteria for microcystin toxins
is calculated as follows:
Recreational value (ng/L) = RfD
RSC x BW
Ingestion Rate
Where:
RfD (|ig/kg/d)
RSC
BW (kg)
Ingestion rate
(L/d)
0.05 ng/kg/d (U.S. EPA 2015d)
0.8 (U.S. EPA 2000a)
mean body weight of children 6 to < 11 years (31.8 kg)
(U.S. EPA 2011)
recreational water incidental ingestion rate for children
(0.33 L/d) at approximately the 90th percentile (U.S. EPA
2011; U.S. EPA 1997)
0.8 X 31.8 kg
Microcystins recreational value = 0.05 |j,g/kg/d x = 4 |j,g/L
0.33 L/d
6.2 Cylindrospermopsin Magnitude
The magnitude of the recreational criteria and swimming advisory values for
cylindrospermopsin is calculated as follows:
RSC x BW
Recreational value (|_ig/L) = RfD
Where:
RfD (|ig/kg/d)
RSC
BW (kg)
Ingestion rate
(L/d)
Ingestion Rate
0.1 (U.S. EPA 2015c)
0.8 (U.S. EPA 2000a)
mean body weight of children 6 to < 11 years (31.8 kg)
(U.S. EPA 2011)
recreational water incidental ingestion rate for children
(0.33 L/d), at approximately the 90th percentile (U.S. EPA
2011; U.S. EPA 1997)
0.8 x 31.8 kg
Cylindrospermopsin recreational value = 0.1 (ig/kg/d x —q 33 — = ^ Mg/L
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6.3 Recommended Swimming Advisory and Recreational Criteria for Microcystins and
Cylindrospermopsin
Recreational criteria and the swimming advisory include a magnitude, duration, and
frequency. Magnitude is the numeric expression of the maximum amount of the contaminant that
may be present in a waterbody that supports the designated use, in this case protecting public
health of recreators. Duration is the period of time over which the magnitude is calculated.
Frequency of excursion describes the maximum number of times the pollutant may be present
above the magnitude over the specified time period (duration). The magnitude, duration, and
frequency in combination protect the designated use (in this case primary contact recreation).
EPA requests public comment on all three of these recommendations.
The magnitude values are based on body weight and intake in children and are considered
protective of adverse health effects for adults. To protect public health of swimmers at a beach,
EPA recommends that the magnitude of the advisory value not be exceeded on any single day.
For adoption as a recreational water quality criterion, EPA recommends using an excursion
frequency of no greater than 10 percent of days per recreational season (up to one year), which is
similar to recommendations for other recreational criteria (U.S. EPA 2012a). The 10 percent
exceedance rate can help inform decisions on identifying impaired and threatened waters. The
seasonal assessment period can take into consideration the temporal variability of HABs in the
waterbody. For example, HABs can occur in some waterbodies earlier and later in the
recreational season, while in other waterbodies HABs can occur and persist as long as conditions
are conducive to their growth. HABs that produce toxins that last for extended periods or that
reoccur across years when conditions are conducive to cyanobacterial growth can signify
waterbodies with excessive nutrient loadings. EPA does not anticipate states using these
cyanotoxin recommendations alone for developing load allocations for Total Maximum Daily
Loads (TMDLs) or for Water Quality-Based Effluent Limits (WQBELs). For permitting
purposes, cyanobacteria or their toxins are not typically present in permitted discharges. Permits
are more likely to be written to address point source discharges of the causal pollutants, such as
nutrients, on a waterbody-specific or watershed basis, where the permit writer has determined
there is a reasonable potential for the causal pollutants in the discharge to cause or contribute to
an exceedance of the cyanotoxin standards.
The recommended recreational criteria or swimming advisory values for the cyanotoxins
microcystins and cylindrospermopsin are presented in Table 6-1.
Table 6-1. Recreational Criteria or Swimming Advisory Recommendations for
Microcystins and Cylindrospermopsin
Application of
Recommended
Values
Microcystins
Cylindrospermopsin
Magnitude
Oig/L)
Frequency
Duration
Magnitude
fag/L)
F rcqucncy
Duration
Swimming
Advisory
4
Not to be
exceeded
One day
8
Not to be
exceeded
One day
Recreational
Water Quality
Criteria
No more than
10 percent of
days
Recreational
season (up to
one calendar
year)
No more than
10 percent of
days
Recreational
season (up to
one calendar
year)
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As an example:
• To protect swimmers, the concentration of total microcystins shall not exceed
4 micrograms per liter in a day.
• To protect the recreational use, the concentration of total microcystins shall not exceed
4 micrograms per liter more than 10 percent of days in a recreational season.
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7.0
EFFECTS CHARACTERIZATION
7.1 Cyanobacterial Cells
Cyanobacterial cell densities can indicate the eutrophic status of a water body, especially
when considering the frequency and severity of HAB occurrence (Yuan & Pollard 2015). Thus,
cyanobacterial cell densities, especially the extent, severity, and frequency of blooms under
environmental conditions conducive to cyanobacterial cell growth, are an indicator of the
ecological health of a water body.
Cyanobacterial cells are associated with two distinct sets of health endpoints. First,
cyanobacteria are associated with toxin-related endpoints. The second set of health effects
associated with cyanobacterial cells are the inflammatory health endpoints including rashes,
respiratory and gastrointestinal distress, and ear and eye irritation, which may be instigated by
direct contact with the cells, bioactive compounds in the cyanobacteria not currently classified as
toxins, or by contact with cyanobacteria-associated microbial commensals via dermal, oral
and/or inhalation exposure routes (Eiler and Bertilsson 2004; Gademann and Portman 2008).
Such effects have been observed in various health studies, including epidemiological and clinical
studies and outbreak reports (Bernstein et al. 2011; Geh et al. 2015; Levesque et al. 2014; Lin et
al. 2015; Pilotto et al. 1997; Pilotto et al. 2004; Stewart et al. 2006a,b). Also, while not all
cyanobacteria produce cyanotoxins, scientists have observed a relationship between
cyanobacteria density and cyanotoxin concentration (Loftin et al. 2016b). Environmental
conditions and ecosystem interactions also affect the production and release of cyanotoxins into
ambient waters. Cyanobacterial cell densities can be an indicator of the potential of a bloom to
produce cyanotoxins. While there is uncertainty and variability associated with the propensity of
a bloom to produce cyanotoxins, cell densities can be used to estimate the potential for
cyanotoxin concentrations to exceed the recommended values presented in section 6.
7.1.1 Cyanobacterial Cells Related to Inflammatory Health Effects
Various health studies, described in more detail in Appendix D, relate recreational
exposure to cyanobacterial cells with specific health endpoints that can be described as acute
inflammatory or allergenic reactions. It is possible that these endpoints could be related to other
biological or biochemical mechanisms that are not yet understood. Studies have (1) examined the
epidemiological relationships of recreational exposure to cyanobacteria in the water to recreator-
reported symptomologies, (2) characterized the allergenic and dermal reactions to exposed
animals and humans in clinical and in vitro studies, and (3) collated information on illness
outbreaks associated with recreational exposure to HABs. However, the reported health
endpoints and cyanobacterial density associated with the inflammatory response health outcomes
are not consistent. Empirical study differences, such as study size, species, strains of
cyanobacteria present, and measurement of possible co-exposures and the cyanobacterial
densities associated with significant health effects, lead to uncertainties in determining what
level of cyanobacteria result in a specific level of inflammatory responses in these studies. The
lack of a described dose-response characterizing cell-related inflammatory health effects could
suggest a "threshold" rather than a specific dose-response relationship (Cochrane et al. 2015;
Stewart et al. 2006b). Allergy is an example of a threshold mechanism, meaning that there is a
level of exposure (i.e., a threshold value) below which the development of sensitization and the
elicitation of an allergic reaction will not occur. Defining accurate numerical values for threshold
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exposure levels is difficult due to lack of validated methods and uncertainties about the
mechanism of sensitization (Cochrane et al. 2015).
WHO recommends the use of cyanobacterial cell densities related to an increasing scale
of the probability of adverse health effects in the Guidelines for Safe Recreational Water
Environments (WHO 2003b). They also estimated microcystin concentrations that could be
associated with these cell density levels. WHO used this approach to differentiate "between the
chiefly irritative symptoms caused by unknown cyanobacterial substances and the potentially
more severe hazard of exposure to high concentrations of cyanotoxins, particularly
microcystins." Therefore, WHO recommended a series of three guideline values associated with
incremental severity and probability of health effects rather than a single guideline value. The
WHO guideline values are:
• Low probabilities of adverse health effects (20,000 cells per mL) can be associated with
irritative or allergenic effects from exposure to cyanobacterial cells (corresponding to
10 |ig chlorophyll a per liter, under conditions of cyanobacterial dominance). The Pilotto
et al. (1997) epidemiological study directly informed the derivation of this cut point.
They also estimated that microcystin concentrations of 2-4 |ig/L can be expected at this
level.
• Moderate probability of the adverse health effects (i.e., 100,000 cells per mL) (equivalent
to approximately 50 |ig chlorophyll a per liter, if cyanobacteria dominate) is associated
with an increased potential for irritative health outcomes and the potential for negative
health impacts associated with exposure to higher cyanotoxin concentrations. The
100,000-cell cut point was informed by (1) modifying the value for the WHO drinking-
water guideline for microcystin-LR for a recreational exposure scenario and (2)
translating microcystin concentrations to cell densities based on the average microcystin
content of Microcystis cells (equivalent to 20 |ig microcystin/L). The WHO estimated that
"at a cell density of100,000 cells per mL, there is the potential for some frequently
occurring species (i.e., microcystis) to form scums, " which can "increase risks for bathers
and others involved in body-contact water sports. "
• The high probability of adverse health effects category is associated with the elevated
potential for exposure to cyanotoxins and the potential for severe health outcomes. "The
presence of cyanobacterial scum in swimming areas represents the highest risk of
adverse health effects due to abundant evidence for potentially severe health outcomes
associated with these scums" (estimated at 50-100 jug microcystin/L).
Epidemiological studies, clinical studies, and recreational water outbreak reports were
identified during searches of the publicly available and peer-reviewed scientific literature that
characterized the human health effects associated with recreating in surface waters where
cyanobacteria were present (see Appendix D). Although these epidemiological studies provide
evidence for statistically significant associations between cyanobacterial cell densities and
possible inflammatory or allergenic health endpoints (Levesque et al. 2014; Levesque et al.
2016; Lin et al. 2015; Pilotto et al. 1997; Stewart et al. 2006b), they do not provide consistent
evidence of associations either at similar densities of cyanobacterial cells or with the associated
health endpoints. The wide range in cyanobacterial cell densities associated with various health
outcomes, either with specific health endpoints or with combined symptom categories, implies
potential variability in the stressor occurrence. Differing cyanobacterial community composition
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and proportions of the more allergenic, non-cyanotoxin-producing strains relative to the
cyanotoxin-producing strains at each site is a factor. Additionally, potential variability in
sensitivity in the study populations, differences among the specific sites studied (e.g., fresh
versus marine beaches), and uncertainty with the potential confounding effects of other microbes
that can co-occur with cyanobacteria were some uncertainties associated with these data.
Additional uncertainties are described below.
The limited size of some studies could have affected the ability to detect significantly
increased rates of illness in individual symptom categories (Pilotto et al. 1997; Stewart et al.
2006b). Small sample size can diminish the statistical power of the study and the ability to detect
an association if one exists (Rothman et al. 2008). The incomplete characterization or
consideration of frank or opportunistic pathogens that could co-occur with cyanobacteria in
ambient waters also could complicate conclusions related to the etiologic agent of the reported
symptoms (Levesque et al. 2014; Lin et al. 2015; Pilotto et al. 1997; Stewart et al. 2006b).
Variability in the reported associations, including with the range of cyanobacterial cell
densities reported and with specific symptom categories, affected the ability to identify a discrete
cyanobacterial cell density value that would provide a consistent level of protection across
different waters. Pilotto et al. (1997) reported a significant association with the occurrence of one
or more symptoms, such as skin rashes, eye irritation, ear irritation, gastrointestinal distress,
fever and respiratory symptoms, and exposure to > 5,000 cells/mL for > 1 hour. Levesque et al.
(2014) observed a significant increase in GI symptoms associated with recreational contact. The
increase in GI symptoms was significant in the > 20,000-cells/mL and > 100,000-cells/mL
categories, and the positive trend for increasing illness with increased cyanobacterial cell
densities also was significant at/? = 0.001. Pilotto et al. (1997), however, in discussing the
significance of the trend of increasing symptom occurrence and with the 5,000 cells/mL cut
point, specifically suggested that the 20,000 cell/mL threshold might be too high to be
adequately protective of recreators (Pilotto et al. 1997). Lin et al. (2015) reported significant
associations between respiratory symptoms and exposure to the 25th to 75th percentile range of
cyanobacterial cells excluding picocyanobacteria (range 37-237 cells/mL) and between reported
respiratory, rash, and earache symptoms and exposure to the highest quartile (range 237-
1,461 cells/mL). The 1,461-cells/mL value was the highest cell density observed in that study
(Lin et al. 2015).
Cyanobacterial cell densities reported in the literature are used by states to provide "safe
to swim" decisions by state and local health departments (see Table 2-3 for a list of states with
cyanobacterial cell density guidelines; see Appendix B for state guidelines and associated
actions). Due to the uncertainties associated with delineating discrete cyanobacterial densities
associated with a specified level of protection for recreators, EPA is not recommending CWA
304(a) criteria that include quantitative cyanobacterial cell densities predictive of the
inflammatory or allergenic health outcomes because available data do not support a consistent
quantitative dose-response relationship at this time. However, EPA recognizes that studies
examining the potential health effects associated with exposure to cyanobacterial cells
demonstrate that exposure to the cells—particularly via dermal and inhalation exposure—can be
associated with numerous health endpoints potentially characterized as inflammatory responses.
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7.1.2 Cyanobacterial Cells as Indicators for Potential Toxin Production
Available information suggests that cyanobacterial cell density could be used as an
indicator of the potential for a cyanobacterial HAB to produce cyanotoxins at the concentrations
discussed in section 6. Although EPA is not recommending criteria at this time that address
inflammatory health effects based on cyanobacterial cell density, many states already use cell-
based guidelines based on recommendations from the World Health Organization (WHO 2003a).
States use the cell density information gleaned from their monitoring efforts to inform decision-
making. Also, remote sensing techniques using satellite-based imagery to observe cyanobacterial
blooms are of increasing interest to states (Schaeffer et al. 2012; 2013). This approach detects the
level of chlorophyll a or phycocyanins in the water and converts that to a cell density estimate.
Therefore, a cell density value corresponding to the cyanotoxin criteria value is needed to
interpret the remote sensing data. Below, EPA has used a similar approach as WHO to calculate
a cyanobacterial cell density with the potential to produce the cyanotoxin at the criteria
concentration.
The WHO guidelines were developed for microcystin and cyanobacterial cell density at
different probabilities of adverse health effects to support management of recreational waters.
The WHO designated a low probability of adverse health effects category associated with the
cyanobacterial cell-related inflammatory response health endpoints (see section 7.1.1). The
probability of adverse health effects increased to moderate and high levels based on the risk
associated with the potential of the cyanobacterial cells to produce microcystin. For example, at a
level of 100,000 cyanobacterial cells per mL, WHO estimated that a concentration of 20 jag
microcystin per L is possible if those cells were predominantly Microcystis sp. and each cell
contained an average of 0.2 pg microcystin per cell (WHO 2003).
Using this approach, EPA calculates a cyanobacterial density associated with the
recommended microcystins criteria/ swimming advisory concentration as follows:
Cyanobacterial Cell Density (CCD)
Ambient cyanotoxin concentration (ACC)
Cell toxin amount (CTA)
Where:
CCD
calculated cell density associated with a specific toxin
concentration
ACC
specific toxin concentration target in ambient water (e.g., AWQC
value)
CTA
amount of toxin produced in a cyanobacterial cell
For the microcystin produced by Microcystis sp.:
ACC
CTA
4 (J,g/L; recommended recreational criteria value for microcystins
0.2 pg/cell; reported mean concentration of microcystin in a cell of
Microcystis species
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Adding in the conversion factors to convert units, the equation is:
CCD = ACC (ng/L) x 106 pg/|ig L
CTA (0.2 pg/cell) 1,000 mL
Adding in the values,
4 |j,g/L x 106 pg/|J,g 1 L
CCD = — , ,f x = 20,000 cells/mL
0.2 pg/cell 1,000 mL
Thus, a Microcystis sp. cell density of 20,000 cells/mL has the potential to result in a microcystin
concentration of 4 (J,g/L.
There is variability in the estimate of cyanotoxin concentrations associated with cell
density. WHO acknowledged that various cyanobacterial species could contain more or less
microcystin per cell. Species that contain more microcystin could result in much higher water-
column concentrations of the cyanotoxin at a similar cyanobacterial cell density. Cyanobacterial
community differences between locations could affect the level of cyanotoxin that is present. For
example, WHO discussed that a bloom dominated by Planktothrix could result in 10 to 20 times
higher water-column cyanotoxin concentrations given the same cell density (WHO 2003b). The
same cell density applied at different locations could result in inconsistent levels of health
protection for recreators at those locations.
EPA surveyed the published peer-reviewed scientific literature for information on the
amount of microcystin and cylindrospermopsin produced by or contained in a cell from a variety
of freshwater blooms reported around the world. Laboratory-based culture studies with
numerous clones of Microcystis aeruginosa, Cylindrospermopsis raciborskii, Planktothrix
agardhii, and Planktothrix rubescens were also found. Many of these references also included
either biomass-toxin conversions or graphic data which would support conversion factors from
cyanobacterial cell density (expressed in a variety of units including cells L"1, biovolume (|im3)
L"1, chlorophyll a L"1) to toxin concentrations for these species. Cyanotoxin concentration is
generally related to cyanobacterial cell abundance, which is determined by nutrient availability
(Welker 2008), so nutrient concentration is often correlated to cyanotoxin concentration.
Information gleaned from this literature search also suggests that cyanotoxin amounts can vary
with genetic factors (i.e., some isolates lack the genes involved in toxin production,
physiological factors (e.g., growth rate, growth stage, photosynthetic rate, and allelopathic
factors), trophic factors (e.g., grazing interactions), and environmental factors (e.g., temperature,
salinity, carbon dioxide concentration, light intensity, macronutrient [i.e., nitrogen, phosphorus]
and micronutrient [e.g., trace metal concentrations]). Most data available are for microcystins
rather than for cylindrospermopsin. Please refer to Appendix F for additional information on
cyanotoxin amounts per cell and conversion factors found in the literature survey.
States that currently have guidelines for HABs in recreational waters consider cell
densities, cyanotoxin concentrations, or both. Decisions to issue recreational water
warnings/advisories, or initiate monitoring for cyanotoxins based on the cyanobacterial cell
density only once a bloom is observed (i.e., green, discolored water and/or scum
formation/accumulation associated with high densities of cells) may overlook situations where
extracellular toxins are present. Cells may accumulate in locations different from where the
bloom originated (e.g., by wind and/or wave action, or transport downstream). A cell density of
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20,000 cells/mL (corresponding to the recommended AWQC value) is lower than that typically
associated with a bloom (WHO 2003b). Decision points contingent on visually confirmed
blooms may miss or delay the identification of the hazardous condition associated with exposure
to elevated cyanotoxins. States may wish to consider using visual identification of blooms
preferentially for waterbodies with a previous history of HAB events and/or microcystin
detections.
7.2 Enhanced Risk or Susceptibility
Children recreating are likely to spend more time in direct contact with waters and
measured incidental ingestion data while swimming indicate that children between 6 and
11 years ingest on average more water than older children and adults (Evans et al. 2006). No
measured incidental ingestion data are available for children younger than 6 years old. A study
by Schets et al. (2011), described in more detail in section 7.3, provides incidental ingestion
volumes for children ages 0 to 14 years, but this study relied on surveyed parents' estimates of
the amount their children incidentally ingested. Although this study used a qualitative approach
that is less certain than the studies that used analytical methods, Schets et al. (2011) identified
an average incidental ingestion volume for children aged 0 to 14 years that was the same as the
mean ingestion volume reported by Dufour et al. (2006) for children aged 6 to 18 years (37 mL).
Children ages 5 to 11 years also tend to spend more time in the water compared to younger and
older life stages (U.S. EPA 1997). The significant differences between life-stages in the volume
of water ingested while recreating and duration of exposure can translate to increased risk of
exposure to cyanotoxins for children compared to adults.
Based on the available studies in animals, individuals with liver and/or kidney disease
may be more susceptible than the general population since the detoxification mechanisms in the
liver and impaired excretory mechanisms in the kidney may be compromised. Data from an
episode in a dialysis clinic in Caruaru, Brazil where microcystins (and possibly
cylindrospermopsin) were not removed by treatment of dialysis water, identify dialysis patients
as a population of potential concern in cases where the drinking water source was contaminated
with cyanotoxins.
The data on red blood cell acanthocytes suggest that individuals that suffer from anemia
(e.g., hemolytic or iron-deficiency) might be a potentially sensitive population. Several rare
genetic defects such as abetalipoproteinemia (rare autosomal recessive disorder that interferes
with the normal absorption of fat and fat-soluble vitamins from food) and
hypobetalipoproteinemia are associated with abnormal red blood cell acanthocytes, which
appears to result from a defect in expression of hepatic apoprotein B-100, a component of serum
low density lipoprotein complexes (Kane & Havel 1989). Individuals with either condition might
be sensitive to exposure to cylindrospermopsin.
Available animal data are not sufficient to determine if there is a definitive difference in
the response of males versus females following oral exposure to microcystins. Fawell et al.
(1999) observed a slight difference between male and female mice in body weight and serum
proteins, but no sex-related differences in liver pathology. Available animal data are not
sufficient to determine if there is a definitive difference in the response of males versus females
following oral exposure to cylindrospermopsin.
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7.3 Other Studies of Ingestion While Swimming
EPA used the recommended incidental ingestion while recreating values discussed in the
Exposure Factors Handbook (2011), which cites Dufour et al. (2006) as the basis for its default
recreational ingestion values. Dufour et al. (2006) measured the incidental ingestion of water
while participants were swimming in a pool and found that children under the age of 18 ingested
higher volumes of water while swimming than adults. The values presented in EPA's Exposure
Factors Handbook (2011) adjusted the Dufour et al. (2006) data from a per event basis to an
hourly ingestion rate.
In addition to Dufour et al. (2006), five other studies (Dorevitch et al. 2011; Evans et al.
2006; Schets et al. 2011; Schijven & de Roda Husman 2006; Suppes et al. 2014) evaluated
recreation-associated incidental ingestion. See Table 7-1 for a summary overview of the
available studies of incidental ingestion while recreating.
Evans et al. (2006) presented results from an observational study of incidental water
ingestion during recreational swimming activities using the same methodology as the Dufour et
al. (2006) pilot study. They cited the methods published in the Dufour et al. (2006) pilot study,
which involved using cyanuric acid as an indicator of pool water ingestion to estimate the
amount of water ingested by boys (n = 107) and girls (n = 80) ages 6-18 years who were
directed to stay in the pool and actively swim for 45 to 60 minutes. Evans et al. (2006) reported
that children ages 6-18 years incidentally ingested a mean volume of 47 mL per swimming
event (boys: 48 mL/event; girls: 47 mL/event). Consistent with Dufour et al. (2006), Evans et
al. (2006) found that children ingested higher volumes of water than both adults and the entire
study population combined. Adults (both genders combined) incidentally ingested a mean
volume of 24 mL. Adult men and adult women incidentally ingested 30 mL and 19 mL,
respectively. The entire study population had a mean incidental ingestion volume of 32 mL. The
Evans et al. (2006) reported study has not been peer reviewed.
Suppes et al. (2014) evaluated incidental water ingestion rates using cyanuric acid as an
indicator of pool-water ingestion, and found that children on average ingested pool water at a
higher rate than adult participants. Total time in water, quantified by viewing videos, was used
to adjust pool-water ingestion volumes to obtain rates. After adjustments for false-positive
measurements were applied, the mean rate at which adults ingested water was 0.0035 L/hr with
range 0-0.051 L. The mean rate at which children ingested water was 0.026 L/hr with range
0.0009-0.106 L/hr.
Taking a different approach, a study in the Netherlands by Schets et al. (2011) used
questionnaires to collect estimates of water swallowed while swimming/bathing in freshwater,
marine water, and swimming pools and found children had higher ingestion volumes. Two
rounds of surveys were conducted, one in 2007 and another in 2009. Of the 8,000 adults who
completed the questionnaire, 1,924 additionally provided estimates for their eldest child
(<15 years of age). The participants estimated the amount of water they or their children
swallowed while swimming. Schets et al. (2011) also conducted a series of experiments to
measure the amount of water that corresponded to a mouthful of water and converted the data in
the four response categories to volumes of water ingested. Depending on the water type, adult
men swallowed, on average, 0.027-0.034 L per swimming event and women swallowed 0.018-
0.023 L. Children swallowed more than adults on average, 0.031-0.051 L per swimming event
(Schets et al. 2011). Although the incidental ingestion data reported by Schets et al. (2011) were
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based primarily on participant-reported estimates, the mean values were similar to those reported
in Dufour et al. (2006).
Schijven and de Roda Husman (2006) studied sport and occupational diver incidental
ingestion. The types of water studied for occupational divers (n = 37 divers) were open sea and
coastal marine water, and freshwater. For sport divers (n = 483 divers), the types of water
considered were open sea and coastal marine water, fresh recreational water, canals and rivers,
city canals, and swimming pools. The divers were asked to estimate how much water they
swallowed in terms of: none, few drops, shot glass, coffee cup, or soda glass. The authors
translated the description of volumes from the questionnaires into average volumes.
Occupational divers reported incidentally ingesting more water per dive in marine water (mean:
0.0098 L/dive; maximum: 0.1 L/dive) compared to freshwater (mean: 0.0057 L/dive; maximum:
0.025 L/dive). Sports divers wearing an ordinary diving mask reported incidentally ingesting the
most water per dive in swimming pools (mean: 0.02 L/dive; maximum: 0.19 L/dive), followed
by recreational freshwater (mean: 0.013 L/dive; maximum: 0.19 L/dive) and coastal marine
water (mean: 0.0099 L/dive; maximum: 0.19 L/dive). Sports divers wearing a full face mask
reported incidentally ingesting less water than sports divers wearing an ordinary diving mask.
The age of the divers was not included in the study report. Duration of dives was also not
reported.
Dorevitch et al. (2011) evaluated incidental ingestion associated with multiple types of
water contact activities in both surface water and in pools. Volume of ingestion was self-reported
via interviews (3,367 participants), and the authors used a subset of the pool exposures to assess
cyanuric acid in urine to determine the accuracy of the self-reported ingestion volumes. There
was strong agreement between self-reported results and cyanuric acid measurement (none =
0.0014±0.008 L; drop to teaspoon = 0.0094±0.011 L; mouthful = 0.026±0.037 L). In surface
water, participants ages 6 and above incidentally ingested the most water while canoeing and
capsizing compared to any other activity assessed (median = 0.0036 L; mean = 0.006 L; Upper
95 percent CI = 0.0199 L). In swimming pool water, participants ages 6 and above incidentally
ingested the most water while swimming compared to any other activity assessed (median =
0.006 L; mean = 0.01 L; Upper 95 percent CI = 0.0348 L). Swimmers in a pool were more than
50 times as likely to report swallowing a teaspoon of water compared to people who canoed or
kayaked in surface waters. Duration of activities was not reported, so the ingestion volumes are
on a per event basis.
Additional estimates of incidental water ingestion rates while swimming in pools have
been identified by EPA's Office of Pesticide Programs (OPP). OPP calculates people's
exposures to pool chemicals while they swim using its SWIMODEL (U.S. EPA 2003).
SWIMODEL uses incidental ingestion values for children that are twice the values used for
adults. Incidental ingestion rates among adults while swimming competitively and
noncompetitively are 0.0125 L/hr and 0.025 L/hr, respectively. The model assumes an incidental
ingestion rate of 0.050 L/hr for children ages 7-10 and 11-14 years while swimming
noncompetitively. The 0.050-L/hr value is the value used in EPA OPP's Standard Operating
Procedures (2000b) and is based on recommendations from EPA's Risk Assessment Guidance for
Superfund, Part A (ACC 2002; U.S. EPA 1989; U.S. EPA 1997; U.S. EPA 2000b; U.S. EPA
2003). SWIMODEL assumes that noncompetitive swimmers incidentally ingest water at twice
the rate as competitive swimmers, which is based on recommendations from ACC (2002), which
is unpublished.
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Table 7-1. Studies of Incidental Ingestion Volumes or Rates While Recreating
Reference
Number of
Participants,
Water Type
Dufour et n = 53
al. (2006) Swimming
pool
Recreational
Activity
Swimming
Measurement
Methodology
Cyanuric acid was
measured in pool
water and urine
samples
Measure-
ment
Parameter
Ingestion
volume per
event
Parameter
Provided
Mean
Age Group(s)
Children (6—<18
years old)a
Adults
All ages
Value
37 mL
16 mL
32 mL
Mean
Duration
of Event
>45 min
Mean Rate
of Ingestion
(mL/hr)
49
21
43
Evans et n = >500
al. (2006) Swimming
pool
Swimming
Cyanuric acid was
measured in pool
water and urine
samples, and ingestion
rate was calculated
based on duration of
swimming event
Ingestion
volume per
event
Ingestion
rate
Mean (upper Children (6-18
95 percent CI) years old)a
Mean (95
percent CI)
Mean
Adults
6-15 years
16+ years
Children and
adults
47 mL (142 mL)
24 mL (2-84 mL)
42 mL/hr
28 mL/hr
33 mL/hr
>45 min
63
32
42
28
33
Dorevitch
etal.
(2011)
n = 3,367
Surface water
Swimming
pool
Canoeing and
capsizing
Kayaking and
capsizing
Swimming
Kayaking and
capsizing
Canoeing and
capsizing
Estimates of amount of
water swallowed were
self-reported
Estimates of amount of
water swallowed were
self-reported; cyanuric
acid was measured in
urine in a subset of
participants
Ingestion
volume per
event
Median; Mean 6+yearsb
(upper 95
percent CI)
3.6 mL;
mL)
2.9 mL;
mL)
6.0 mL;
mL)
4.8 mL;
mL)
3.9 mL;
mL)
6 mL (19.9 No duration
constraints
5 mL (16.5
10 mL (34.8 60 min
7.9 mL (26.8
6.6 mL (22.4
10
7.9
6.6
Schets et n = 8,000 Swimming
al. (2011) adults, 1,924
children
Freshwater
Marine water
Descriptive estimates
of the amount of water
swallowed were self-
reported by
participants or parents
of participants, and
estimates were
converted to volumes
Ingestion
volume per
event
Mean (95
percent CI)
0-14 years3 37 mL (0.14-170 mL) 79 min 28
Adults, males 27 mL (0.016-140 54 min 30
mL)
Adults, females 18 mL (0.022-86 mL) 20
0-14 years3 31 mL (0.08-140 mL) 65 min 29
Adults, males 27 mL (0.016-140 45 min 36
mL)
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Number of
Reference Participants,
Water Type
Swimming
pool
Recreational
Activity
Measurement
Methodology
Measure-
ment
Parameter
Parameter
Provided
Age Group(s)
Adults, females
0-14 years3
Adults, males
Adults, females
Value
18 mL (0.022-90 mL)
51 mL (0.62-200 mL)
34 mL (0.022-170
mL)
23 mL (0.033-110
mL)
Mean
Duration
of Event
41 min
81 min
68 min
67 min
Mean Rate
of Ingestion
(mL/hr)
26
38
30
21
Suppes et n = 38
al. (2014) Swimming
pool
Swimming
Cyanuric acid was
measured and total
time in water was
quantified using videos
to adjust ingestion
volumes to rates;
authors adjusted
Ingestion volumes to
correct for potential
false positive
measurements from
cyanuric acid carry-
over between sample
injections
Ingestion
rate,
adjusted
Ingestion
rate,
unadjusted
Mean
(Standard
deviation);
Range
Mean;
Maximum
Mean
Children (5-17
years old)a
Adults
Children and
adults
Children (5-17
years old)3
Adults
Children and
adults
26 mL/hr (29 mL/hr);
0.9-106 mL/hr
3.5 mL/hr (11.7
mL/hr); 0-51 mL/hr
14 mL/hr (24 mL/hr);
0-106 mL/hr
59 mL/hr; 225 mL/hr
9 mL/hr
32 mL/hr
>45 min
26
3.5
14
59
9
32
Schijven
and de
Roda
Husman
(2006)
n = 37
Freshwater
Marine water
Coastal
marine water4
n = 483
Swimming
pool
Recreational
freshwater
Coastal
marine water
Swimming
pool
Diving,
occupational
Diving,
recreational with
ordinary diving
mask
Descriptive estimates
of the amount of water
swallowed were self-
reported, and estimates
were converted to
volumes
Ingestion
volume per
event
Mean;
Maximum
Adults
5.7 mL; 25 mL
9.8 mL; 100 mL
12 mL; 100 mL
20 mL; 190 mL
13 mL; 190 mL
9.9 mL; 190 mL
13 mL; 190 mL
60-95 min 3.6-5.7
6.2-9.8
7.6-12
42-52 min
23-29
15-19
11-14
15-19
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Number of
Reference Participants,
Water Type
Recreational
Activity
Measurement
Methodology
Measure-
ment
Parameter
Parameter „ ...
„ ... Age Group(s) Value
Provided " 1 w
Mean
Duration
of Event
Mean Rate
of Ingestion
(mL/hr)
Coastal and
Diving,
1.3 mL; 15 mL
1.5-1.9
open marine
recreational with
water
full face mask
aData cannot be separated by different age groups among children.
b Results were not reported in children and adult categories.
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Although these studies used different methodologies and have limitations with respect to
reporting information for different age group categories, their results corroborate the Dufour et
al. (2006) data. Similar to Dufour et al. (2006), the studies that included children confirmed that
children ingested more than adults. The freshwater ingestion results reported by Schets et al.
(2011) included parent estimates of children's ingestion of water while swimming in freshwater
that are most similar to the Dufour et al. (2006) findings. Schets et al. (2011) found a mean
ingestion volume for children aged 0 to 14 years of 37 mL, which is the same as the mean
ingestion volume reported by Dufour et al. (2006) for children. The adult self-reported ingestion
volumes in Schets et al. (2011) were also similar to the Dufour et al. (2006) adult value. Schets et
al. (2011) reported adult values ranging from 18 and 27 mL for females and males, respectively,
while Dufour et al.'s adult ingestion volume was 16 mL. Schijven and de Roda Husman (2006)
found adult divers mean ingestion volumes while diving recreationally in a swimming pool or in
freshwater ranged between 13 and 20 mL, varying depending on mask type used. Dorevitch et al.
(2011) also evaluated self or parent estimates of ingestion volumes while swimming and found a
mean ingestion volume for all ages of 10 mL. Suppes et al. (2014) used a similar measurement
method as Dufour et al. (2006), i.e., measuring cyanuric acid as an indicator of pool water
ingestion, to estimate the amount of water ingested by 16 children ages 5 to 17 years. After
adjustment for false positives, the mean rate at which child participants ingested water was
26 mL/hr, just about half of the Dufour et al. (2006) normalized ingestion rate of 50 mL/hr.
7.4 Distribution of Potential Recreational Health Protective Values by Age
To evaluate the parameters used to calculate the cyanotoxin recreational AWQC, EPA
compiled and evaluated available information for various lifestages. This section discusses
potential health protective values for children and adults based on alternative data sets (section
7.4.1) and considers younger children's exposure parameters (section 7.4.2).
7.4.1 Evaluation of Criteria Related Lifestages
Using the ingestion rates for each age-group from EPA's Exposure Factors Handbook
(U.S. EPA 2011), EPA estimated recreational health protective values for microcystins and
cylindrospermopsin (plotted on Figure 7-1) to demonstrate the variability due to body weight,
recreational water incidental ingestion, and exposure duration by lifestage.
EPA derived the recreational AWQC based on children's recreational exposures because
this life stage has higher recreational exposures relative to adult recreators and the general
population as a whole (i.e., all ages). As Figure 7-1 demonstrates, the calculated values for
children (4 |ig/L for microcystins and 8 |ig/L for cylindrospermopsin) are protective of adults
and the general population. EPA calculated for comparison recreational health protective values
for adults using (1) 80 kg as the body weight (U.S. EPA 2011), (2) the maximum observed
incidental ingestion value for adults (0.07 L/h) which EPA's Exposure Factors Handbook (2011)
recommended due to the limited size of the data set, and (3) the mean recreational exposure
duration for the 18- to 64-year age group (2.0 hr/d) (U.S. EPA 1997). The estimated recreational
health protective values for adults are 23 |ig/L for microcystins and 46 |ig/L for
cylindrospermopsin, Therefore, the recreational criteria and swimming advisories EPA
calculated to be protective of children are protective of adults.
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Figure 7-1. Comparison of Recreational Health Protective Values for Microcystins and
Cylindrospermopsin for Children, Adults, and General Population
50
< 45 i : : : : :-
3.
a) 40 :::::::::::
Rec AWQC
Rec AWQC
Microcystins Cylindrospermopsin
¦ Children DAdults 0General Population
The parameters used to calculate health protective values for children include incidental
ingestion values for children less than 18 years, mean body weight for children ages 6 to
11 years, and recreational exposure duration for children ages 5 to 11 years. The Dufour et al.
(2006) incidental ingestion data are limited to children older than 6 years but less than 18 years.
Schets et al. (2011) surveyed individuals 15 years and older to estimate their incidental ingestion
of freshwater while recreating and asked those who had children to estimate incidental ingestion
of their oldest child aged 0 to < 15 years. The ingestion volumes were initially binned into
exposure classes and then translated into volumes using the results of a second study that
quantified the distribution of volumes associated with ingested mouthfuls of water (Schets et al.
2011). The Schets et al. (2011) results for ages 0 to < 15 years were similar to estimates for
6 to < 18 years in Dufour et al. (2006). In both studies, children ingested more than adults on
average and in the range of volumes (see Table 7-1). Based on the qualitative nature of the data
available for the youngest children and given that the mean values were similar, EPA concludes
that the values reported in the Exposure Factors Handbook are protective of children of all ages,
including those younger than 6 years.
Evans et al. (2006) reported results of a full-scale study using the same methodology
reported in Dufour et al. (2006); results of this study were presented by Evans et al. (2006) at an
EPA recreational waters conference in 2006. The full-scale study included a study population
sufficient to break out age categories that included younger children (6 to 10 years old), older
children (11 to 17 years old), and adults. The number of study participants in the Dufour et al.
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(2006) pilot study was 53, while their full-scale study evaluated more than 500 participants
(Evans et al. 2006). Similar to the results reported in the pilot study, children (<18 years old)
ingested significantly more than adults. Additionally, Evans et al. (2006) reported that younger
children (6 to 10 years old) ingest significantly more than older children (11 to 17 years old), or
adults. Data quality standards require EPA to use independently peer reviewed and published
data within our recommendations. Until this data set is published, EPA cannot include it in its
analysis.
Table 7-2 presents a comparison of the daily ingestion rates (i.e., hourly ingestion rate
times the exposure duration in hours) for the Dufour et al. (2006) study compared to Evans et al.
(2006). While rates for "children" (<18 years old) are similar between the studies, the ingestion
rates using information from the newer study and duration rates from the EPA's Exposure
Factors Handbook (2011) indicate a significantly higher exposure for younger children
compared to older children or adults. A Kruskal-Wallis statistical test indicated that ingestion
rates differed significantly between groups (p-value < 0.001). The pairwise Wilcoxon test with
Bonferroni correction also indicated that ingestion rates in younger children (aged 6 to 10) were
significantly different from ingestion rates in older children (p-value < 0.001). However, there is
no difference between ingestion rates between older children (aged 11 to 17) and adults (p-value
>0.05).
Table 7-2. Comparison of Daily Ingestion Rates While Recreating between Dufour et al.
(2006) and Evans et al. (2006)a
Ajjc
Group
Parameter Tvpeb
Dufour et al. (2006) Based
Daily Ingestion Rate (L/d)
Evans et al. (2006) Based Daily Ingestion
Rate (L/d)
Children
Mean
6 to < 18 years
0.13
0.14
6 to 10 years
0.22
11 to 17 years
0.09
Upper percentile
0.33
0.34
6 to 10 years
0.50
11 to 17 years
0.23
Adults
Mean
18+ years
0.04
0.06
Upper percentile
0.14
0.12
a The results reported in Evans et al. (2006) are not yet published. It is EPA's policy to use peer-reviewed study
results to inform its regulatory efforts. The Evans et al. (2006) results are included within this effects
characterization to provide context to the parameter values EPA used in the criteria derivation and because these
results were presented publicly at the National Recreational Water Conference in 2006.
b The calculations of daily ingestion rate all used the mean exposure duration; the parameter type refers to the hourly
ingestion rate.
Table 7-3 presents alternative recreational values derived based on the more specific
children's age groups available from Evans et al. (2006). Figure 7-2 provides a chart of this
information for comparison purposes.
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67
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Table 7-3. Alternative Recreational Criteria Values for Microcystins and
Cylindrospermopsin Calculated based on Alternative Ingestion Data from Evans et al.
(2006)
Age Group
Body Weight
(kg)
Ingestion Rate
(L/d)a
Alternative
Health Protective Recreational
Cyanotoxin Value (ju.g/L)
Microcystins
Cylindrospermopsin
Children 6 to 10 years
31.8
0.50
3
5
Children 11 to 17 years
56.8
0.23
10
20
Adults 18 to 64
80
0.12
26
52
General Population
60
0.20
12
24
a Ingestion rate is the product of incidental ingestion volume normalized to one hour (L/lir) and the recreational
duration (lir/d). Scenario uses mean body weight and ingestion rate based on upper percentile or maximum value for
ingestion rate fromEPA's Exposure Factors Handbook (U.S. EPA 2011) and mean value for recreational exposure
duration in the older version of EPA's Exposure Factors Handbook (U.S. EPA 1997).
Figure 7-2. Comparison of Alternative Health Protective Recreational Values and
Recreational AWQC for Microcystins and Cylindrospermopsin Calculated based on Evans
et al. (2006)a
55
Cj 50
~3>
3 45
a>
"I 40
>
«j 35
O
% 30
a>
Rec AWQC
Rec AWQC
Microcystins Cylindrospermopsin
¦ 6-10 years IS 11-17 years ~ 18 -81 years ~ General Population
a The results reported in Evans et al. (2006) are not yet published. It is EPA's policy to use peer-
reviewed study results to inform its regulatory efforts. The Evans et al. (2006) results are included
within this effects characterization to provide context to the parameter values EPA used in the criteria
derivation and because these results were presented publicly at the National Recreational Water
Conference in 2006.
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68
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7.4.2 Evaluation of Younger Children's Exposure Factors
In the calculation of the cyanotoxin values reported in section 6, EPA utilized exposure
parameters reported in the Exposure Factors Handbook (U.S. EPA 1997; U.S. EPA 2011).
Information on children's mean body weights were available for children's age groups including
0 to 1 year, as well as 1 to < 2 year, 2 to < 3 years, etc. Using the body weight data provided in
U.S. EPA (2011), weighted mean for the age groups 0 to < 6 years and 1 to < 6 years were
calculated.
The available values from the Exposure Factors Handbook (1997, 2011) for incidental
ingestion volume and exposure duration, however, were limited to specific age ranges. For
incidental ingestion, the data reported were limited to children 6 years old and older. U.S. EPA
(2011) recommends using the 97th percentile ingestion volume for children <18 years based on
the Dufour et al. (2006) measured incidental ingestion volume normalized to 1 hour. The Dufour
et al. (2006) study did not include children younger than 6 years. The 97th percentile is
recommended because the study had a small number of participants. U.S. EPA (1997) provided a
recreational exposure duration for children ages 1 to 4 years (1.4 hr/d). This duration is shorter
than the duration for children ages 5 to < 11 years (2.7 hr/d). Values for exposure duration were
not available for children younger than 1 year.
To evaluate potential health-protective water quality values specifically for children
younger than 6 years, EPA searched for additional exposure parameter information in the peer-
reviewed and published scientific literature. Table 7-4 shows data availability and differences
between the exposure parameters used in the microcystin and cylindrospermopsin recreational
AWQC calculation (ages 6 to < 11 years) and estimates for younger lifestages. The younger
lifestages include children aged 0 to 6 years, children aged 1 to 6 years, and children younger
than 1 year.
EPA found one other study that characterized incidental ingestion for children. Schets et
al. (2011) reported incidental ingestion volumes for children ages 0 to < 15 years. However, the
study did not further divide this cohort into younger children and older children. These data for
children represent parental estimates of volumes of freshwater incidentally ingested by their
children. The ingestion volumes were initially binned into exposure classes and then translated
into volumes using the results of a second study that quantified the distribution of volumes
associated with ingested mouthfuls of water (Schets et al. 2011). Because of the initial binning
and then the translation step to arrive at a distribution of ingestion volume for each exposure
class, there is some uncertainty associated with the estimates. However, these estimates represent
a different methodological approach compared to the approach used by Dufour et al. (2006). To
facilitate comparing the results between the studies, EPA calculated an hourly incidental
ingestion volume based on the Schets et al. (2011) data using the mean freshwater recreational
durations reported in the same study. Because this study was not limited in size, as was the case
with Dufour et al. (2006), EPA calculated both the 90th percentile and 97th percentile hourly
ingestion volume. The 97th percentile was calculated to facilitate a more direct comparison with
the results from Dufour et al. (2006) and the 90th percentile was calculated to provide a value
that would typically be used to calculate health-protective values for a pollutant (U.S. EPA
2000a).
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Table 7-4. Comparison of Younger Children's Exposure Factors and Incidental Ingestion Data Sets
I'Aposlll'C
Parameter
Body weight
(U.S. EPA 2011)
I sed in Recreational
AWQC Calculation
l)ulour el al.
Children 1 lo <
(i \ ea rs
(200(0 Incidental
Children 0 lo
< (i \ ea rs
Ingestion Data
Children
< 1 jear
Scliels el al. (
Children 1 lo <
0 jears
2011) Incidental 1
Children 0 lo <
0 j ears
ngcslion Dala
Children
< 1 jear
31.8 kg = mean body
weight of children 6 to
<11 years
15 (> ku
weighted mean
body weight of
children 1 to
< 6 years
1 ^ 4 ku
weighted mean
body weight of
children 0 to
< 6 years
- S ku
weighted mean
body weight of
children 0 to
< 1 year
15.6 kg =
weighted mean
body weight of
children 1 to
< 6 years
13.4 kg =
weighted mean
body weight of
children 0 to
< 6 years
7.8 kg =
weighted mean
body weight of
children 0 to
< 1 year
Incidental
ingestion volume
normalized to
incidental ingestion
per hour
0.12 L/hr = upper 97th percentile
calculated based on study that included children 6 to < 18 years3
0.07 L/hr (90th percentile ingestion volume)
0.12 L/hr (97th percentile ingestion volume)
calculated based on parent surveys for children 0 to <
15 yearsb
Recreational
exposure duration
(U.S. EPA 1997)
2.7 hr/d = mean
recreational exposure
duration for children
ages 5 to 11 years
1.4 hr/d = mean recreational exposure duration for children ages 1 to 4 years0
Ingestion rate
0.33 L/d
0.17 L/d
0.10 L/d (90th percentile ingestion volume)
0.17 L/d (97th percentile ingestion volume)
a Hourly ingestion rate for children is from EPA Exposure Factors Handbook (2011) Table 3-5: Ingestion of Water and Other Select Liquids fromDufour et al.
(2006). The Dufour et al. (2006) pilot study measured incidental ingestion of water of participants who spent time swimming or playing in a swimming pool for
at least a 45-minute period (n = 53; 41 children ages 6 to < 18 years; 12 adults > 18 years). This study did not include children younger than 6 years. U.S. EPA
(2011) reported an hourly ingestion rate, which EPA calculated by normalizing the Dufour et al. (2006) ingestion volume per 45 minutes to an ingestion volume
per hour (hourly ingestion rate) and also recommended using the 97th percentile as the "upper percentile" for children.
b Hourly ingestion rate for children is based on Schets et al. (2011) survey of Dutch parents' estimates of recreational duration and incidental ingestion volume
while recreating in surface water; 486 of the survey respondents reported their children recreated in fresh water. The incidental ingestion volume 90th and 97th
percentiles for children, 0.10 and 0.16 L/event, respectively, were calculated based on the distribution parameter reported in the paper. These volumes per event
were normalized to an hourly ingestion rate by dividing these values by the mean duration per recreational event reported for children by Schets et al. (2011),
which was 79 minutes or 1.3 hours.
0 Recreational exposure duration values reported in EPA's Exposure Factors Handbook (1997) are limited to children > 1 year old. Children aged 1 to 4 years are
the youngest life stage for which duration data are reported in Table 15-119 of U.S. EPA (1997).
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When comparing the values for the normalized mean hourly ingestion volume (Table
7-4), both studies estimated a similar incidental ingestion volume of 0.12 L/hr. This is notable
because Dufour et al. (2006) characterized the 6 to < 18-year age group and Schets et al.
characterized the 0 to < 15-year age group. The similar volumes between the cohorts could
indicate that the children younger than 6 years old were not contributing substantially to the
distribution of incidental ingestion volume. Likewise, the same could be said for the 15 to 18-
year age group. The results in Table 7-2 provide evidence that 6 to 10-year-old children
incidentally ingest significantly more than 11 to < 18-year-old children or adults. The 90th
percentile for incidental ingestion by 0 to < 15-year-old children reported by Schets et al. (2011)
is approximately 40 percent lower than the 97th percentile volume for 6 to < 18-year-old
children reported by Dufour et al. (2006).
EPA relied on the incidental ingestion volume recommended in the Exposure Factors
Handbook (2011), which discusses the use of the 97th percentile ingestion volume reported in
Dufour et al. (2006), because that study directly quantifies the water incidentally ingested while
recreating. EPA included the Schets study in this discussion because it provides valuable context
for characterizing children's incidental ingestion while recreating.
For children younger than 1 year, specific information is only available for body weights.
Combining this parameter with ingestion volumes and exposure duration times reported for older
age groups creates an exposure profile that would not seem to be representative of this early life
stage. This combination of factors is presented in Table 7-4 for comparison purposes only. The
available information is a better fit for children 1 to < 6 years.
Children 1 to 4 years are exposed for less time compared to children 5 to 11 years old,
1.4 hr/d compared to 2.7 hr/d, respectively (U.S. EPA 1997). Calculating the mean incidental
ingestion rate per day for children younger than 6 years old based on results from Dufour et al.
(2006) (0.17 L/d) or Schets et al. (2011) (0.10 L/d) results in lower estimated mean incidental
ingestion rates compared to children ages 6 to < 11 years (Table 7-4). However, these estimates
have large uncertainties given the lack of measured incidental ingestion data specifically for
children younger than 6 years. Information on exposure durations for children < 1-year-old is
also lacking. Because ingestion rates are greatest for 5 to 11 year olds, EPA concluded that
calculating the ingestion rate using a higher duration was protective of children younger than
6 years old as indicated in Table 7-4 (Dufour et al. 2006; Schets et al. 2011).
7.5 Other Recreational Exposures
This section compares primary and secondary contact exposures and discusses tribal
considerations for cyanotoxin and cyanobacterial cell exposure.
7.5.1 Other Recreational Exposure Pathways
EPA selected primary contact activities and incidental ingestion of water as the primary
exposure pathway for derivation of the recreational criteria and swimming advisories. Alternative
exposure parameter data could be considered in this approach as described in section 7.4. In this
section, EPA evaluated potential cyanotoxin exposures via inhalation of aerosols and dermal
contact. Inhalation and dermal toxicity data were not available; however, there are limited
available data to estimate inhalation and dermal exposure. EPA conducted analyses to estimate
inhalation and dermal exposure and compared those estimates to incidental ingestion of the
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cyanotoxins while recreating. Section 7.5.1.1 compares recreational ingestion and inhalation
exposures to microcystins. Similarly, section 7.5.1.2 compares recreational ingestion and dermal
exposure.
7.5.1.1 Inhalation of Cyanotoxins
Volatilization of microcystins and cylindrospermopsin from water to air is not expected
due to their size and charges. Both cyanotoxins are rather large compared to volatile chemicals.
Microcystins' acid groups are charged at the pH of normal surface waters. Cylindrospermopsin
features both negative and positive changes and like other zwitterions, do not volatilize
significantly into the air from water (Butler et al. 2012).
EPA did an analysis to determine if the criteria/swimming advisory values based on
incidental ingestion are protective of recreational inhalation exposures. Although the recreational
use is primary contact recreation, such as swimming, data are available for secondary contact
activities such as jet skiing or boating and white-capped wave, bubble-bursting action, which can
result in cyanotoxins becoming aerosols (microscopic liquid or solid particles suspended in air).
Cheng et al. (2007) collected via personal samples and found that volunteers recreating on a lake
with a 1 |ig/L concentration in water were exposed to air concentrations of microcystin-LR of
approximately 0.08 ng/m3 in their breathing zone.
Using the information from Cheng et al. (2007) and inhalation exposure parameters
provided in EPA's Exposure Factors Handbook (201 1), EPA compared the microcystin inhaled
dose (ng/d) to the ingested dose. The parameters and calculations for this analysis are presented
in Table 7-5. Using conservative assumptions for inhalation rates (i.e., moderate intensity and
95th percentile) and inhalation exposure duration (i.e., 5 hr/d) and comparing with mean
incidental ingestion rates, the estimated ingested dose is 151 times higher than the estimated
inhaled dose for children and 43 times higher than the estimated inhaled dose for adults.
This analysis supports the conclusion that inhalation exposure is negligible compared to
incidental ingestion while recreating. The inhalation toxicity is unknown for microcystin, but if it
is equal to ingestion toxicity, the values based on oral ingestion should be protective of
recreational inhalation exposures. EPA did not conduct a similar analysis for cylindrospermopsin
because published measured air concentration data for this cyanotoxin were not available.
The California Environmental Protection Agency (CalEPA) came to a similar conclusion
for water skiers (Butler et al. 2012). They cited Cheng et al. (2007) and noted that their results
showed that a liter of water contains 700,000 to 800,000 times the amount of cyanotoxins as in a
cubic meter of air. CalEPA calculated that this concentration is equivalent to 1.3 to 1.4 [xL
aerosolized microcystin/m3. Compared to the ingestion assumptions used for swimmers in the
calculation of their recreational guideline (i.e., 50 mL/hr), CalEPA calculated that a water-skier
would have to inhale at least 35,000 m3/hr while skiing to achieve a dose equal to the swimmer,
which is 17,000 times the inhalation rate of a marathon runner. CalEPA concluded that a water
skier would not inhale enough aerosol to receive a dose similar to what a swimmer gets from
ingestion.
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Table 7-5. Comparison of Recreational Exposure Ingested Dose to Inhaled Dose of Microcystin
Age
Group
Inhalation
Rate
(nrVmin)"
Inhalation
Rate per
Hour
(m3/hr)
| volume
per min x
60 min/hr]
Du ration of
Inhalation
Exposure
per Dav
(hr/d)
Daily
Inhalation
Rate
Adjusted for
Duration of
Exposure
(m3/d)
Concen-
tration in
Air
(ng/m3)b
Inhaled
Dose
(ng/day)
[daily rate
x conc. in
air]
Inhaled
Dose
(jig/day)
Ingestion
Rate (L/d)
Concen-
tration
in Water
(jj-g/L)
Ingestion
Dose
(|a.g/day)
|water conc.
x daily
ingestion
rate]
Ratio of
Ingested Dose
to Inhalation
Dose
[ingested
dose/inhaled
dose]
Assumed 95th percentile short-term inhalation exposure, moderate intensity activity level inhalation rate (U.S. EPA 2011), a 24-hour inhalation
exposure duration, and mean ingestion rate (U.S. EPA 2011), mean recreational exposure duration (U.S. EPA 1997), and water concentration of 1 ng/L
(Cheng et al. 2007)
Children
0.037
2.2
5.0
53
0.08
4.3
0.004
0.13
1
0.13
151
Adults
0.040
2.4
5.0
58
0.08
4.6
0.005
0.04
1
0.04
43
11 EPA's Exposure Factors Handbook (2011) did not report recommended short term, moderate intensity activity level inhalation rate values for children or adults
in aggregate; used highest inhalation rate listed for children and adult age groups for this conservative screen. For children, it was the age group 16 to < 21 years,
and for adults, it was 51 to < 61 years.
b Cheng et al. (2007) measured 0.08 ng/m3 in air near surface waters with a concentration of 1 |ag/L microcystins. Assuming a linear relationship of water
concentration to air concentration based on Cheng et al. (2007), the concentration in air at the recreational AWQC concentration for microcystins (i.e., 4 |.ig/L) is
calculated by multiplying 4 |.ig/L by the ratio of (0.08 ng/m3)/(l |ig/L). which equals 0.32 ng/m3.
Human Health Recreational Ambient Water Quality Criteria or 73
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Another comparison considers spray exposures from jet-ski and boat spray. Sinclair et al.
(2016) modeled a water-spray exposure scenario and observed much lower exposures than those
resulting from swimming or limited-contact recreational activities reported in the previous
study. Thus, EPA expects that the comparison above based on exposure from secondary contact
recreation is protective of primary contact recreation. Sinclair et al. (2016) also measured
urinary concentrations of cyanuric acid after 26 participants' exposure to spray in a simulated
10-minute car wash situation. Each subject wore a protective coverall with hood, vinyl gloves,
waterproof footwear, and safety glasses to ensure that only their face and mouths were exposed.
The estimated median and 90th percentile ingestion volumes were 0.18 and 1.89 mL,
respectively. Converted to a duration of 1 hour, the amounts would be 1.08 mL and 11.3 mL,
which are much lower than the incidental ingestion intakes per hour.
7.5.1.2 Dermal Absorption
EPA did not find any peer reviewed measured data for microcystin or
cylindrospermopsin dermal absorption. EPA's Dermal Exposure Assessment: A Summary of
EPA Approaches (U.S. EPA 2007) states that to get through the skin, a chemical must dissolve
into the stratum corneum, which is a stabilized lipid barrier; therefore, lipid solubility is required
initially (U.S. EPA 2007).
EPA used the dermal exposure equations in its Risk Assessment Guidance for Superfund
(U.S. EPA 2004) to estimate the potential absorbed dose of microcystins and compare it to the
incidentally ingested dose. Octanol-water partition coefficients required by these equations are
available for four microcystins, including microcystin-LR. Ward and Codd (1999) estimated the
log octanol-water partition coefficients of microcystin-LR, -LY, -LW and -LF using high
performance liquid chromatography (HPLC) as 2.16, 2.92, 3.46, and 3.56, respectively.
Cylindrospermopsin dermal absorption could not be predicted due to the lack of these
lipophilicity parameters.
The equation to estimate skin permeability coefficient from U.S. EPA (2004) is
Log Kp= -2.80 + 0.66 x log KOW-0.0056 x MW
Where:
Kp = Dermal permeability coefficient of compound in water (cm/hr)
Kow = Octanol-water partition coefficient (dimensionless)
MW = molecular weight (g/mole)
The equation to estimate dermal absorbed dose for highly ionized organic chemicals from
U.S. EPA (2004) is:
DA = Kp x Cw x t
Where:
DA = Absorbed dose per event (mg/cm2-event)
Kp = Dermal permeability coefficient of compound in water (cm/hr)
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Cw
t
The estimated microcystins absorbed dose based on these calculations and the exposure
parameters used for microcystins are presented in Table 7-6. Although this analysis is based on
very limited data, it supports the hypothesis that the dermal absorbed dose of microcystins is
likely to be negligible compared to incidentally ingested doses during recreational activities.
CalEPA also concluded dermal absorption of microcystins and cylindrospermopsin while
swimming is not expected to be significant due to the large size and charged nature of these
molecules (Butler et al. 2012). CalEPA eliminated the dermal absorption pathway from its risk
assessment of microcystins and cylindrospermopsin citing evidence that similarly large
molecules such as antibiotics have not been able to be formulated in a way to penetrate the skin
(Butler et al. 2012). A U.S. Army-contracted in vitro study by Kemppainen et al. (1990)
measured microcystin dermal penetration in 48 hours through excised human abdominal skin and
found 0.9 (±0.3) percent of the total dose in water penetrated through the skin; however, this
study has not been peer reviewed.
7.5.2 Tribal Considerations
EPA considered alternative exposure scenarios tribal communities might have, given
their cultural practices. Native American food foraging customs or cultural or religious
ceremonies can put them into primary or secondary contact with cyanotoxins. Primary contact
ceremonial use may include the use of a surface water body for religious or traditional purposes
by members of a tribe, involving immersion and intentional or incidental ingestion of water
(Eastman 2007).
It is uncertain whether these activities would lead to cyanotoxin exposures higher than
the primary recreational contact assumptions for incidental ingestion and exposure duration used
in this assessment.
7.6 Livestock and Pet Concerns
The world's first scientific report of adverse effects to animals from cyanobacteria was
written by George Francis, who described in 1878 the rapid death of stock animals at Lake
Alexandrina, a freshwater lake at the mouth of the Murray River in South Australia (Francis
1878). Since then, there have been numerous descriptions of mammal and bird mortalities
associated with exposure to cyanobacteria (Backer et al. 2015; Hilborn & Beasley 2015). The
literature throughout the 20th century includes reports from all inhabited continents (Stewart et al.
2008). However, the impacts of cyanotoxins on domestic and companion animals are likely
under-recognized because many cases are misdiagnosed, few cases are biochemically confirmed,
and even fewer are reported in the scientific literature or to animal health systems (Zaias et al.
2010).
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Table 7-6. Comparison of Recreational Exposure Ingested Dose to Dermal Absorbed Dose of Microcystins
Microcystin
LogKow
(Ward &
Codd
1999)
Molecular
Weight
Log Skin
Perm-
eability
Coefficient
(Log Kp)
Skin Perm-
eability
Coefficient
(KP)
(cm/hr)
Chemical
Cone, in
Water
(mg/cm3)
Assuming
Rcc AWQC
Level
Event
Duration"
(hr/cvcnt)
(mean for
5-to 11-
vear-olds)
Dermal
Absorbed
Dose per
Event
(mg/em2-
cvent)
Total Body
Surface Area
(cm2) (U.S.
EPA 2011)
95th percentile
Children 6 to <
11 Years
Dermal
Absorbed
Dose per
Event
(mg/cvcnt)
Dermal
Absorbed
Dose per
Event (mg/
event)
Ratio of
Ingested
Dose to
Dermal
Absorbed
Dose
Microcystin-LR
2.16
995.17
-6.95
1.1E-07
4.00E-06
2.7
1.2E-12
1.48E+04
2E-08
2E-05
71,824
Microcystin-LY
2.92
1002.16
-6.48
3.3E-07
4.00E-06
2.7
3.6E-12
1.48E+04
5E-08
5E-05
24,764
Microcystin-LW
3.46
1025.2
-6.26
5.5E-07
4.00E-06
2.7
6.1E-12
1.48E+04
9E-08
9E-05
14,670
Microcystin-LF
3.56
986.16
-5.97
1.1E-06
4.00E-06
2.7
1.2E-11
1.48E+04
2E-07
2E-04
7,618
11 Event duration is defined as 24-hour cumulative time spent at home in outdoor pool or spa as reported in EPA's Exposure Factors Handbook (U.S. EPA 1997).
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Livestock and pets can potentially be exposed to higher concentrations of, or have
increased exposure to, cyanotoxins than humans because they are known to consume
cyanobacterial scum and mats and drink cyanobacteria-contaminated water (Backer et al. 2013).
Dogs are additionally at risk, as they may lick cyanobacterial cells from their fur after swimming
in a water body with an ongoing bloom. Mats and scums can represent thousand-fold to million-
fold concentrations of cyanobacterial cell populations, and published microcystin concentrations
have ranged up to 24 mg microcystin/L from scum material (Chorus & Bartram 1999). Common
signs of HAB cyanotoxin poisonings in pets include repeated vomiting, diarrhea, loss of appetite,
abdominal swelling, stumbling, seizures, convulsions, disorientation, inactivity, or skin rashes
and hives (New York Sea Grant 2014; Trevino-Garrison et al. 2015). Although reports of
livestock deaths are relatively rare, in extreme cases death can occur minutes after drinking from
a contaminated water source. Acute symptoms of cyanotoxin poisoning can include loss of
appetite, weakness, staggering, or inflammation of the muzzle, ear, or udder. Higher levels of
cyanotoxins can lead to severe liver damage, the development of jaundice, and severe
photosensitization. Often livestock or pets that recover from these ailments can then suffer from
chronic failure to thrive (Australia Department of Economic Development Jobs Transport and
Resources 2013; Robinson & Alex 1987).
7.6.1 States and Animal HAB Guidelines
A few states have guideline levels specific to the protection of animals from cyanotoxin
poisoning (Appendix G). California has dog and cattle action levels for the cyanotoxins
microcystin, anatoxin-a, and cylindrospermopsin (Butler et al. 2012). For both dogs and cattle,
California estimated drinking water ingestion rates based on two publications by the National
Research Council, Nutrient Requirements for Beef Cattle and Nutrient Requirements for Dogs
and Cats. The animal specific RfD for each cyanotoxin was divided by the final water and
cyanobacterial biomass intake exposure levels, providing a cyanotoxin concentration that would
result in exposure at the RfD level or below. These calculations were performed for an acute
(lethal) and a subchronic scenario. Oregon has dog-specific guideline values for the cyanotoxins
anatoxin-a, cylindrospermopsin, microcystin, and saxitoxin based on the CalEPA method.
However the dog-specific guideline value for saxitoxins was modified by applying an
uncertainty factor for interspecies differences in sensitivity between humans (the species in the
critical study) and dogs (Oregon Health Authority 2016). Grayson County in Texas gives
information for domestic animals at current advisory levels for microcystin and
cylindrospermopsin. Advisories levels of 20 ppb for microcystin and cylindrospermopsin are
calculated as gallons of water that can be consumed for 10 and 80 pound dogs that will cause a
lethal or near-lethal dose. This does not include additional dose amounts that could be ingested
by a dog while self-grooming cyanobacteria scum off its fur (Lillis et al. 2012).
Other states mention animal poisoning in their guideline documents but do not give
guideline values specific to livestock or companion animals. For example, Utah and Washington
report that animal illness or death can be reason to issue or accelerate a HAB advisory warning
(Hardy & Washington State Department of Health 2008; Utah Department of Environmental
Quality and Department of Health 2015). However, Ohio issues the disclaimer that thresholds
used are protective of human exposure and may or may not be protective of animals such as dogs
or livestock (Kasich et al. 2015). Several other states including Connecticut, Idaho, Kansas,
Massachusetts, Nebraska, Vermont, and Virginia provide informational pamphlets, warn about
Human Health Recreational Ambient Water Quality Criteria or
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harm to pets or other animals, or post about harm to animals in their beach warnings and
advisory signage (Connecticut Department of Public Health: Connecticut Energy Environment
2013; IDEQ 2015; Kansas Department of Health and Environment 2016; Massachusetts Bureau
of Environmental Health 2015; Nebraska Department of Environmental Quality and Nebraska
Department of Health and Human Services: Division of Public Health 2016; Vermont
Department of Health 2015; Virginia Department of Health 2012).
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APPENDIX A. INTERNATIONAL RECREATIONAL WATER GUIDELINES FOR CYANOTOXINS AND
CYANOBACTERIA
Jurisdiction
Recreational Water Guideline Level
Recommended Action
Australia"
cyanobacteria (total): >10 mm3/L
(where known toxins are not present)
red level action mode; level 2 guideline:
• Immediately notify health authorities for advice on health risk.
• Make toxicity assessment or toxin measurement of water if this has not already been
done.
• Health authorities warn of risk to public health (i.e., the authorities make a health risk
assessment considering toxin monitoring data, sample type and variability).
cyanobacteria (total): > 4 mm3/L (where
a known toxin producer is dominant in
the total biovolume)
red level action mode; level 1 guideline:
• Immediately notify health authorities for advice on health risk.
• Make toxicity assessment or toxin measurement of water if this has not already been
done.
• Health authorities warn of risk to public health (i.e., the authorities make a health risk
assessment considering toxin monitoring data, sample type and variability).
cyanobacteria (total): > 0.4 to < 10
mm3/L (where known toxin producers
are not present)
amber level alert mode:
• Increase sampling frequency to twice weekly where toxigenic species are dominant
within the alert level definition (i.e., total biovolume).
• Monitor weekly or fortnightly where other types are dominant.
• Make regular visual inspections of water surface for scums.
• Decide on requirement for toxicity assessment or toxin monitoring.
cyanobacteria (total): > 0.4 to <4 mm3/L
(where a known toxin producer is
dominant in the total biovolume)
amber level alert mode:
• Increase sampling frequency to twice weekly where toxigenic species are dominant
within the alert level definition (i.e., total biovolume).
• Monitor weekly or fortnightly where other types are dominant.
• Make regular visual inspections of water surface for scums.
• Decide on requirement for toxicity assessment or toxin monitoring.
cyanobacteria (total): > 0.04 to <0.4
mm3/L
green level surveillance mode:
• Weekly sampling and cell counts at representative locations in the water body where
known toxigenic species are present; or
• Fortnightly for other types including regular visual inspection of water surface for scums.
cyanobacterial scums consistently
present
red level action mode; level 2 guideline:
• Immediately notify health authorities for advice on health risk.
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Jurisdiction
Recreational Water Guideline Level
Recommended Action
• Make toxicity assessment or toxin measurement of water if this has not already been
done.
• Health authorities warn of risk to public health (i.e., the authorities make a health risk
assessment considering toxin monitoring data, sample type and variability).
microcystins (total): >10 |ig/L
red level action mode; level 1 guideline:
• Immediately notify health authorities for advice on health risk.
• Make toxicity assessment or toxin measurement of water if this has not already been
done.
• Health authorities warn of risk to public health (i.e., the authorities make a health risk
assessment considering toxin monitoring data, sample type and variability).
Microcystis aeruginosa (total): > 50,000
cells/mL
red level action mode; level 1 guideline:
• Immediately notify health authorities for advice on health risk.
• Make toxicity assessment or toxin measurement of water if this has not already been
done.
• Health authorities warn of risk to public health (i.e., the authorities make a health risk
assessment considering toxin monitoring data, sample type and variability).
Microcystis aeruginosa (total): > 5,000
to < 50,000 cells/mL
amber level alert mode:
• Increase sampling frequency to twice weekly where toxigenic species are dominant
within the alert level definition (i.e., total biovolume).
• Monitor weekly or fortnightly where other types are dominant.
• Make regular visual inspections of water surface for scums.
• Decide on requirement for toxicity assessment or toxin monitoring
Microcystis aeruginosa (total): > 500 to
< 5,000 cells/mL
green level surveillance mode:
• Weekly sampling and cell counts at representative locations in the water body where
known toxigenic species are present; or
• Fortnightly for other types including regular visual inspection of water surface for scums.
Canadad
cyanobacteria (total): > 100,000
cells/mL
issue swimming advisory
detection of a cyanobacterial bloom
issue beach closure
microcystins (total): > 20 |ig/L
(expressed as microcystin-LR)
issue swimming advisory
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Jurisdiction
Recreational Water Guideline Level
Recommended Action
Cuba0
any report of toxic effect in humans or
animals
action (in red): as for "Alert", but with increased actions for public communication
benthic mats: < 40 percent coverage of
surfaces with any cyanobacteria; > 20
percent with toxicogenic cyanobacteria;
> 50 percent with potentially toxicogenic
cyanobacteria (particularly where they
are visibly detaching and accumulating
in scum)
alert: increased sampling (weekly and more sites); daily inspection; notification to public
health unit and local managers; report to local government; warning of the public
cyanobacteria: < 500 cells/mL
monthly visual inspection
cyanobacteria: > 1 of the species known
as potentially toxic
alert: increased sampling (weekly and more sites); daily inspection; notification to public
health unit and local managers; report to local government; warning of the public
phytoplankton cells: >_20.000 to <
100,000 cells/mL, > 50 percent of cells
cyanobacteria
alert: increased sampling (weekly and more sites); daily inspection; notification to public
health unit and local managers; report to local government; warning of the public
phytoplankton: > 0 to < 1,500 cells/mL
monthly visual inspection and sampling at least four months per year
scum consistently present; confirmed
bloom persistence
action (in red): as for "Alert", but with increased actions for public communication
Czech Republic0
cells: > 100,000 cells/mL
2nd warning level: closure for public recreation
cells: > 20,000 cells/mL
1st warning level (not otherwise specified)
Denmark0
chlorophyll a: > 50 |.ig/L. dominated by
cyanobacteria
relevant authorities are informed and decide when and how the public should be informed;
warnings include signs, media and contact to local user groups such as kindergardens,
scouts, water sports clubs
visible surface scum
relevant authorities are informed and decide when and how the public should be informed;
warnings include signs, media and contact to local user groups such as kindergardens,
scouts, water sports clubs
European Unionf
cyanobacterial proliferation (occurrence)
when cyanobacterial proliferation occurs and a health risk has been identified or presumed,
adequate management measures shall be taken immediately to prevent exposure, including
information to the public
cyanobacterial proliferation (potential
for)
appropriate monitoring shall be carried out to enable timely identification of health risks.
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Jurisdiction
Recreational Water Guideline Level
Recommended Action
Finland0
algae (includes cyanobacteria): detected
level 1: Possibly microscopic examination and even toxin analysis if there is a specific
cause such as very popular beach or reports of adverse health effects or animal deaths
algae (includes cyanobacteria): high
amount
level 2: Preferably microscopical examination; toxin analysis; warning of the public is
compulsory
algae (includes cyanobacteria): very high
amount
level 3: Preferably microscopical examination; toxin analysis; warning of the public is
compulsory
France0
bloom, scum, change in water color
microscopy examination. If cyanobacteria are absent: no further action. If present: counting
and genus identification
cyanobacteria: < 20,000 cells/mL (±20
percent)
active daily monitoring. Counting at least on a weekly basis. Normal recreational activity at
the site
cyanobacteria: > 100,000 cells/mL (±20
percent)
bathing and recreational activities are restricted. Public is informed.
cyanobacteria: >_20.000 to < 100,000
cells/mL (±20 percent)
active daily monitoring. Counting on a weekly basis. Recreationalactivities are still
allowed; the public is informed by posters on site.
microcystins (MC): 25 |ig/L (±5 percent)
if MC < 25 (ig/L bathing and recreational activities are restricted. If MC > 25 |ig/L bathing
is banned and recreational activities are restricted. In either case, public is informed.
visible scum or foam in recreational or
bathing area
all water activities in this area are prohibited. Restrictions do not necessarily apply to the
whole recreational site. Other areas without scum may still be open.
Germany0
Secchi Disk reading > 1 m AND
biovolume: < 1 mm3/L
monitor further cyanobacterial development
Secchi Disk reading > 1 m AND
biovolume: > 1 mm3/L
publish warnings, discourage bathing, consider temporary closure
Secchi Disk reading > 1 m AND
chlorophyll a (with dominance by
cyanobacteria): < 40 (ig/L
monitor further cyanobacterial development
Secchi Disk reading > 1 m AND
chlorophyll a (with dominance by
cyanobacteria): > 40 |ig/L
publish warnings, discourage bathing, consider temporary closure
Secchi Disk reading > 1 m AND
microcystins: <10 |ig/L
monitor further cyanobacterial development
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Jurisdiction
Recreational Water Guideline Level
Recommended Action
Secchi Disk reading > 1 m AND
microcystins: >10 |ig/L
publish warnings, discourage bathing, consider temporary closure
visible heavy scums and/or microcystins:
> 100 (ig/L
publish warnings, discourage bathing, temporary closure is recommended
Hungary0
cell count: > 50,000 to < 100,000
cells/mL
no recommended actions listed, water body classification: Acceptable
cell count: < 20,000 cells/mL
no recommended actions listed, water body classification: Excellent
cell count: > 20,000 to < 50,000
cells/mL
no recommended actions listed, water body classification: Good
cell count: > 100,000 cells/mL
no recommended actions listed, water body classification: Unacceptable
chlorophyll a (with dominance by
cyanobacteria): <10 (ig/L
no recommended actions listed, water body classification: Excellent
chlorophyll a (with dominance by
cyanobacteria): > 10 to < 25 |ig/L
no recommended actions listed, water body classification: Good
chlorophyll a (with dominance by
cyanobacteria): > 25 to < 50 |ig/L
no recommended actions listed, water body classification: Acceptable
chlorophyll a (with dominance by
cyanobacteria): > 50 (ig/L
no recommended actions listed, water body classification: Unacceptable
microcystins: > 4 to < 10 |ig/L
no recommended actions listed, water body classification: Good
microcystins: > 10 to < 20 (ig/L
no recommended actions listed, water body classification: Acceptable
microcystins: < 4 (ig/L
no recommended actions listed, water body classification: Excellent
microcystins: > 20 (ig/L
no recommended actions listed, water body classification: Unacceptable
Italy0
cyanobacterial cell count (combined
with identification of genus and, if
possible, species): < 20,000 cells/mL
if possible, daily visual observation; weekly counting
cyanobacterial cell count (combined
with identification of genus and, if
possible, species): > 100,000 cells/mL
bathing prohibited until quantification of microcystins; information to the public; at least
weekly counting
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Jurisdiction
Recreational Water Guideline Level
Recommended Action
cyanobacterial cell count (combined
with identification of genus and, if
possible, species): >_20.000 to < 100,000
cells/mL
daily visual observation; at least weekly counting; information to the public; quantification
of microcystins
microcystins: >25 |ig/L
bathing prohibited
visible scums
bathing prohibited until quantification of microcystins; warning notice; scum drift
monitoring
Netherlands0
biovolume (cyanobacterial cell count):
>0 to < 2.5 mm3/L
surveillance level: continue fortnightly monitoring
biovolume (cyanobacterial cell count): >
15 mm3/L (if 80 percent dominance of
microcystin-producers and microcystin <
20 |ig/L, revert to Alert Level 1).
alert level 2: weekly monitoring and advice against bathing (by public authority): "You are
advised not to bathe in this water;" prohibition by local authority is possible.
biovolume (cyanobacterial cell count): >
2.5 to < 15 mm3/L
alert level 1: weekly monitoring and issue warning (by site operator) for duration of that
week: "Toxic blue-green algae. Risk of skin irritation or intestinal problems." In case of
daily site inspection, reevaluate the warning on a daily basis.
chlorophyll a: > 0 to < 12.5 (ig/L
surveillance level: continue fortnightly monitoring
chlorophyll a: >15 |ig/L
alert level 2: weekly monitoring and advice against bathing (by public authority): "You are
advised not to bathe in this water;" prohibition by local authority is possible.
chlorophyll a: ^12.5 to < 75 (ig/L
alert level 1: weekly monitoring and issue warning (by site operator) for duration of that
week: "Toxic blue-green algae. Risk of skin irritation or intestinal problems." In case of
daily site inspection, reevaluate the warning on a daily basis.
surface scum: category 1
surveillance level: continue fortnightly monitoring
surface scum: category 2
alert level 1: weekly monitoring and issue warning (by site operator) for duration of that
week: "Toxic blue-green algae. Risk of skin irritation or intestinal problems". In case of
daily site inspection, reevaluate the warning on a daily basis.
surface scum: category 3
alert level 2: weekly monitoring and advice against bathing (by public authority): "You are
advised not to bathe in this water"; prohibition by local authority is possible.
New Zealand11
cyanobacteria (benthic): 20-50 percent
coverage of potentially toxigenic
cyanobacteria attached to substrate
alert (amber mode):
• Notify the public health unit.
• Increase sampling to weekly.
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Jurisdiction
Recreational Water Guideline Level
Recommended Action
• Recommend erecting an information sign.
• Consider increasing the number of survey sites.
• If toxigenic cyanobacteria dominate the samples, testing for cyanotoxins is advised. If
cyanotoxins are detected in mats or water samples, consult the testing laboratory to
determine if levels are hazardous.
cyanobacteria (benthic): greater than
50 percent coverage of potentially
toxigenic cyanobacteria attached to
substrate
action (red mode) situation 1:
• Immediately notify the public health unit
• If potentially toxic taxa are present (see Table 2) then consider testing samples for
cyanotoxins
• Notify the public of the potential risk to health
cyanobacteria (benthic): Up to 20
percent coverage of potentially toxigenic
cyanobacteria attached to substrate
surveillance (green mode):
• Undertake fortnightly surveys between spring and autumn at representative locations in
the water body where known mat proliferations occur and where there is recreational use
cyanobacteria (benthic): up to 50 percent
where potentially toxigenic
cyanobacteria are visibly detaching from
the substrate, accumulating as scums
along the river's edge or becoming
exposed on the river's edge as the river
level drops.
action (red mode) situation 2:
• Immediately notify the public health unit
• If potentially toxic taxa are present (see Table 2) then consider testing samples for
cyanotoxins.
• Notify the public of the potential risk to health
cyanobacteria (total): <0.5 mm3/L
(biovolume equivalent of the combined
total of all cyanobacteria)
surveillance (green mode):
• Undertake weekly or fortnightly visual inspection and sampling of water bodies where
cyanobacteria are known to proliferate between spring and autumn
cyanobacteria (total): < 500 cells/mL
surveillance (green mode):
• Undertake weekly or fortnightly visual inspection and sampling of water bodies where
cyanobacteria are known to proliferate between spring and autumn
cyanobacteria (total): >1.8 mm3/L
(biovolume equivalent of potentially
toxic cyanobacteria)
action (red mode) situation 1:
• Continue monitoring as for alert (amber mode)
• If potentially toxic taxa are present (see Table 1), then consider testing samples for
cyanotoxins
• Notify the public of a potential risk to health
cyanobacteria (total): >_0.5 to < 1.8
mm3/L (biovolume equivalent of
potentially toxic cyanobacteria)
alert (amber mode):
• Increase sampling frequency to at least weekly
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Jurisdiction
Recreational Water Guideline Level
Recommended Action
• Notify the public health unit
• Multiple sites should be inspected and sampled
cyanobacteria (total): > 0.5 to < 10
mm3/L (total biovolume of all
cyanobacterial material where the
cyanobacterial population has been
tested and shown not to contain known
toxins)
alert (amber mode):
• Increase sampling frequency to at least weekly.
• Notify the public health unit.
• Multiple sites should be inspected and sampled.
cyanobacteria (total): >_10 mm3/L (total
biovolume of all cyanobacterial material
where the cyanobacterial population has
been tested and shown not to contain
known toxins)
action (red mode) situation 2:
• Continue monitoring as for alert (amber mode)
• If potentially toxic taxa are present (see Table 1), then consider testing samples
for cyanotoxins
• Notify the public of a potential risk to health
cyanobacterial scums consistently
present for more than several days in a
row
action (red mode) situation 3:
• Continue monitoring as for alert (amber mode)
• If potentially toxic taxa are present (see Table 1), then consider testing samples for
cyanotoxins
• Notify the public of a potential risk to health
microcystins (total): >12 |ig/L
action (red mode) situation 1:
• Continue monitoring as for alert (amber mode)
• If potentially toxic taxa are present (see Table 1), then consider testing samples for
cyanotoxins
• Notify the public of a potential risk to health
Poland0
visible blooms
sampling of bathing sites not less than 4 times per season (the interval between sampling
does not exceed one month), including responses to cyanobacteria if blooms are observed.
Scotland®
chlorophyll a: > 10 |ig/L with
dominance of cyanobacteria
1. watch for scum or conditions conducive to scums.
2. discourage bathing and further investigate hazard.
3. post on-site risk advisory signs.
4. inform relevant authorities.
cyanobacteria: > 20,000 cells /mL
1. watch for scum or conditions conducive to scums.
2. discourage bathing and further investigate hazard.
3. post on-site risk advisory signs.
4. inform relevant authorities.
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Jurisdiction
Recreational Water Guideline Level
Recommended Action
cyanobacterial scum
formation in bathing areas
1. immediate action to control contact with scums; possible prohibition of swimming and
other water-contact activities.
2. public health follow-up investigation.
3. inform public and relevant authorities.
Singapore0
chlorophyll a: < 50 |ig/L (of 95 percent
of a 3-year rolling period)
status of the sites reviewed annually. If the assessment is that the water body is unsuitable
for primary water contact activities, the public is notified.
Spain0
cyanobacteria proliferation potential
(High, Medium, Low)
criteria for assessment of health risk and response are set locally; some health authorities
use WHO scheme, others include further risk parameters (such as number of users, type of
use); temporary closure has occasionally occurred based on the abundance of
cyanobacteria.
Turkey0
cells: < 20,000 cells/mL
level 1: recreational activities are allowed to continue and users are informed by posters on
site. Monitoring (sampling, counting and species identification) should be done fortnightly.
cells: 20,000-100,000 cells/mL
level 2: At > 20 000 cells/mL, microcystins are analyzed. If microcystin-LR equivalents
>25 |ig/L, immediate action to inform relevant authorities and public. Discourage users
from swimming and other water-contact activities by advisory signs on site.
chlorophyll a (if dominated by
cyanobacteria): <10 |ig/L
level 1: recreational activities are allowed to continue and users are informed by posters on
site. Monitoring (sampling, counting and species identification) should be done fortnightly.
microcystin-LR: < 10 |ig/L equivalents
level 1: recreational activities are allowed to continue and users are informed by posters on
site. Monitoring (sampling, counting and species identification) should be done fortnightly.
microcystin-LR: > 25 |ig/L equivalents
level 2: At > 20,000 cells/mL, microcystins are analyzed. If microcystin-LR equivalents
>25 |ig/L. immediate action to inform relevant authorities and public. Discourage users
from swimming and other water-contact activities by advisory signs on site.
visible scum in bathing area
level 3: all activities in the water may be prohibited
World Health
Organization (WHO)b g
chlorophyll a: 10 |ig/L with dominance
of cyanobacteria
low risk: post on-site advisory signs, inform relevant authorities
chlorophyll a: 50 (ig/L with dominance
of cyanobacteria
moderate risk: watch for scums or conditions conducive to scums, discourages swimming
and further investigate hazard, post on-site risk advisory signs, inform relevant authorities
cyanobacteria: 100,000 cells/mL
moderate risk: watch for scums or conditions conducive to scums, discourages swimming
and further investigate hazard, post on-site risk advisory signs, inform relevant authorities
cyanobacteria: 20,000 cells/mL
low risk: post on-site advisory signs, inform relevant authorities
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
A-9
-------
Jurisdiction
Recreational Water Guideline Level
Recommended Action
cyanobacterial scum formation in areas
where whole-body contact and/or risk of
ingestions/aspiration occur
high risk: immediate action to control contact with scums, possible prohibition of
swimming and other water contact activities, public health follow-up investigation, inform
public and relevant authorities
a Australian Government National Health and Medical Research Council (2008). Guidelines for Managing Risk in Recreational Water.
b Chorus, I. and Bartram, J. (eds.) (1999). Toxic cyanobacteria in water: A guide to public health significance, monitoring and management. E. & F.N. Spon /
Chapman & Hall, London, United Kingdom.
c Federal Environment Agency (Germany) (2012). Current approaches to Cyanotoxin risk assessment, risk management and regulations in different countries.
d Health Canada (2012). Guidelines for Canadian Recreational Water Quality, Third Edition. Water, Air and Climate Change Bureau, Healthy Environments and
Consumer Safety Branch, Health Canada, Ottawa, Ontario. (Catalogue No H129-15/2012E).
e Scottish Government Health and Social Care Directorates Blue-Green Algae Working Group (2012). Cyanobacteria (Blue-Green Algae) in Inland and Inshore
Waters: Assessment and Minimization of Risks to Public Health.
'European Parliament and the Council of the European Union (2006). Directive 2006/7/EC of the European Parliament and of the Council of 15 February 2006
concerning the management of bathing water quality and repealing Directive 76/160/EEC.
g WHO (World Health Organization) (2003). Guidelines for Safe Recreational Water Environments: Volume 1: Coastal and Fresh Waters. World Health
Organization.
h Wood S; Hamilton, D; Sail K; Williamson, W. (2008). New Zealand Guidelines for Cyanobacteria in Recreational Fresh Waters: Interim Guidelines. New
Zealand Ministry for the Environment and Ministry of Health.
Human Health Recreational Ambient Water Quality Criteria or A-10
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
-------
APPENDIX B. STATE RECREATIONAL WATER GUIDELINES FOR CYANOTOXINS
AND CYANOBACTERIA
EPA compiled the information presented in this appendix based on searches of state
websites for publicly available information regarding guidelines or action levels for cyanotoxins
and cyanobacteria. The website research was completed in November 2015. Subsequent direct
personal communication of state guidelines revealed some updates for a few states later in 2015
and in early 2016.
Table B-l. Summary Counts of State Recreational Water Guidelines for Cyanotoxins and
Cyanobacteria by Type and Scope of Guidelines
Recreational Water
Guideline Tvpe
and Scope
Number of States and
List of States
Additional Information
Quantitative guidelines for
cyanobacteria only
6 states
Arizona, Connecticut, Idaho,
Maine, New Hampshire,
Wisconsin
Measurements for these criteria include
cyanobacterial cell densities, proportion of
toxigenic cyanobacteria, chlorophyll concentration,
and Secchi disk depth measurements.
Quantitative guidelines for
cyanotoxins only
7 states
California, Colorado, Illinois,
Iowa, Nebraska, North Dakota,
Ohio
State guidelines address four cyanotoxins in order
from most to least common:
microcystins (20 states)
anatoxin-a (9 states)
cylindrospermopsin (7 states)
saxitoxin (4 states)
Quantitative guidelines for
cyanotoxins and either
quantitative or qualitative
guidelines for
cyanobacteria
14 states
Indiana, Kansas, Kentucky,
Maryland, Massachusetts, New
York, Oklahoma, Oregon, Rhode
Island, Texas, Utah, Vermont,
Virginia, Washington
Qualitative guidelines only
6 states
Delaware, Florida, Montana,
North Carolina, North Dakota,
West Virginia
Examples include:
presence of surface scum
visible discoloration
presence of potentially toxic algae
Note: EPA found that Texas and North Carolina published guidelines in the past, but the guidelines are no longer
found on their websites.
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
B-l
-------
Table B-2. Summary Counts of State Recreational Water Guidelines for Cyanotoxins and
Cyanobacteria by Basis of Guidelines
Recreational Water Guideline
Basis of Guideline Category
Number of States and List of States
Based on WHO
6 states
Colorado, Indiana, Kentucky, Oklahoma, Utah, Wisconsin
Modified WHO
8 states
Illinois, Indiana, Iowa, Kentucky, Oklahoma, Oregon, Utah, Virginia
Jurisdiction-specific (i.e., based on
risk assessments or site-specific
monitoring information)
8 states
California, Colorado, Kansas, Massachusetts, Ohio, Oregon, Vermont,
Washington
Based on studies or guidelines other
than WHO
3 states
Idaho, Indiana, Utah
Qualitative evaluations or narrative
criteria application (includes states
with insufficient documentation to
categorize the source of their
guideline)
21 states
Arizona, Connecticut, Delaware, Florida, Maine, Maryland, Massachusetts,
Montana, Nebraska, New Hampshire, New York, North Carolina, North
Dakota, Oregon, Rhode Island, Texas, Utah, Virginia, Washington, West
Virginia, Wisconsin
Note: Some states are listed in more than one category because they had more than one guideline (e.g., both
cyanobacterial cell and cyanotoxin guidelines), and these guidelines fit into different categories.
Human Health Recreational Ambient Water Quality Criteria or B-2
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
-------
Table B-3. State Recreational Water Quality Guideline for Cyanotoxins and Cyanobacteria Sorted by Type
State
Recreational Water Guideline Level
Recommended Action
Reference
States with Guidelines Based on Cyanobacteria Only
Arizona
blue-green algae (mean value based on a
minimum of two sample events within one
peak season): 20,000 cells/mL
and
chlorophyll a result (mean value based on a
minimum of two sample events within one
peak season) in target range
violation of the Narrative Nutrient Standard
Arizona Department of Environmental
Quality (2008). Narrative Nutrient
Standard Implementation Procedures
for Lakes and Reservoirs.
httD://www.azdea.eov/environ/water/s
tandards/download/draft nutrient.txtf.
Last Accessed: 08/03/2016.
Connecticut
visual rank category 1: visible material is not
likely cyanobacteria or water is generally
clear
no action
Connecticut Department of Public
Health: Connecticut Energy
Environment (2013). Guidance to
Local Health Departments For Blue-
Green Algae Blooms in Recreational
Freshwaters.
httD://www.ct.eov/deeD/lib/deeD/water
/water tiualifv standards/guidance Hi
m l>ga bloonx -n!3.ixif. Last
visual rank category 2: cyanobacteria present
in low numbers; there are visible small
accumulations but water is generally clear;
OR blue-green algae cells > 20,000 cells/mL
and < 100,000 cells/mL
notify Connecticut Department of Public Health (CT
DPH), Connecticut Department of Energy and
Environmental Protection (CT DEEP)
visual rank category 3: cyanobacteria present
in high numbers; scums may or may not be
present; water is discolored throughout; large
areas affected; color assists to rule out
sediment and other algae; OR blue-green
algae cells > 100,000 cells/mL
update/inform CT DPH & CT DEEP and expand risk
communication efforts; POSTED BEACH
CLOSURE: if public has beach access, alert water
users that a blue-green algae bloom is present;
POSTED ADVISORY: at other impacted access
points
Accessed: 08/03/2016.
Idaho
Microcystis or Planktothrix: >40,000 cells/mL
public health advisory posting by Public Health
District in conjunction with water body operator
IDEQ (Idaho Department of
Environmental Quality) (2015). Blue-
Green Algae Bloom Response Plan:
Final.
httD://www.eDa.illinois.gov/toDics/wat
sum of all potentially toxigenic taxa: >
100,000 cells/mL
public health advisory posting by Public Health
District in conjunction with water body operator
visible surface scum that is associated with
toxigenic species
public health advisory posting by Public Health
District in conjunction with water body management
agency
er-qual i tv/mo nitori ng/alga 1-
. Last Accessed: 08/03/2016.
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
B-3
-------
State
Recreational Water Guideline Level
Recommended Action
Reference
Maine
Secchi disk reading < 2 meters caused by
algae
body of water considered impaired, but still safe to
swim
Maine Department of Environmental
Protection (2013). Reports of Algal
Blooms.
lit! d ://www. ma i ne. eov/deD/wate r/lakes
/reob loo m. lit nil. Last Accessed:
08/03/2016.
New Hampshire
cyanobacteria: > 50 percent of total cell
counts from toxigenic cyanobacteria
post beach advisory
New Hampshire Department of
Environmental Services (2014). Beach
Advisories.
httD://des. nh.gov/organization/division
s/water/wmb/beaches/advisories.htm.
Last Accessed: 08/03/2016.
Wisconsin
cyanobacteria: > 100,000 cells/mL
post health advisory and possible beach closure
Wisconsin Department of Natural
Resources (2012). Draft Blue-Green
Algae Section of 303 (d) Report-
7/3/2012: Harmful Algal Blooms.
httD://dnr. wi.gov/lakes/bluegreenalgae
/documents/Harmful AlgalBloomsvs2.
pdf. Last Accessed: 08/03/2016.
visible scum layer
post health advisory and possible beach closure
Werner M, & Masnado R (2014).
Guidance for Local Health
Departments: Cyanobacteria and
Human Health.
lit! d ://c i tv. mi lwaukee. gov/I ma geLib ra r
v/G roiros/liealtliAutlio rs/DCP/PDF s/C
vanobacteriaLHD.pdf. Last Accessed:
08/03/2016.
States with Guidelines Based on Cyanotoxin(s) Only
California
anatoxin-a: 90 |ig/L
Unclear
Butler N, Carlisle J, Kaley KB, &
Linville R (2012). Toxicological
Summary and Suggested Action
Levels to Reduce Potential Adverse
Health Effects of Six Cyanotoxins.
cylindrospermopsin: 4 |ig/L
Unclear
microcystins: 0.8 |ig/L
Unclear
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
B-4
-------
State
Recreational Water Guideline Level
Recommended Action
Rct'ercncc
i ssues/pro gra ms/oeer review/docs/ca li
f cvaix)toxins/cvaiiotoxins()53112.pdf.
Last Accessed: 08/03/2016.
Colorado
anatoxin-a: > 7 |ig/L
issue toxic algae caution:
a. post sign with "caution" language.
b. perform routine testing for toxin levels.
bi. if test results are below caution thresholds, test
at least once per week until algae visually subsides,
bii. if test results are above caution thresholds, test
at least twice per week until toxin levels are below
caution thresholds for two consecutive tests.
c. notify drinking water providers and county
health department if toxin levels exceed the caution
thresholds.
d. toxic algae caution ends when there is no visual
evidence of algae and toxin levels are non-
detectable for two consecutive weeks.
di. notify drinking water providers and county
health department that bloom has ended,
dii. remove "caution" sign.
Colorado Department of Public Health
& Environment. Algae bloom risk-
management toolkit for recreational
waters.
httDs://drive.gooele.com/file/d/OBOtm
P067k3 N VN 2U 4 VHZB c W xPN OE/vi
ew. Last Accessed: 10/21/2016
cylindrospermopsin: > 7 |ig/L
issue toxic algae caution:
a. post sign with "caution" language.
b. perform routine testing for toxin levels.
bi. if test results are below caution thresholds, test
at least once per week until algae visually subsides,
bii. if test results are above caution thresholds, test
at least twice per week until toxin levels are below
caution thresholds for two consecutive tests.
c. notify drinking water providers and county
health department if toxin levels exceed the caution
thresholds.
d. toxic algae caution ends when there is no visual
evidence of algae and toxin levels are non-
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
B-5
-------
State
Recreational Water Guideline Level
Recommended Action
Reference
detectable for two consecutive weeks,
di. notify drinking water providers and county
health department that bloom has ended,
dii. remove "caution" sign.
microcystin-LR: > 10 |ig/L and < 20 j.ig/L
issue toxic algae caution:
a. post sign with "caution" language.
b. perform routine testing for toxin levels.
bi. if test results are below caution thresholds, test
at least once per week until algae visually subsides,
bii. if test results are above caution thresholds, test
at least twice per week until toxin levels are below
caution thresholds for two consecutive tests.
c. notify drinking water providers and county
health department if toxin levels exceed the caution
thresholds.
d. toxic algae caution ends when there is no visual
evidence of algae and toxin levels are non-
detectable for two consecutive weeks.
di. notify drinking water providers and county
health department that bloom has ended,
dii. remove "caution" sign.
microcystin-LR: > 20 j.ig/L
issue toxic algae warning:
a. immediately post sign with "warning" language.
b. take necessary steps to prevent contact with
water in affected area for humans and pets.
c. notify drinking water providers and county
health department if toxin levels exceed warning
thresholds.
d. test at least twice per week until toxin levels are
below warning thresholds for two consecutive
tests.
e. posting can be reduced to "caution" language
when microcystin test results drop below the
warning threshold and no new human illness or pet
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
B-6
-------
State
Recreational Water Guideline Level
Recommended Action
Reference
deaths have been reported for two consecutive
weeks.
saxitoxin: > 4 |ig/L
issue toxic algae caution:
a. post sign with "caution" language.
b. perform routine testing for toxin levels.
bi. if test results are below caution thresholds, test
at least once per week until algae visually subsides,
bii. if test results are above caution thresholds, test
at least twice per week until toxin levels are below
caution thresholds for two consecutive tests.
c. notify drinking water providers and county
health department if toxin levels exceed the caution
thresholds.
d. toxic algae caution ends when there is no visual
evidence of algae and toxin levels are non-
detectable for two consecutive weeks.
di. notify drinking water providers and county
health department that bloom has ended,
dii. remove "caution" sign.
potentially toxic algae are visible
issue toxic algae caution:
a. post sign with "caution" language.
b. perform routine testing for toxin levels.
bi. if test results are below caution thresholds, test
at least once per week until algae visually subsides,
bii. if test results are above caution thresholds, test
at least twice per week until toxin levels are below
caution thresholds for two consecutive tests.
c. notify drinking water providers and county
health department if toxin levels exceed the caution
thresholds.
d. toxic algae caution ends when there is no visual
evidence of algae and toxin levels are non-
detectable for two consecutive weeks.
di. notify drinking water providers and county
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
B-7
-------
State
Recreational Water Guideline Level
Recommended Action
Reference
health department that bloom has ended,
dii. remove "caution" sign.
Illinois
microcystin-LR: > 10 |ig/L
appropriate lake management personnel and Illinois
EPA staff will be notified; follow-up monitoring by
the Illinois EPA may occur as professional judgment
dictates and staff laboratory, and financial resources
allow
Illinois Environmental Protection
Agency (2015). 2013 Statewide
Harmful Algal Bloom Program,
http ://epa. illinois.gov/topics/water-
quality/monitoring/algal-bloom/2013-
program/index. Last Accessed:
08/03/2016.
California
anatoxin-a: 90 |ig/L
unclear
Butler N, Carlisle J, Kaley KB, &
Linville R (2012). Toxicological
Summary and Suggested Action
Levels to Reduce Potential Adverse
Health Effects of Six Cyanotoxins.
httD://www. waterboards.ca.gov/water
cylindrospermopsin: 4 j.ig/L
unclear
microcystins: 0.8 j.ig/L
unclear
issues/Drograms/Deer review/docs/cali
f cvanotoxins/cvanotoxins( rff.
Last Accessed: 08/03/2016.
Illinois
microcystin-LR: > 10 |ig/L
appropriate lake management personnel and Illinois
EPA staff will be notified; follow-up monitoring by
the Illinois EPA may occur as professional judgment
dictates and staff laboratory, and financial resources
allow
Illinois Environmental Protection
Agency (2015). 2013 Statewide
Harmful Algal Bloom Program,
http ://epa. illinois.gov/topics/water-
quality/monitoring/algal-bloom/2013-
program/index. Last Accessed:
08/03/2016.
Iowa
microcystin: > 20 |ig/L
warnings are posted at state park beaches
Iowa Environmental Council (2015).
State Park Beach Advisories Report.
Updated September 3, 2015.
httD://www. iaenvironinent.org/webres/
F i le/P to gra m%20Pub li ca ti o ns/2015 %
20State%20Park%20Beach%20Advis
ories%20Retx)rt.txtf. Last Accessed:
08/03/2016.
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
B-8
-------
State
Reereational Water Guideline Level
Recommended Aetion
Ret'crcnee
Nebraska
microcystin: > 20 |ig/L
health alert; signs posted advising public to use
caution; affected swimming beaches will be closed;
boating and other recreational activities will be
allowed, but public advised to use caution and avoid
prolonged exposure to the water
Nebraska Department of
Environmental Quality and Nebraska
Department of Health and Human
Services: Division of Public Health
(2016). Fact Sheet: Precautions and
facts regarding toxic algae at Nebraska
Lakes.
httD://dea.ne.gov/NDEOProe.nsi70nW
eb/ENV042607. Last Accessed:
08/03/2016.
North Dakota
blue-green algae bloom is present AND
microcystin-LR: < 10 |ig/L
issue advisory
North Dakota Department of Health:
Division of Water Quality (2016).
Blue-green algae advisories and
warnings.
httD://www. ndhealth. eov/WO/SW/H A
B s/H AB s I nfo rma t i o n/B lue -
blue-green algae bloom is present over a
significant portion of the lake AND
microcystin-LR: > 10 |ig/L
issue warning
greenLakeListings 20160808.txtf.
Last Accessed: 10/18/2016.
Ohio
anatoxin-a: 300 |ig/L
issue no contact advisory
Kasich JR, Taylor M, Butler CW,
Zehringer J, & Hodges R (2016). State
of Ohio Harmful Algal Bloom
Response Strategy For Recreational
Waters.
fatt o ://e oa. ofai o. gov/oo rta ls/3 S/faab/HA
anatoxin-a: 80 |ig/L
issue recreational public health advisory
cylindrospermopsin: 20 j.ig/L
issue no contact advisory
cylindrospermopsin: 5 j.ig/L
issue recreational public health advisory
microcystin-LR: 20 |ig/L
issue no contact advisory
BResDonseSt.rategv.Ddf. Last
Accessed: 08/03/2016.
microcystin-LR: 6 j.ig/L
issue recreational public health advisory
saxitoxin: 0.8 j.ig/L
issue recreational public health advisory
saxitoxin: 3 |ig/L
issue no contact advisory
Nebraska
microcystin: > 20 (ig/L
health alert; signs posted advising public to use
caution; affected swimming beaches will be closed;
boating and other recreational activities will be
Nebraska Department of
Environmental Quality and Nebraska
Department of Health and Human
Services: Division of Public Health
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
B-9
-------
State
Reereational Water Guideline Level
Recommended Aetion
Ret'crcnee
allowed, but public advised to use caution and avoid
prolonged exposure to the water
(2016). Fact Sheet: Precautions and
facts regarding toxic algae at Nebraska
Lakes.
httD://dea.ne.gov/NDEOProe.nsi70nW
eb/ENV042607. Last Accessed:
08/03/2016.
Ohio
anatoxin-a: 300 |ig/L
issue no contact advisory
Kasich JR, Taylor M, Butler CW,
Zehringer J, & Hodges R (2015). State
of Ohio Harmful Algal Bloom
Response Strategy For Recreational
Waters.
fat! d ://e oa. olii o. eov/do rta ls/3 5/faab/HA
anatoxin-a: 80 |ig/L
issue recreational public health advisory
cylindrospermopsin: 20 j.ig/L
issue no contact advisory
cylindrospermopsin: 5 j.ig/L
issue recreational public health advisory
microcystin-LR: 20 j.ig/L
issue no contact advisory
BResDonseStrateev.Ddf. Last
Accessed: 08/03/2016.
microcystin-LR: 6 j.ig/L
issue recreational public health advisory
saxitoxin: 0.8 |ig/L
issue recreational public health advisory
saxitoxin: 3 jrg/L
issue no contact advisory
Vermont
anatoxin-a: > 10 j.ig/L
close recreational beaches
Vermont Department of Health
(2015). Cyanobacteria (Blue-green
Algae) Guidance for Vermont
Communities.
lit!d ://healthvermo fit. eov/e nvi ro/b g al
Accessed: 08/03/2016.
cylindrospermopsin: > 10 |ig/L
close recreational beaches
microcystin-LR (equivalents): > 6 j.ig/L
close recreational beaches
visible known blue-green algae bloom/scum
or an unknown, potentially blue-green algae
(i.e., not pollen), bloom/scum
close recreational beaches
Washington
anatoxin-a: 1 |ig/L
tier 2: local health posts WARNING sign; local
health takes additional site-specific steps; minimum
weekly sampling. In addition, if history of high
toxicity, or reports of illness, pet death than tier 3:
local health posts DANGER sign; lake closed
Hardy J, & Washington State
Department of Health (2008).
Washington State Recreational
Guidance for Microcystins
(Provisional) and Anatoxin-a
(Interim/Provisional).
httD://www.doh.wa.eov/Portals/l/Doc
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
B-10
-------
State
Recreational Water Guideline Level
Recommended Action
Reference
uments/4400/2 reeguide.pdf.
Last Accessed: 08/03/2016.
bloom is forming or a bloom scum is visible
(toxic algae may be present); toxin levels do
not exceed thresholds
tier 1: local health posts CAUTION sign; samples
taken and sent for toxicity tests; weekly sampling
until bloom dissipates
Hardy J, & Washington State
Department of Health (2011).
Washington State Provisional
Recreational Guidance for
Cylindrospermopsin and Saxitoxin.
lift d ://www. doli. wa. gov/po rta Is/ 1/docu
ments/4400/32
cvlindrosax%20reDort.txtf. Last
Accessed: 08/03/2016.
cylindrospermopsin: 4.5 |ig/L
tier 2: local health posts WARNING sign; local
health takes additional site-specific steps; minimum
weekly sampling. In addition, if history of high
toxicity, or reports of illness, pet death than tier 3:
local health posts DANGER sign; lake closed.
microcystins: 6 |ig/L
tier 2: local health posts WARNING sign; local
health takes additional site-specific steps; minimum
weekly sampling. In addition, if history of high
toxicity, or reports of illness, pet death than tier 3:
local health posts DANGER sign; lake closed.
Hardy J, & Washington State
Department of Health (2008).
Washington State Recreational
Guidance for Microcystins
(Provisional) and Anatoxin-a
(Interim/Provisional).
httD://www.doh.wa.eov/Portals/l/Doc
iiinents/4400/j reeguide.pdf.
Last Accessed: 08/03/2016.
Washington
(continued)
saxitoxin: 75 |ig/L
tier 2: local health posts WARNING sign; local
health takes additional site-specific steps; minimum
weekly sampling. In addition, if history of high
toxicity, or reports of illness, pet death than tier 3:
local health posts DANGER sign; lake closed.
Hardy J, & Washington State
Department of Health (2011).
Washington State Provisional
Recreational Guidance for
Cylindrospermopsin and Saxitoxin.
lit! p ://www. doli. wa. eov/po rta Is/ 1/docu
ments/4400/332-118-
cvlindrosax%20report.pdf. Last
Accessed: 08/03/2016.
States with Guidelines Based on Cyanobacteria and Cvanotoxin(s)
Indiana
anatoxin-a: 80 |ig/L
issue recreation advisory
Indiana Department of Environmental
Management (2015). Addressing
blue-green algae: 100,000 cells/mL
issue recreation advisory
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
B-ll
-------
State
Recreational Water Guideline Level
Recommended Action
Reference
Concerns About Blue-Green Algae:
Indiana Reservoir and Lake Update.
htto://www. in. gov/idem/aleae/2310.lit
cylindrospermopsin: 5 |ig/L
issue recreation advisory
microcystin-LR: 20 |ig/L
close beaches
microcystin-LR: 6 (ig/L
issue recreation advisory
m. Last Accessed: 08/03/2016.
Kansas
cyanobacteria: > 10,000,000 cells/mL
recommended that all in-lake recreation cease and
that picnic, camping and other public land activities
adjacent to affected waters be closed
Kansas Department of Health &
Environment (2015). Guidelines for
Addressing Harmful Algal Blooms in
Kansas Recreational Waters.
littp://www.kdlieks.gov/aleae-
cyanobacteria: > 250,000 cells/mL
issue public health warning
cyanobacteria: > 80,000 and < 250,000
cells/mL
issue public health watch
illness/download/t icv.odf.
Last Accessed: 08/03/2016.
microcystin: > 2,000 |ig/L
recommended that all in-lake recreation cease and
that picnic, camping and other public land activities
adjacent to affected waters be closed
microcystin: > 20 j.ig/L
issue public health warning
microcystin: > 4 and < 20 |ig/L
issue public health watch
Kentucky
blue-green algae: > 100,000 cells/mL
issue an harmful algal bloom (HAB) advisory
Kentucky Department for
Environmental Protection (2014).
Harmful Algal Blooms: Background.
lit! d ://wa te r.kv. eov/wate raua li fv/Docu
ir F s/H AB%20Backgr
ound%20Fact%20Sheet.odf. Last
Accessed: 08/03/2016.
microcystins: > 20 |ig/L
issue recreational use advisory
Commonwealth of Kentucky:
Department for Environmental
Protection Division of Water (2015).
Harmful Algal Blooms.
httD://water.kv.eov/wateraualitv/Daees
/HABS.asDx. Last Accessed:
08/03/2016.
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
B-12
-------
State
Recreational Water Guideline Level
Recommended Action
Reference
Maryland
Microcystis aeruginosa or other potential
microcystin producing blue-green algae
> 40,000 cells/mL, and samples contain
microcystins: >10 ppb
put up signs advising public of health risk, notify
local press (through joint DHMH, DNR, MDE press
release) and coordinate with local health department,
place advisory information on DNR web site (Eyes
on the Bay), Maryland Healthy Beaches web site if a
swimming beach is affected, or other local web site.
MDE will initiate emergency closure to shellfish
harvesting if warranted, and coordinate with DNR
Natural Resource Police
Maryland Department of Natural
Resources (2010). Harmful Algal
Bloom (HAB) Monitoring and
Management SOP. SOP document
(with edits) sent via email
correspondence with Catherine
Wazniak, Program Manager at the MD
DNR, on February 22, 2016.
presence of potentially toxic algae
issue algae bloom beach alert
Maryland Department of Natural
Resources (2010). Harmful Algal
Bloom (HAB) Monitoring and
Management SOP. SOP document
(with edits) sent via email
correspondence with Catherine
Wazniak, Program Manager at the MD
DNR, on February 22, 2016.
Massachusetts
blue-green algae: > 50,000 cells/mL
toxin testing of lysed cells should be done to ensure
that guideline of 14 ppb is not exceeded
Massachusetts Bureau of
Environmental Health (2015). MDPH
Guidelines for Cyanobacteria in
Freshwater Recreational Water Bodies
in Massachusetts. Boston,
Massachusetts.
lifto://www. mass, gov/eolilis/docs/doli/
e nvi ro nine nta 1/exDOSiire/oro toco 1-
cvanobacteria.Ddf. Last Accessed:
08/03/2016.
blue-green algae: > 70,000 cells/mL
post an advisory against contact with the water
microcystins: >14 |ig/L
post an advisory against contact with the water
visible cyanobacteria scum or mat is evident
MDPH recommends an immediate posting by the
local health department, state agency, or relevant
authority to advise against contact with the water
body
New York
visible HAB
prohibit wading, swimming, diving and any water
contact activities in the swimming area; post beach
closure and advisory signs at the beach and other
shoreline access areas; contact local health
department
June, Stephanie. Senior Sanitarian at
the New York State Department of
Health. Email correspondence on Feb.
23,2016.
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
B-13
-------
State
Recreational Water Guideline Level
Recommended Action
Reference
microcystins: 10 |ig/L
prohibit wading, swimming, diving and any water
contact activities in the swimming area; post beach
closure and advisory signs at the beach and other
shoreline access areas; contact local health
department; to reopen swim areas:
• water must be visibly clear of HABs or associated
material for one day
• at that time, a water sample is to be collected and
tested for microcystins
• if the sample indicates toxin levels <10 |ig/L and
the HAB has not returned to the swim area, the signs
may be removed and the beach may be reopened
June, Stephanie. Senior Sanitarian at
the New York State Department of
Health. Email correspondence on Feb.
23,2016.
bloom: credible report or digital imagery of a
bloom determined as likely to be potentially
toxic cyanobacteria by the Department of
Environmental Conservation (DEC) or
Department of Health (DOH) staff; a
descriptive field report from professional staff
or trained volunteer may be used as a report in
absence of digital images; for all other
surveillance reports received from the general
public, lay monitors, etc., DEC HABs
Program staff will determine if a bloom is
suspicious and whether collection of a sample
is feasible or warranted
post DEC blue-green algal bloom notice: suspicious
bloom
Gorney, Rebecca. Research Scientist
at New York State Department of
Environmental Conservation. Email
correspondence on Feb. 23, 2016.
blue green chlorophyll a: >25-30 |ig/L
post DEC blue-green algal bloom notice: confirmed
bloom
Gorney, Rebecca. Research Scientist
at New York State Department of
Environmental Conservation. Email
correspondence on Feb. 23, 2016.
potential toxin-producing cyanobacteria taxa:
>50% of algae present
post DEC blue-green algal bloom notice: confirmed
bloom
Gorney, Rebecca. Research Scientist
at New York State Department of
Environmental Conservation. Email
correspondence on Feb. 23, 2016.
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
B-14
-------
State
Reereational Water Guideline Level
Recommended Aetion
Ret'crcnee
microcystin-LR: 4 |ig/L
post DEC blue-green algal bloom notice: confirmed
bloom
Gorney, Rebecca. Research Scientist
at New York State Department of
Environmental Conservation. Email
correspondence on Feb. 23, 2016.
anatoxin-a or other cyanotoxins: high risk of
exposure based on consult among DEC or
DOH staff
post DEC blue-green algal bloom notice: confirmed
bloom
Gorney, Rebecca. Research Scientist
at New York State Department of
Environmental Conservation. Email
correspondence on Feb. 23, 2016.
microcystin-LR: 10 |ig/L in open water
sample
post DEC blue-green algal bloom notice: confirmed
with high toxins
Gorney, Rebecca. Research Scientist
at New York State Department of
Environmental Conservation. Email
correspondence on Feb. 23, 2016.
microcystin-LR: 20 |ig/L in shoreline sample
post DEC blue-green algal bloom notice: confirmed
with high toxins
New York State Department of
Environmental Conservation. Water
Clarity Fact Sheet.
liftd://www.dec. nv. eov/docs/water txtf
/cslaDlkDara.txtf. Last Accessed:
10/23/2015.
Oklahoma
cyanobacteria: 100,000 cell/mL
issue advisory
Oklahoma Legislature (2012). SB 259
Bill Summary.
httD://webserverl.lsb.state.ok.us/CF/2
12%20SUPPORT%20DOCUMENTS/
iuse/SB259%20ccr%20a
%20billsum.doc. Last Accessed:
08/03/2016.
microcystin: > 20 |ig/L
issue advisory
Oregon
anatoxin-a: > 20 j.ig/L
issue public health advisory
Oregon Health Authority (2016).
Oregon Harmful Algae Bloom
Surveillance (HABS) Program Public
Health Advisory Guidelines: Harmful
Algae Blooms in Freshwater Bodies.
httDs://Dublic.health.oreeon.eov/Healt
cylindrospermopsin: > 20 |ig/L
issue public health advisory
microcystin: >10 j.ig/L
issue public health advisory
Microcystis: > 40,000 cells/mL
issue public health advisory
Planktothrix: > 40,000 cells/mL
issue public health advisory
hvEnvironments/Recreation/HarmfulA
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
B-15
-------
State
Reereational Water Guideline Level
Recommended Aetion
Ret'crcnee
lgaeBlooms/Documents/HABPublicH
ealthAdvisorvGuidelines.txif. Last
Accessed: 08/03/2016.
saxitoxin: > 10 |ig/L
issue public health advisory
toxigenic species: > 100,000 cells/mL
issue public health advisory
visible scum with documentation and testing
issue public health advisory
Rhode Island
cyanobacteria: > 70,000 cells/mL
issue health advisory
Rhode Island Department of
Environmental Management, & Rhode
Island Department of Health (2013).
Cyanobacteria Related Public Health
Advisories in Rhode Island.
httD://www.health. ri.gov/Dublications/
da tareDorts/2013CvanobacteriaB looms
InRliodelsland.Ddf. Last Accessed:
08/03/2016.
microcystin-LR: > 14 (ig/L
issue health advisory
visible cyanobacteria scum or mat
issue health advisory
Texas
>100,000 cell/mL of cyanobacterial cell
counts and >20|ig/L microcystin
blue-green algae awareness level advisory
U.S. EPA (United States
Environmental Protection Agency)
(2016). What are the Standards or
Guidelines for
Cyanobacteria/Cyanotoxin in
Recreational Water.
https://www.epa.gov/nutrient-policy-
data/guidelines-and-
recommendations#what3. Last
Accessed: 08/03/2016.
Utah
anatoxin-a: > 20 (ig/L
issue caution advisory; post CAUTION sign; weekly
sampling recommended
Utah Department of Environmental
Quality and Department of Health
(2015). Utah Guidance for Local
Health Departments: Harmful Algal
Blooms and Human Health.
httD://www.dea.utah.eov/ToDics/Wate
r/Hea IthAdviso rvPa nel/docs/07 Jitl/Ha r
nifiilAlgalBlootns.pdf. Last Accessed:
08/03/2016.
blue-green algae: >10,000,000 cells/mL
issue danger advisory; post DANGER sign; weekly
sampling recommended; consider closure
blue-green algae: 100,000-10,000,000
cells/mL
issue warning advisory; post WARNING sign;
weekly sampling recommended
blue-green algae: 20,000-100,000 cells/mL
issue caution advisory; post CAUTION sign; weekly
sampling recommended
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
B-16
-------
State
Reercational Water Guideline Level
Rceommended Aetion
Ret'crcnee
large scum mat layer
issue danger advisory; post DANGER sign; weekly
sampling recommended; consider closure
microcystin: > 2,000 |ig/L
issue danger advisory; post DANGER sign; weekly
sampling recommended; consider closure
microcystin: 20-2,000 |ig/L
issue warning advisory; post WARNING sign;
weekly sampling recommended
microcystin: 4-20 (ig/L
issue caution advisory; post CAUTION sign; weekly
sampling recommended
reports of animal illnesses or death
issue warning advisory; post WARNING sign;
weekly sampling recommended
reports of human illness
issue danger advisory; post DANGER sign; weekly
sampling recommended; consider closure
Virginia
blue-green algal "scum" or "mats" on water
surface
immediate public notification to avoid all
recreational water contact where bloom is present;
continue weekly sampling
Virginia Department of Health
(Division of Environmental
Epidemiology) (2012). Virginia
Recreational Water Guidance for
Microcystin and Microcystis Blooms:
Provisional Guidance.
httDs://www.vdh.vi rginia.gov/eDide mi
microcystin: > 6 |ig/L
immediate public notification to avoid all
recreational water contact where bloom is present;
continue weekly sampling
Microcystis: > 100,000 cells /mL
immediate public notification to avoid all
recreational water contact where bloom is present;
continue weekly sampling
o lo gv/dee/H AB S/docume nts/VDHMic
rocvstisGuidance.Ddf. Last Accessed:
08/03/2016.
Microcystis: 20,000 to 100,000 cells/mL
notify public through press release and/or signage;
advise people and pet-owners that harmful algae are
present; initiate weekly water sampling
Microcystis: 5,000 to < 20,000 cells/mL
local agency notification; initiate bi-weekly water
sampling
States with Qualitative Guidelines Only
Delaware
thick green, white, or red scum on surface of
pond
post water advisory signs
Delaware Department of Natural
Resources and Environmental Control:
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
B-17
-------
State
Recreational Water Guideline Level
Recommended Action
Reference
Division of Water. Blue-Green Algae
in Delaware.
htt d ://www. dnrec .delaware. eov/wr/IN
FORMATION/OTHERINFO/Pages/B
lue-GreenAlgae.asDx. Last Accessed:
08/03/2016.
Florida
cyanobacteria bloom
health advisory
Florida Department of Environmental
Protection (2016). South Florida Algal
Bloom Monitoring and Response.
httDs://deDiiewsroom.wordDress.com/s
Giitli-flo rida-algal-b loo m-mo ni to ri ng-
and-resDonse/. Last Accessed:
08/16/2016.
Florida Department of Health (2016).
Blue-Green Algae (Cyanobacteria).
lift d ://www.:flo ridahealth. gov/e nvi ro n
me nta 1-liea Itli/aauat ic-
toxins/cv anobacteria.html. Last
Accessed: 08/16/2016.
Montana
reservoirs that seem stagnated and harbor
large quantities of algae
the Montana Department of Environmental Quality
advises people to avoid swimming in ponds, lakes, or
reservoirs
State of Montana Newsroom (2015).
DEQ Issues Advisory on Blue-Green
Algae Blooms: Ponds, Lakes, and
Reservoirs Most Often Affected.
httD://news. int. gov/Home/ArtMID/244
Advisory-on-Blue-Green-Algae-
Blooms. Last Accessed: 08/03/2016.
North Carolina
visible discoloration or surface scum
Microcystin testing
North Carolina Health and Human
Services: Division of Public Health
(2014). Occupational &
Environmental Epidemiology:
Cyanobacteria (Blue-green Algae).
httD://eDi.Dublichealth.nc.eov/oee/a z/
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
B-18
-------
State
Reereational Water Guideline Level
Rceommended Aetion
Ret'crcnee
algae.html. Last Accessed:
08/03/2016.
West Virginia
blue-green algal blooms observed and
monitored
issue public health advisory
West Virginia Department of Health &
Human Resources (2015). DHHR
Continuing to Monitor Blue-Green
Algal Blooms on the Ohio River:
Residents Advised to Adhere to Public
Health Advisory.
httD://www.dhhr.wv. eov/News/201S/P
a ges/DHHR-Co nti nui ng-to -Mo ni to r-
Blue-Green-Algal-Blooms-on-tlie-
Ohio-River%3B-Residents-Advised-
to-Adhere-to-Public-Health-
Advisorv.asDx. Last Accessed:
08/03/2016.
Human Health Recreational Ambient Water Quality Criteria or B-19
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
-------
Human Health Recreational Ambient Water Quality Criteria or B-20
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
-------
APPENDIX C. LITERATURE SEARCH DOCUMENTATION
The recreational ambient water quality criteria document for microcystins,
cylindrospermopsin, and cyanobacteria relied significantly on information identified, reviewed,
and synthesized in U.S. EPA's Health Effects Support Document for the Cyanobacterial Toxin
Microcystins, Heath Effects Support Document for the Cyanobacterial Toxin
Cylindrospermopsin, Drinking Water Health Advisory for the Cyanobacterial Microcystin
Toxins, and Drinking Water Health Advisory for the Cyanobacterial Toxin Cylindrospermopsin
((U.S. EPA (2015c); U.S. EPA (2015d)); (U.S. EPA 2015a; U.S. EPA 2015b). EPA conducted
supplemental literature searches to answer additional questions related to recreational exposures,
exposure factors, and to identify new health data.
For the Health Effects Support Documents, EPA conducted a comprehensive literature
search from January 2013 to May 2014 using Toxicology Literature Online (TOXLINE),
PubMed, and Google Scholar. EPA assembled available information on occurrence;
environmental fate; mechanisms of toxicity; acute, short-term, subchronic, and chronic toxicity
and cancer in humans and animals; and toxicokinetics and exposure. For a detailed description of
the literature review search and strategy, see the Health Effects Support Documents for
microcystins and cylindrospermopsin (U.S. EPA 2015c; U.S. EPA 2015d).
EPA conducted supplemental literature searches in September 2015 to capture references
published since the completion of the Health Effects Support Documents literature searches and
to account for the recreational exposure scenario. The specific questions investigated include:
1. What levels of anatoxin-a, cylindrospermopsin, or microcystins are humans—of all
ages, including children—exposed to through recreational use (activities) in
freshwaters or marine waters from incidental ingestion, inhalation, and dermal
exposure routes?
2 What health effects information for humans or animals exposed to
cylindrospermopsin or microcystins (through ingestion, inhalation, and dermal
exposure routes) has been published since the health effects literature searches were
conducted for EPA's 2015 Health Effects Support Documents for cylindrospermopsin
and microcystins?
3. What recreational water use safety levels or criteria have been set for microcystins or
cylindrospermopsin by states or international governments, and how did they derive
them?
4. What new information, if any, is available regarding how aquatic recreational
exposure ingestion rates in children differ among age groups between 0 and 18 years?
5. What incidents of companion animal (e.g., dogs, horses) or livestock poisonings,
including mortality or adverse health effects, due to exposure to cyanotoxins in
freshwaters, marine waters, or beaches have occurred in the past 15 years?
Specifically, when and where did these incidents occur, to which cyanotoxin were the
animals exposed, how were they exposed, and what were the weights and breeds of
the affected animal(s)?
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
C-l
-------
EPA implemented a unique literature search strategy to address each research question.
Trial searches were conducted, and results were evaluated to refine the search strategies (e.g., to
reduce retrieval of citations unrelated to the research questions). The search strings were refined
to improve the relevancy of the results. The literature search strategies implemented for each
research question are subsequently detailed.
Research Question 1: What levels of anatoxin-a, cylindrospermopsin, or microcystins are
humans—of all ages, including children—exposed to through recreational use (activities) in
freshwaters or marine waters, from incidental ingestion, inhalation, and dermal exposure
routes?
EPA searched the bibliographic databases, PubMed and Web of Science (WoS), to
identify candidate journal article literature relevant to human exposure to anatoxin-a,
cylindrospermopsin, or microcystins through recreational activities. PubMed and WoS contain
peer-reviewed journal abstracts and articles on various biological, medical, public health, and
chemical topics. The WoS search string differs slightly from the PubMed search string due to
how the search engines treat search terms with more than one word. Both search strings are
presented below.
Results
The searches returned 321 journal articles after removing duplicates between PubMed
and WoS results. Based on a screening review of each article's title and abstract, EPA retrieved 9
articles that appeared to be studies that measured, reviewed, or estimated human recreational
exposure to cyanotoxins.
PubMed Search:
("A. lemmermannii Raphidiopsis mediterranean OR Anabaena flos-aquae OR jlos-aquae OR
anatoxin-a OR Aphanizomenon OR cylindrospermopsin OR "C. raciborskii" OR Cuspidothrix
OR Cylindrospermopsis OR Cylindrospermum OR "Cylindrospermopsis raciborskii" OR
Dolichospermum OR "M aeruginosa" OK Microcystis OR microcystin OR microcystins OR
Oscillatoria OR Planktothrix OR Phormidium OR Tychonema OR Woronichinia)
AND
("boogie board" OR "boogie boarding" OR "jet ski" OR "jet skier" OR "jet skiers" OR "jet
skiing" OR "water ski" OR "water skier" OR "water skiers" OR "water skiing" OR aerosol OR
boat OR boating OR boats OR bodyboard OR bodyboarding OR canoe OR canoeing OR canoes
OR capsize OR capsized OR dermal OR inhalation OR inhale OR kayak OR kayaker OR
kayaking OR kayaks OR kneeboard OR kneeboarding OR paddle OR paddling OR raft OR
rafting OR rafts OR recreation OR recreational OR rowing OR skin OR surf OR surfer OR
surfing OR swim OR swimmer OR swimmers OR swimming OR tubing OR wading OR
wakeboarding OR wakeboard)
AND
("marine water" OR "surface water" OR beach OR beaches OR estuaries OR estuarine OR
estuary OR "fresh water" OR freshwater OR lake OR lakes OR ocean OR oceans OR pond OR
ponds OR reservoir OR reservoirs OR river OR rivers OR sea OR stream OR streams OR water)
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
C-2
-------
Filters: English
Date search was conducted: 10/9/2015
Publication dates searched: 1/1/1995 - 10/9/2015
Web of Science Search:
("lemmermannii Raphidiopsis mediterranean OR Anabaena flos-aquae OR jlos-aquae OR
anatoxin OR Aphanizomenon OR cylindrospermopsin OR "C. raciborskii" OR Cuspidothrix OR
Cylindrospermopsis OR Cylindrospermum OR "Cylindrospermopsis raciborskii" OR
Dolichospermum OR "M aeruginosa" OK Microcystis OR microcystin OR microcystins OR
Oscillatoria OR Planktothrix OR Phormidium OR Tychonema OR Woronichinia)
AND
("boogie board" OR "boogie boarding" OR "jet ski" OR "jet skier" OR "jet skiers" OR "jet
skiing" OR "water ski" OR "water skier" OR "water skiers" OR "water skiing" OR aerosol OR
boat OR boating OR boats OR bodyboard OR bodyboarding OR canoe OR canoeing OR canoes
OR capsize OR capsized OR dermal OR inhalation OR inhale OR kayak OR kayaker OR
kayaking OR kayaks OR kneeboard OR kneeboarding OR paddle OR paddling OR raft OR
rafting OR rafts OR recreation OR recreational OR rowing OR skin OR surf OR surfer OR
surfing OR swim OR swimmer OR swimmers OR swimming OR tubing OR wading OR
wakeboarding OR wakeboard)
AND
("marine water" OR "surface water" OR beach OR Beaches OR estuaries OR estuarine OR
estuary OR "fresh water" OR freshwater OR lake OR lakes OR ocean OR oceans OR pond OR
ponds OR reservoir OR reservoirs OR river OR rivers OR sea OR stream OR streams OR water)
Filters: English
Date search was conducted: 10/9/2015
Publication dates searched: 1/1/1995-10/9/2015
Research Question 2: What health effects information for humans or animals exposed to
microcystins, cylindrospermopsin, or anatoxin-a (through ingestion, inhalation, and
dermal exposure routes) has been published since the health effects literature searches
were conducted for EPA's 2015 Health Effects Support Documents for Cylindrospermopsin
and Microcystins?
EPA searched PubMed and WoS to identify candidate journal article literature relevant to
health effects associated with exposure to anatoxin-a, cylindrospermopsin, or microcystins. The
WoS search string differs slightly from the PubMed search string due to how the search engines
treat search terms with more than one word. Both search strings are presented below.
Results
The searches returned 1,000 journal articles after removing duplicates between PubMed
and WoS results. Based on a screening review of each article's title and abstract, EPA retrieved
40 articles that appeared to be prospective human epidemiological studies (n = 1), ecological
human epidemiologic studies (n = 2), reviews of human health effects (n = 4), in vivo animal
studies (n = 30), or reviews of in vivo animal studies (n = 3).
PubMed Search:
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
C-3
-------
("A. lemmermannii Raphidiopsis mediterranean OR Anabaena flos-aquae OR flos-aquae OR
anatoxin-a OR Aphanizomenon OR cylindrospermopsin OR "C. raciborskii" OR Cuspidothrix
OR Cylindrospermopsis OR Cylindrospermum OR "Cylindrospermopsis raciborskii" OR
Dolichospermum OR "M aeruginosa" OK Microcystis OR microcystin OR microcystins OR
Oscillatoria OR Planktothrix OR Phormidium OR Tychonema OR Woronichinia)
AND
("non cancer" OR "blurred vision" OR "cell damage" OR "cellular damage" OR "health effect"
OR "health endpoint" OR "health outcome" OR "health risk" OR "loss of protein" OR "loss of
water" OR "micronucleated binucleate cell" OR abdominal pain OR ache OR acute OR alanine
aminotransferase OR allergic OR allergies OR allergy OR aspartate aminotransferase OR blister
OR blistered OR blisters OR cancer OR carcinogen OR carcinogenic OR carcinogens OR
chronic OR clinical OR cough OR dermal OR detoxification OR detoxify OR develop OR
development OR developmental OR dialysis OR diarrhea OR disease OR DNA OR dyspnea OR
electrolyte OR emergency room OR enzyme OR enzymes OR epidemiologic OR
epidemiological OR epidemiology OR epilepsy OR epileptic OR epithelium OR eye OR failure
OR fever OR gastrointestinal OR genetox OR genotoxic OR glutamyltransferase OR head OR
hematologic OR hematological OR hepatic OR histopathologic OR histopathological OR
histpathology OR hospital OR hospitalizations OR hospitals OR hospitalization OR ill OR
illness OR illnesses OR intoxicate OR intoxicated OR irritate OR irritated OR kidney OR larynx
OR lesion OR lesions OR liver OR lung OR lymph OR lymph nodes OR lymphatic OR
metabolic OR metabolism OR mucosa OR mutate OR mutated OR mutation OR mutations OR
nausea OR necrosis OR neonatal OR neonate OR neonates OR neoplasm OR neurologic OR
neurological OR noncancer OR oral OR organ OR pain OR placenta OR pneumonia OR
polymorphism OR polymorphisms OR prenatal OR red blood cell OR renal OR reproduction OR
respiratory OR seizure OR sick OR sickness OR skin OR stomach OR subacute OR subchronic
OR symptom OR symptoms OR teratogen OR teratogenic OR teratogens OR throat OR toxic
OR toxicity OR trachea OR tumor OR tumors OR urinary OR urine OR vomit OR vomiting OR
conjugate OR conjugated OR diagnose OR diagnosis OR diagnosed OR diagnoses)
Filters: English
Date search was conducted: 10/9/2015
Publication dates searched: 1/1/2014-10/9/2015
Web of Science Search:
("lemmermannii Raphidiopsis mediterranean OR Anabaena flos-aquae OR flos-aquae OR
anatoxin OK Aphanizomenon OR cylindrospermopsin OR "C. raciborskii" OR Cuspidothrix OR
Cylindrospermopsis OR Cylindrospermum OR "Cylindrospermopsis raciborskii" OR
Dolichospermum OR "M aeruginosa" OK Microcystis OR microcystin OR microcystins OR
Oscillatoria OR Planktothrix OR Phormidium OR Tychonema OR Woronichinia)
AND
("non cancer" OR "blurred vision" OR "cell damage" OR "cellular damage" OR "health effect"
OR "health endpoint" OR "health outcome" OR "health risk" OR "micronucleated binucleate
cell" OR abdominal pain OR ache OR acute OR alanine aminotransferase OR allergic OR
allergies OR allergy OR aspartate aminotransferase OR blister OR blistered OR blisters OR
cancer OR carcinogen OR carcinogenic OR carcinogens OR chronic OR clinical OR cough OR
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
C-4
-------
dermal OR detoxification OR detoxify OR develop OR development OR developmental OR
dialysis OR diarrhea OR disease OR DNA OR dyspnea OR electrolyte OR emergency room OR
enzyme OR enzymes OR epidemiologic OR epidemiological OR epidemiology OR epilepsy OR
epileptic OR epithelium OR eye OR failure OR fever OR gastrointestinal OR genetox OR
genotoxic OR glutamyltransferase OR head OR hematologic OR hematological OR hepatic OR
histopathologic OR histopathological OR histpathology OR hospital OR hospitalizations OR
hospitals OR hospitalization OR ill OR illness OR illnesses OR intoxicate OR intoxicated OR
irritate OR irritated OR kidney OR larynx OR lesion OR lesions OR liver OR lung OR lymph
OR lymph nodes OR lymphatic OR metabolic OR metabolism OR mucosa OR mutate OR
mutated OR mutation OR mutations OR nausea OR necrosis OR neonatal OR neonate OR
neonates OR neoplasm OR neurologic OR neurological OR noncancer OR oral OR organ OR
pain OR placenta OR pneumonia OR polymorphism OR polymorphisms OR prenatal OR red
blood cell OR renal OR reproduction OR respiratory OR seizure OR sick OR sickness OR skin
OR stomach OR subacute OR subchronic OR symptom OR symptoms OR teratogen OR
teratogenic OR teratogens OR throat OR toxic OR toxicity OR trachea OR tumor OR tumors OR
urinary OR urine OR vomit OR vomiting OR conjugate OR conjugated OR diagnose OR
diagnosis OR diagnosed OR diagnoses)
Filters: English
Date search was conducted: 10/9/2015
Publication dates searched: 1/1/2014-10/9/2015
WoS research areas searched: Environmental Sciences Ecology OR Marine Freshwater Biology
OR Toxicology OR Pharmacology Pharmacy OR Public Environmental Occupational Health OR
Microbiology OR Immunology OR Biotechnology Applied Microbiology OR Biochemistry
Molecular Biology OR Research Experimental Medicine OR Water Resources OR Infectious
Disease OR Science Technology Other Topics OR Life Sciences Biomedicine Other Topics OR
Gastroenterology Hepatology OR Pediatrics.
Research Question 3: What recreational water use safety levels or criteria have been set for
microcystins or cylindrospermopsin by states or international governments and how did
they derive them?
To identify state-level recreational guidelines for cyanobacteria and cyanotoxins, EPA
searched the websites of state-level departments of public health, environmental health, and
natural resources for all 50 U.S. states. If relevant recreational guidelines were not found by
searching state-level websites, EPA conducted Google searches of the internet using state names,
key terms for cyanobacteria and cyanotoxins (e.g., harmful algal bloom, blue green algae,
microcystin, cylindrospermopsin), and key terms for guidelines (e.g., advisory, guidance,
guideline, standard, regulation). For international governments, EPA used the 2012 report,
Current Approaches to Cyanotoxin Risk Assessment, Risk Management and Regulations in
Different Countries, by Dr. Ingrid Chorus, Federal Environment Agency, Germany, to identify
international government recreational safety levels for cyanobacteria and cyanotoxins. In
addition, EPA implemented the same search strategy as used for U.S. states to identify updated
international recreational guidelines or guideline levels not featured in the 2012 report by Dr.
Ingrid Chorus.
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
C-5
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Research Question 4: What new information, if any, is available regarding how aquatic
recreational exposure ingestion rates in children differ among age groups between 0 and 18
years?
Search of Bibliographic Databases
EPA searched PubMed, WoS, and Google Scholar to identify literature that has cited, or
is similar (based on terms identified in the titles and abstracts) to, the studies that provide water
ingestion data for swimmers or during water recreational activities in EPA's (2011) Exposure
Factors Handbook (i.e.,(Dorevitch et al. 2011; Dufour et al. (2006); Schets et al. 2011). The
PubMed and WoS searches were conducted on 10/9/2015, the publication dates searched were
1/1/2011 to 10/9/2015, and an English filter was applied. The Google Scholar search was
conducted on 10/9/2015 and could not be limited by year or language.
Results
Together all three searches returned 341 journal articles. Duplicates were removed
between PubMed and Wos, but this total might include duplicates between Google Scholar
results and WoS/PubMed results. Based on a screening review of each article's title and abstract,
EPA retrieved 5 articles, 4 of which were published between 2013-2015 and appeared to
measure or estimate incidental water ingestion. EPA also retrieved one 2012 study that assessed
duration of non-swimming recreational water exposure by using novel time lapse photography
technology.
Google Search of Internet:
In addition, EPA conducted a Google search of the internet focused on specified URL
domains (listed in Table C-l) to identify candidate gray literature (e.g., state, federal, or
international government reports or guidance). The Google search string is presented below. The
Google search of the internet could not be limited by year or language.
Table C-l. Internet URL Domains Searched for Research Question 4
Organization
URL Domain
U.S. Government
•gov
.us
All U.S. States
Google Custom Search Engine
Centers for Disease Control and Prevention, including Agency
for Toxic Substances and Disease Registry
cdc.gov
Australia, including Australian Department of Health
gov.au
Canada, including Health Canada
gc.ca
European Union, including
• European Chemicals Agency
• European Commissions on Environment, Public Health,
Food, and Health and Consumers
europa.eu
Public Health England
hpa.org.uk
United Kingdom
gov.uk
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
C-6
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Organization
URL Domain
Germany
.de
Education websites
.edu
HERA (Human and Environmental Risk Assessment) Project
heraproject.com
World Health Organization
who.int
Results:
The Google search returned 390 results after removing duplicates. Based on a
preliminary screen of each result, EPA retrieved two documents which appeared to either derive
or cited an incidental ingestion rate while recreating which had not previously been identified
during the literature search process.
Google Search of Internet (conducted separately for each URL domain listed in Table C-l)
(pool OR swim OR swimmer OR swimmers OR swimming OR recreation OR recreational)
AND
(adolescents OR boys OR child OR children OR girls OR kids OR teenagers)
AND
("activity-related ingestion" OR "incidental ingestion" OR "activity-related ingestion" OR
"ingestion of water" OR "water ingestion")
AND
rate
AND
inurl:.
Filters: None
Date search was conducted: 10/9/2015
Dates searched: Not specified
Web browser: Internet Explorer
Research Question 5: What incidents of companion animal (e.g., dogs, horses) or livestock
poisonings, including mortality or adverse health effects, due to exposure to cyanotoxins in
freshwaters, marine water, or beaches have occurred in the past 15 years? Specifically,
when and where did these incidents occur, to which cyanotoxin were the animals exposed,
how were they exposed, and what were the weights and breeds of the affected animal(s)?
EPA searched PubMed, WoS, and Agricola to identify candidate journal article literature
relevant to companion animal or livestock poisoning due to exposures to cyanobacterial cells,
anatoxin-a, cylindrospermopsin, or microcystins. EPA first searched PubMed and WoS with a
focus on dogs. EPA conducted two additional searches in PubMed, WoS, and Agricola focused
on livestock, and on cats and birds. The search strings for each search iteration are presented
below.
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
C-l
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Results
The number of journal articles returned by the three searches is provided in Table C-2.
Based on a screening review of the article's title and abstract, EPA retrieved 5 of the 35 journal
articles retrieved during the search focused on dogs. These 5 articles appeared to provide
information about an incident of cyanotoxin exposure to an animal where the authors confirm
that the animal was exposed to a cyanotoxin by either measuring the concentration of cyanotoxin
found in the animal and/or by sampling the body of water to which the animal had contact.
Table C-2. Number of Journal Articles Returned by Three Search Strategies for Research
Question 5
Search Strategy Focus
Number of Results Returned from PubMed, WoS, and
Agricola Searches
Dogs
35a
Livestock
100
Cats and birds
169b
a Search conducted in PubMed and WoS only.
b Duplicates between PubMed/WoS results and Agricola results were not removed. Therefore, the cats and birds
search might include duplicates between Agricola results and PubMed/WoS results.
C.l Search strategy focused on dogs
PubMed Search:
("A lemmermannii Raphidiopsis mediterranean OR flos-aquae OR anatoxin-a OR
Aphanizomenon OR cylindrospermopsin OR "C. raciborskii" OR Cuspidothrix OR
Cylindrospermopsis OR Cylindrospermum OR "Cylindrospermopsis raciborskii" OR
Dolichospermum OR "M aeruginosa" OK Microcystis OR microcystin OR microcystins OR
Oscillatoria OR Planktothrix OR Phormidium OR Tychonema OR Woronichinia OR
Cyanobacteria OR cyanotoxin OR Cyanotoxins OR "harmful algae" OR "harmful algal bloom"
OR blue green algae)
AND
("health effect" OR "health endpoint" OR "health outcome" OR dead OR death OR deaths OR
died OR disease OR diseased OR diseases OR exposed OR exposure OR ill OR illness OR
illnesses OR infect OR infected OR infection OR infections OR morbidity OR mortality OR
poison OR poisoned OR poisoning OR poisonings OR sick OR sickness OR toxic OR toxicity
OR diagnose OR diagnosis OR diagnosed OR diagnoses)
AND
(canine OR canines OR dog OR dogs OR "Canis lupus familiaris" OR "Canis familiaris")
Filters: English
Date search was conducted: 10/5/2015
Publication dates searched: 1/1/2012-10/5/2015
Web of Science Search:
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
C-8
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("lemmermannii Raphidiopsis mediterranean OR flos-aquae OR anatoxin OR Aphanizomenon
OR cylindrospermopsin OR "C. raciborskii" OR Cuspidothrix OR Cylindrospermopsis OR
Cylindrospermum OR "Cylindrospermopsis raciborskii" OR Dolichospermum OR "M
aeruginosa" OK Microcystis OR microcystin OR microcystins OR Oscillatoria OR Planktothrix
OR Phormidium OR Tychonema OR Woronichinia OR Cyanobacteria OR cyanotoxin OR
Cyanotoxins OR "harmful algae" OR "harmful algal bloom" OR blue green algae)
AND
("health effect" OR "health endpoint" OR "health outcome" OR dead OR death OR deaths OR
died OR disease OR diseased OR diseases OR exposed OR exposure OR ill OR illness OR
illnesses OR infect OR infected OR infection OR infections OR morbidity OR mortality OR
poison OR poisoned OR poisoning OR poisonings OR sick OR sickness OR toxic OR toxicity
OR diagnose OR diagnosis OR diagnosed OR diagnoses)
AND
(canine OR canines OR dog OR dogs OR "Canis lupus familiaris" OR "Canis familiaris")
Filters: English
Date search was conducted: 10/5/2015
Publication dates searched: 1/1/2012-10/5/2015
C.2 Search strategy focused on livestock
PubMed and Agricola Searches:
("A. lemmermannii Raphidiopsis mediterranean OR flos-aquae OR anatoxin-a OR
Aphanizomenon OR cylindrospermopsin OR "C. raciborskii" OR Cuspidothrix OR
Cylindrospermopsis OR Cylindrospermum OR "Cylindrospermopsis raciborskii" OR
Dolichospermum OR "M aeruginosa" OK Microcystis OR microcystin OR microcystins OR
Oscillatoria OR Planktothrix OR Phormidium OR Tychonema OR Woronichinia OR
Cyanobacteria OR cyanotoxin OR Cyanotoxins OR "harmful algae" OR "harmful algal bloom"
OR blue green algae)
AND
("health effect" OR "health endpoint" OR "health outcome" OR dead OR death OR deaths OR
died OR disease OR diseased OR diseases OR exposed OR exposure OR ill OR illness OR
illnesses OR infect OR infected OR infection OR infections OR morbidity OR mortality OR
poison OR poisoned OR poisoning OR poisonings OR sick OR sickness OR toxic OR toxicity
OR diagnose OR diagnosis OR diagnosed OR diagnoses)
AND
(alpaca OR alpacas OR bronco OR broncos OR buffalo OR bull OR bulls OR cattle OR colt OR
colts OR cow OR cows OR bovine OR bison OR oxen OR donkey OR donkeys OR duck OR
ducks OR equine OR ewe OR ewes OR fillies OR filly OR foal OR foals OR gelding OR
geldings OR heifer OR heifers OR horse OR horses OR lamb OR lambs OR livestock OR llama
OR llamas OR mare OR mares OR mule OR mules OR mustang OR mustangs OR ponies OR
pony OR ram OR rams OR sheep OR stallion OR stallions OR steer OR pig OR pigs OR piglet
OR piglets)
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
C-9
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Filters: English
Date search was conducted: 11/25/2015
Publication dates searched: 1/1/2012-11/25/2015
Web of Science Search:
("lemmermannii Raphidiopsis mediterranean OR flos-aquae OR anatoxin OR Aphanizomenon
OR cylindrospermopsin OR "C. raciborskii" OR Cuspidothrix OR Cylindrospermopsis OR
Cylindrospermum OR "Cylindrospermopsis raciborskii" OR Dolichospermum OR "M
aeruginosa" OK Microcystis OR microcystin OR microcystins OR Oscillatoria OR Planktothrix
OR Phormidium OR Tychonema OR Woronichinia OR Cyanobacteria OR cyanotoxin OR
Cyanotoxins OR "harmful algae" OR "harmful algal bloom" OR blue green algae)
AND
("health effect" OR "health endpoint" OR "health outcome" OR dead OR death OR deaths OR
died OR disease OR diseased OR diseases OR exposed OR exposure OR ill OR illness OR
illnesses OR infect OR infected OR infection OR infections OR morbidity OR mortality OR
poison OR poisoned OR poisoning OR poisonings OR sick OR sickness OR toxic OR toxicity
OR diagnose OR diagnosis OR diagnosed OR diagnoses)
AND
(alpaca OR alpacas OR bronco OR broncos OR buffalo OR bull OR bulls OR cattle OR colt OR
colts OR cow OR cows OR bovine OR bison OR oxen OR donkey OR donkeys OR duck OR
ducks OR equine OR ewe OR ewes OR fillies OR filly OR foal OR foals OR gelding OR
geldings OR heifer OR heifers OR horse OR horses OR lamb OR lambs OR livestock OR llama
OR llamas OR mare OR mares OR mule OR mules OR mustang OR mustangs OR ponies OR
pony OR ram OR rams OR sheep OR stallion OR stallions OR steer OR pig OR pigs OR piglet
OR piglets)
Filters: English
Date search was conducted: 11/25/2015
Publication dates searched: 1/1/2012-11/25/2015
C.3 Search strategy focused on cats and birds
PubMed and Agricola Searches:
("A. lemmermannii Raphidiopsis mediterranean OR flos-aquae OR anatoxin-a OR
Aphanizomenon OR cylindrospermopsin OR "C. raciborskii" OR Cuspidothrix OR
Cylindrospermopsis OR Cylindrospermum OR "Cylindrospermopsis raciborskii" OR
Dolichospermum OR "M aeruginosa" OK Microcystis OR microcystin OR microcystins OR
Oscillatoria OR Planktothrix OR Phormidium OR Tychonema OR Woronichinia OR
Cyanobacteria OR cyanotoxin OR Cyanotoxins OR "harmful algae" OR "harmful algal bloom"
OR blue green algae)
AND
("health effect" OR "health endpoint" OR "health outcome" OR dead OR death OR deaths OR
died OR disease OR diseased OR diseases OR exposed OR exposure OR ill OR illness OR
illnesses OR infect OR infected OR infection OR infections OR morbidity OR mortality OR
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
C-10
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poison OR poisoned OR poisoning OR poisonings OR sick OR sickness OR toxic OR toxicity
OR diagnose OR diagnosis OR diagnosed OR diagnoses)
AND
(feline OR felines OR cat OR cats OR kitten OR kittens OR Catus" OR "Felis Catus" OR
bird OR birds OR avian OR waterfowl)
Filters: English
Date search was conducted: 2/1/2016
Publication dates searched: 1/1/2012-2/1/2016
Web of Science Search:
("lemmermannii Raphidiopsis mediterranean OR flos-aquae OR anatoxin OR Aphanizomenon
OR cylindrospermopsin OR "C. raciborskii" OR Cuspidothrix OR Cylindrospermopsis OR
Cylindrospermum OR "Cylindrospermopsis raciborskii" OR Dolichospermum OR "M
aeruginosa" OK Microcystis OR microcystin OR microcystins OR Oscillatoria OR Planktothrix
OR Phormidium OR Tychonema OR Woronichinia OR Cyanobacteria OR cyanotoxin OR
Cyanotoxins OR "harmful algae" OR "harmful algal bloom" OR blue green algae)
AND
("health effect" OR "health endpoint" OR "health outcome" OR dead OR death OR deaths OR
died OR disease OR diseased OR diseases OR exposed OR exposure OR ill OR illness OR
illnesses OR infect OR infected OR infection OR infections OR morbidity OR mortality OR
poison OR poisoned OR poisoning OR poisonings OR sick OR sickness OR toxic OR toxicity
OR diagnose OR diagnosis OR diagnosed OR diagnoses)
AND
(feline OR felines OR cat OR cats OR kitten OR kittens OR Catus" OR "Felis Catus" OR
bird OR birds OR avian OR waterfowl)
Filters: English
Date search was conducted: 2/1/2016
Publication dates searched: 1/1/2012-2/1/2016
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
C-ll
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Human Health Recreational Ambient Water Quality Criteria or C-12
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
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APPENDIX D. REVIEW OF THE STATE OF THE SCIENCE ON CYANOBACTERIAL
CELLS HEALTH EFFECTS
D.l Introduction
This appendix provides information gathered and reviewed to determine the state of the
science on health effects from cyanobacterial cells. EPA conducted literature searches to identify
studies relevant to the health effects from cyanobacterial cells. Detailed information on the
design and implementation of these searchers is provided in Appendix C. Results from these
literature searches were reviewed for relevance to cyanobacterial cell exposures and health
effects. This appendix builds on the cyanobacterial bloom information included in the main
document by discussing additional detail on the nature of cyanobacterial cells as stressors and, in
particular, the health effects associated with exposures to cyanobacterial cells.
D. 1.1 Animal Studies
Cyanobacterial cells cause allergenicity and irritation in animals, independent of whether
the cyanobacterial cells produce toxin. Three animal studies (Shirai et al. 1986; Stewart et al.
2006c; Torokne et al. 2001) demonstrated hypersensitivity reactions and dermal and eye
irritation in several species. Results from Torokne et al. (2001) indicated that hypersensitization
reactions do not correlate with microcystin content. Although the number of studies is limited
and different species were evaluated in each study, these studies provide evidence to support
hypersensitivity reactions in animals from exposure to cyanobacteria when cyanotoxins are not
present (Shirai et al. 1986; Torokne et al. 2001) and when they are (Stewart et al. 2006c).
Cyanobacteria bloom samples collected from five different lakes or ponds were tested for
allergenic and irritative effects in guinea pigs and rabbits, respectively (Torokne et al. 2001). The
microcystin content (presumed to be total LR, RR, and YR) ranged from not detected to
2.21 mg/g. To determine sensitization, guinea pigs were initiated with an intradermal injection of
freeze-dried cyanobacteria followed 7 days later by topical application at the injection site.
Sensitization was moderate to strong in 30-67 percent of guinea pigs and did not correlate with
microcystin content. The Aphanizomenon ovalisporum sample (a nontoxin-producing strain)
sensitized 91percent of the animals and was the strongest allergen. Skin irritation tests in albino
rabbits showed slight or negligible irritation, except for Aphanizomenon ovalisporum, which
showed moderate irritation. The eye irritation evaluation in rabbits was positive for four of the
five samples containing Microcystis.
Shirai et al. (1986) reported that C3H/HeJ mice, immunized i.p. with either sonicated or
live cells from a Microcystis water bloom, developed delayed-type hypersensitivity when
challenged 2 weeks later with a subcutaneous injection sonicated Microcystis cells. A positive
reaction, as assessed by footpad swelling, was seen in mice immunized with either live cells or
sonicated cells. Both toxic and nontoxic Microcystis cells induced delayed-type hypersensitivity
in this mouse study. Because this strain of mouse is unresponsive to lipopolysaccharide (LPS),
the footpad delayed-type hypersensitivity was not related to LPS, thus, the antigenic component
of the sonicated cyanobacterial cells is not known.
Stewart et al. (2006c) conducted a mouse ear swelling test in which cylindrospermopsin
and C. raciborskii solutions generated irritation of the abdominal skin exposed during induction
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
D-l
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(2 percent w/v lysed cell solution containing 73 [j,g/mL cylindrospermopsin). Subsequent dermal
exposures to the C. raciborskii solution produced hypersensitivity reactions (p = 0.001). The
cyanobacteria Microcystis aeruginosa and Anabaena circinalis elicited no responses in this test.
Two of the cyanobacterial cell studies in animals found that rodents became sensitized
after exposure and subsequent challenge to nontoxin strains (Shirai et al. 1986; Torokne et al.
2001). Torokne et al. (2001) found that a nontoxic strain was more sensitizing and irritating than
the toxic strains evaluated. These experiments support the conclusion that there is no relationship
between the cyanotoxin content and the allergenic effect of cyanobacteria.
D. 1.2 Clinical and Laboratory Human Studies
Several types of studies and reports provide information on associations between
cyanobacteria exposure and health effects. Clinical and in vitro studies (Bernstein et al. 2011;
Geh et al. 2015; Pilotto et al. 2004; Stewart et al. 2006a) have been able to assess associations
between cyanobacteria exposure and human health effects including dermal and allergenic
reactions. Three clinical studies assessed dermal exposure to cyanobacteri al cells using skin-
patch or skin-prick testing in humans (Bernstein et al. 2011; Pilotto et al. 2004; Stewart et al.
2006a). Some of the exposed individuals showed mild irritation or allergenicity. No statistically
significant dose-response relationships were found between skin irritation and increasing
cyanobacteri al cell concentrations. The allergenicity study suggests that cyanobacteria are
allergenic, particularly among people with chronic rhinitis (Bernstein et al. 2011).
Skin-patch testing in humans was performed by Pilotto et al. (2004) with laboratory-
grown cylindrospermopsin-producing C. raciborskii cells, both whole and lysed, which were
applied using adhesive patches at concentrations ranging from < 5,000 to 200,000 cells/mL to the
skin of 50 adult volunteers. After 24 hours, patches were removed and evaluation of the
erythematous reactions were graded. Analysis of participants' reactions to patches treated with
whole cells showed an OR of 2.13 and a 95% Confidence Interval (CI) of 1.79-4.21 (p < 0.001).
Lysed cells patch analysis showed an OR of 3.41 and a 95% CI of 2.00-5.84 (p < 0.001). No
statistically significant increase or dose-response between skin reactions and increasing cell
concentrations for either patches (whole or lysed) was observed. Subjects had skin reactions to
the cylindrospermopsin, and positive control patches more frequently than to the negative control
patches. The mean percentage of subjects with a reaction was 20% (95% CI: 15-31%). When
subjects reacting to negative controls (39) were excluded, the mean percentage was 11 percent
(95% CI: 6-18%>). Evaluation of erythematous reactions showed that mild irritations (grade 2)
were resolved in all cases within 24 to 72 hours.
Stewart et al. (2006a) conducted a skin-patch test with 39 volunteers (20 dermatology
outpatients; 19 controls) who were exposed to 6 cyanobacteri al suspensions, including toxigenic
species, nontoxigenic species, mixed suspensions, and two cyanobacteri al LPS extracts. All
cyanobacteri al suspensions of lyophilized cells were tested at three concentrations, 0.25 percent
w/v, 0.05 percent w/v, 0.005 percent w/v, and the estimated doses of cyanotoxins were 2.4 ng/kg
cylindrospermopsin and 2.6 ng/kg microcystins. Only one subject showed significant responses
to cyanobacteri al suspensions, specifically to two suspensions of cyanobacteri al cells: C.
raciborskii and mixed M. aeruginosa and C. raciborskii, both of which contained one or more
cyanotoxins. This subject showed no evidence of any dose-response effect in the dermal
reactions. None of the participants reacted to the cyanobacteri al LPS extracts, which ranged from
Human Health Recreational Ambient Water Quality Criteria or
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D-2
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260 ppb to 31 ppm. This small clinical study demonstrated that dermal hypersensitivity reactions
to cyanobacteria exposure occur infrequently, and further research into risk factors for
predisposition to this type reaction could be beneficial.
Bernstein et al. (2011) studied skin sensitization to non-toxic extracts of M aeruginosa in
259 patients with chronic rhinitis over 2 years. Patients were evaluated with aeroallergen skin
testing and skin-prick testing. The authors found that 86 percent of the subjects had positive skin
prick tests to Microcystis aeruginosa, and that patients with existing allergic rhinitis were more
likely to have reactions and sensitization to cyanobacteria than the controls (non-atopic health
subjects). This study indicated that cyanobacterial allergenicity is associated with the non-toxic
portion of the cyanobacteria.
Geh et al. (2015) studied the immunogenicity of extracts of toxic and non-toxic strains of
M aeruginosa in patient sera (18 patients with chronic rhinitis and 3 non-atopic healthy subjects
as documented in Bernstein et al. [2011]). Enzyme Linked Immunosorbent Assay (ELISA) test
was used to test IgE-specific reactivity, and gel electrophoresis, followed by immunoblot and
mass spectrometry, was done to identify the relevant sensitizing peptides. The authors found an
increase in specific IgE in those patients tested with the non-toxic Microcystis extract than the
extract from the toxic strain. After pre-incubation of the non-toxic extract with various
concentrations of microcystin, the authors found that phycocyanin and the core-membrane linker
peptide were responsible for the release of P-hexosaminidase in rat basophil leukemia cells. The
authors concluded that non-toxin-producing strains of cyanobacteria are more allergenic than
toxin-producing strains in allergic patients, and that the toxin may have an inhibitory effect on
the allergenicity of the cyanobacterial cells.
D.1.3 Epidemiological Studies and Case Reports
Among the epidemiological studies discussed here, some identified significant
associations between cyanobacteria exposure and a range of health outcomes including dermal,
eye/ear, GI, and respiratory effects. Several of these studies also measured one or more
cyanotoxins and found no association between cyanotoxin occurrence or exposure and health
effects. Additional evidence from outbreak and case reports provides support for health effects
associated with cyanobacteria exposure. Overall, these studies provide evidence of significant
associations between cyanobacterial cell exposure and human health effects even in the absence
of cyanotoxins. However, the reported associations between cyanobacterial cell densities and
health outcomes are not consistent. The studies vary in study design, methods used, size of study
population, cyanobacterial species evaluated, health effects identified, and cyanobacterial cell
densities associated with human health effects. Therefore, substantial uncertainty remains
regarding the associations between cyanobacterial cell exposure and human health effects.
Eight epidemiological studies evaluated short-term health effects associated with
recreational exposure to cyanobacterial blooms (El Saadi et al. 1995; Levesque et al. 2014; Lin et
al. 2015; Philipp 1992; Philipp & Bates 1992; Philipp et al. 1992; Pilotto et al. 1997; Stewart et
al. 2006d). See Table D-l for a summary list of these studies. The health outcomes evaluated
included dermal, GI, respiratory, and other acute effects, such as eye or ear symptoms. Seven
studies evaluated recreational exposure to freshwater cyanobacteria, and one evaluated exposure
to marine water cyanobacteria (Lin et al. 2015). Two studies included field sites in the
continental United States or Canada (Levesque et al. 2014; Stewart et al. 2006d), three occurred
Human Health Recreational Ambient Water Quality Criteria or
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D-3
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in the United Kingdom (Philipp 1992; Philipp & Bates 1992; Philipp et al. 1992), and three were
conducted in sub-tropical and tropical regions in Australia (El Saadi et al. 1995; Pilotto et al.
1997) and Puerto Rico (Lin et al. 2015). These epidemiological studies are discussed below in
chronological order.
Table D-l. Cyanobacteria Epidemiological Studies Summary
Lowest
Reference
Study
Design,
Cyanobacteria
Cvanotoxins
Health
Significant
n, and
Identified
Measured
Association"
Cyanobacterial
Cell Density
Location
(cells/mL)
Philipp
Cross-sectional
Microcystis sp.,
-
No statistically
No quantitative
(1992)
n = 246
Gleotrichia sp.
significant health
cyanobacterial
UK (Hampshire)
associations
cell densities
provided
Philipp
Cross-sectional
Microcystis sp.,
-
No statistically
No quantitative
and Bates
n = 382
Gleotrichia sp.
significant health
cyanobacterial
(1992)
UK (Somerset)
associations
cell densities
provided
Philipp et
Cross-sectional
Oscillatoria sp.,
-
No statistically
No quantitative
al. (1992)
n = 246
Aphanizomenon sp.,
significant health
cyanobacterial
UK (Lincolnshire,
Aphanothece sp.,
associations
cell densities
South Yorkshire)
Merismopedia sp.
provided
El Saadi et
Case-control
Anabaena sp.,
-
No statistically
No quantitative
al. (1995)
n cases =102 GI, 86
Aphanizomenon sp.,
significant health
cyanobacterial
dermatological
Planktothrix sp.,
associations
cell densities
n controls = 132
Anabaena circinalis,
provided
Australia (South
Microcystis aeruginosa
Australia)
Pilotto et
Cross-sectional
Microcystis aeruginosa,
Hepatotoxins
Significant positive
> 5,000
al. (1997)
n = 295 exposed
Microcystis sp.,
detected by mouse
association between
n = 43 unexposed
Anabaena sp.,
bioassay
combined symptoms
Australia (South
Aphanizomenon sp.,
(GI, dermal,
Australia, New South
Nodularia spumigena
respiratory, fever, eye
Wales, Victoria)
or ear irritation) and
cyanobacteria
Stewart et
Cohort (prospective)
Cyanobacteria
Microcystins
Significant positive
> 100,000b
al. (2006d)
n= 1,331
identified, species not
detected by high-
association between
Australia
specified
performance liquid
respiratory symptoms
(Queensland, New
chromatography
and cyanobacteria
South Wales) and
(HPLC) with
Significant positive
Florida
photodiode array
association between
detection or ELISA;
combined symptoms
cylindrospermopsin
(GI, dermal,
and anatoxin-a
respiratory, fever, eye
detected by HPLC-
or ear irritation) and
MS/MS; saxitoxins
cyanobacteria
not detected by
HPLC with
fluorescence
detection
Levesque
Cohort (prospective)
Cyanobacteria
Microcystins
Significant positive
20,000-100,000
et al.
n = 466
identified, species not
detected by ELISA
association between GI
(2014)
Canada (Quebec)
specified
symptoms with fever
and cyanobacteria
Human Health Recreational Ambient Water Quality Criteria or
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Reference
Lin et al.
(2015 )c
Study
Design,
n, and
Location
Cohort (prospective)
n= 15,726
Puerto Rico
(Boqueron)
Cyanobacteria
Identified
Cyanophyte filament,
Pseudanabaena sp.,
Picocyanophyte,
Synechococcus sp.,
Synechocystis sp.,
Cyanophyte cell pair,
Phormidium sp.,
Lyngbya sp.,
Trichodesmium sp.,
Aphanothece sp.,
Johannesbaptistia sp.,
Komvophoron sp.,
Cyanophyte colony,
Cyanophyte unicell
sphere
Cyanotoxins
Measured
Health
Association"
Lowest
Significant
Cyanobacterial
Cell Density
(cells/mL)
Lyngbyatoxin-a and Significant positive 36.7-237.4
debromo-
aplysiatoxin
measured but not
detected by
HPLC-MC
association between
respiratory illness and
cyanobacteria other
than picocyanobacteria
significant positive
association between
rash and cyanobacteria
other than
picocyanobacteria
>237.4
a Includes only significant associations between recreational cyanobacteria exposure and health effects.
t> Values were converted from cyanobacterial cell surface area (>12.0 mm2/mL) to cyanobacterial cell density (>100,000
cells/mL) using conversions in NHMRC (2008). Relationship between biomass and cyanobacterial cell density can vary by
species and cell size (Lawton et al. 1999; Stewart et al. 2006d).
cLin et al. (2015) evaluated picocyanobacteria and cyanobacteria other than picocyanobacteria separately.
Three cross-sectional studies were conducted by Philipp et al. (Philipp 1992; Philipp &
Bates 1992; Philipp et al. 1992) to evaluate health effects related to exposure to cyanobacteria
from recreational activities including sailing, windsurfing, and fishing in water bodies in the
United Kingdom. Questionnaires were administered to participants who visited one of six inland
lakes to evaluate exposure and morbidity (including dermal, eye/ear, GI, and respiratory
symptoms). Several species of cyanobacteria were identified and, in some cases, cyanobacterial
levels exceeded the National Rivers Authority threshold for "potential to cause harm." Only
minor morbidity was identified among recreators, and no statistically significant associations
between cyanobacteria exposure and morbidity were identified.
El Saadi et al. (1995) conducted a case-control study in Australia to evaluate exposure to
river water with detectable levels of cyanobacteria and GI and dermatological symptoms
evaluated by a medical practitioner. This river was used as a source for drinking water, domestic
water, and recreational water. The authors found no significant association between recreational
exposure to river water with cyanobacteria and GI or dermatological symptoms. Cyanotoxins
were not measured, but species of cyanobacteria were present that were capable of producing
cyanotoxins.
These four earlier studies (El Saadi et al. 1995; Philipp 1992; Philipp & Bates 1992;
Philipp et al. 1992) provided no quantitative data on cyanobacterial cell densities. Therefore,
they could not help inform determination of a quantitative level associated (or not associated)
with health effects.
Four more recent epidemiological studies assessed the association between exposure to
recreational waters containing cyanobacteria and human health and provide quantitative density
data for cyanobacterial cells (Levesque et al. 2014; Lin et al. 2015; Pilotto et al. 1997; Stewart et
al. 2006d). These studies reported at least one statistically significant association between
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exposure to cyanobacteria and human health outcomes., including GI illness (Levesque et al.
2014), respiratory symptoms (Lin et al. 2015; Stewart et al. 2006d), dermal symptoms (Lin et al.
2015), or combined symptomology (GI, dermal, respiratory, and other symptoms) (Pilotto et al.
1997; Stewart et al. 2006d). These associations were linked to a range of densities of
cyanobacterial cells from as low as > 5,000 cells/mL (Pilotto et al. 1997) to as high as 100,000
cells/mL (analogous to > 12 mm2/mL (NHMRC 2008; Stewart et al. 2006d). In contrast to the
studies that examined all cyanobacteria, Lin et al. (2015) evaluated picocyanobacteria, larger
cyanobacterial cells, and total phytoplankton, and reported health effects associated with 37-
1,461 cells/mL for cyanobacteria other than picocyanobacteria.
Pilotto et al. (1997) investigated the health effects from recreational exposures (including
jet-skiing, water-skiing, swimming, and windsurfing) to cyanobacteria in Australia. The study
included 852 participants, 777 who had water contact and were considered exposed, and 75 not
exposed. There were 338 recreators (295 exposed, 43 not exposed) after exclusion of those who
experienced symptoms or had recreational exposure in the 5 days prior to the initial interview at
the water recreation site (the after exclusion study group). Health outcomes evaluated included
diarrhea, vomiting, flu-like symptoms (e.g., cough), skin rashes, mouth ulcers, fevers, or eye or
ear infections. Water samples were collected for evaluation of cyanobacterial cell counts,
hepatotoxins, and neurotoxins.
In the after exclusion study group, when all symptoms were combined, the authors found
a significant trend of increasing symptom occurrence with duration of exposure at 7 days post-
exposure (p-value for trend =0.03). Similarly, in the after exclusion study group there was a
significant trend of increasing symptom occurrence with increasing cyanobacterial cell count (p-
value for trend = 0.04). To account for the combined effect of duration of exposure and
cyanobacterial cell density, unexposed participants were compared with those exposed for up to
60 minutes and for more than 60 minutes to water with up to 5,000 cells/mL and to water with
more than 5,000 cells/mL. For the after exclusion study group, a significant trend of increasing
symptom occurrence with increasing levels of exposure was identified (p-value for trend
=0.004). In addition, participants with recreational exposure for more than 60 minutes to
cyanobacterial densities above 5,000 cells/mL had a significantly higher symptom occurrence
rate at 7 days post-exposure than unexposed participants (OR = 3.44, CI: 1.09-10.82). In this
study, the significant trends observed in the after exclusion study group were not observed when
all participants were included.
Pilotto et al. (1997) reported toxicity data collected by the Australia Water Quality
Center. Presence or absence of particulate (intracellular) hepatotoxins in concentrated surface
water phytoplankton samples was measured by mouse bioassay. The authors reported that
hepatotoxins were identified at one site on two separate interview days and at three sites for one
day each. No evidence of neurotoxins was detected. They reported that no significant association
was found between the presence of hepatotoxins and symptom occurrence at two and seven days
after exposure. Data and analysis methods were not provided. The authors point out that trends
were observed at seven days and not at two days after exposure and this might suggest a delayed
rather than an immediate allergic response. The authors also stated they could not rule out other
causative factors, such as other microorganisms, that could co-occur with cyanobacteria. The
results from this study informed the recommendations made by WHO in Guidelines for Safe
Recreational Water Environments (WHO 2003).
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Stewart et al. (2006d) conducted a prospective cohort study to investigate the incidence
of acute symptoms in individuals exposed to cyanobacteria via recreational activities in lakes and
rivers in Australia and Florida. This study included 1311 recreators with any water contact-
related activity (e.g., swimming, boat entry/egress). Cyanobacterial cell densities were
characterized in terms of cell surface area rather than cell counts (to normalize for cell size
differences among different species). Authors evaluated incidence of acute symptoms in
recreators exposed to low, medium and high levels of cyanobacteria.
Study subjects were asked to complete a self-administered questionnaire before leaving
for the day after enrollment and to submit to a telephone follow-up interview. The questionnaire
and follow-up interview forms gathered information on various acute illnesses, their onset and
severity. Respiratory symptoms among study participants in the high recreational exposure group
(total cyanobacterial cell surface area >12 mm2/mL on day of recreation) were significantly
greater compared to participants in the low recreational exposure group (< 2.4 mm2/mL)
(adjusted OR = 2.1, 95% CI: 1.1-4.0). Respiratory symptoms were defined as difficulty
breathing, dry cough, productive cough, runny nose, unusual sneezing, sore throat, or wheezy
breathing. Reports of any symptom among study participants in the high exposure group were
significantly greater compared to reports among study participants in the low recreational
exposure group (adjusted OR = 1.7, 95% CI: 1.0-2.9). However, when subjects with recent prior
recreational water exposure were excluded the result remained positive but not significant
(adjusted OR = 1.6, 95% CI: 0.8-3.2). A dose-response relationship between increased
cyanobacterial biomass and increased symptom reporting was not identified. The authors
speculated that the pattern in their data could be due to a threshold effect. No other significant
associations with health effects were identified.
For water samples that contained potentially toxic cyanobacteria, Stewart et al. (2006d)
measured cyanotoxins including microcystins, saxitoxins, cylindrospermopsin and anatoxin-a by
HPLC or HPLC-MS/MS methods. Cyanotoxins were infrequently identified and only at low
levels. Microcystins were detected on two occasions (1 and 12 jag /L). Cylindrospermopsin was
found on seven occasions (ranging from 1-2 jag /L). Anatoxin-a was identified on a single
recruitment day at a concentration of 1 |ig/L. A statistically significant increase in symptom
reporting was found to be associated with anatoxin-a exposure, but the number of exposed
subjects was very low (n =18). No relationship between fecal indicator bacteria (fecal coliforms)
and symptoms was identified.
Levesque et al. (2014) conducted a prospective study of health effects including GI,
respiratory, dermal, eye/ear, and other symptoms associated with cyanobacteria and microcystin
exposure at three lakes in Canada (Quebec), one of which was a local supply of drinking water.
The study evaluated acute symptoms in humans (466 subjects included in analysis) living in
proximity to lakes affected by blooms and analyzed recreational exposure (full and limited
contact) and drinking-water exposure scenarios for both cyanobacterial cells and microcystins.
More severe GI symptoms, defined as diarrhea, vomiting, nausea and fever, or abdominal
cramps and fever, were associated with recreational contact (full and limited) and cyanobacteria.
For the more severe GI symptoms, the adjusted relative risk RR increased with cyanobacterial
cell counts providing evidence of a dose-response relationship (p-value for trend= 0.001,
< 20,000 cells/mL: RR = 1.52, 95% CI: 0.65-3.51; 20,000-100,000 cells/mL: RR = 2.71, 95%
CI: 1.02-7.16; > 100,000 cells/mL: RR = 3.28, 95% CI: 1.69-6.37). No evidence of a dose-
response relationship for cyanobacterial cell counts and the less severe GI symptoms was found.
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No relationship was observed between duration of contact or head immersion and risk of GI
symptoms. A significant increase for both the less and the more severe GI symptoms was found
with contact in the more highly impacted lakes (median cell densities 20,001-21,485 cells/mL),
but not in the less impacted lake (median 1,032 cells/mL). No relationship was observed between
microcystin concentrations and risk of GI symptoms. No significant associations between
recreational exposures to cyanobacteria and health effects other than GI effects were identified.
To evaluate possible co-exposures, authors measured microcystin concentrations and E.
coli as a fecal indicator. Levesque et al. (2014) measured particulate (intracellular) and dissolved
microcystins by ELISA and found that microcystins concentrations varied by lake and by sample
location (littoral vs. limnetic). Microcystin was detected in all three lakes. At Lake William the
median values were below the limit of detection at littoral and limnetic stations, with maximum
values of 0.63 [j,g/L and 0.02 [j,g/L respectively. At Lake Roxton littoral stations, the median
concentration was 0.23 [j,g/L (range: 0.008 [j,g/L-108.8 (J,g/L) and at limnetic stations the median
was 0.12 [j.g/L (range: 0.04 jag/L—1.12 (J,g/L). The Mallets Bay littoral stations had a median of
0.70 [j,g/L (range: under limit of detection - 773 (J,g/L) and the limnetic stations had a median of
0.35 [j,g/L (range: 0.001 [j.g/L-125 (J,g/L).
Levesque et al. (2014) reported that as a whole the microcystin concentrations during
contact were relatively low (1 st tertile: < 0.0012 (J,g/L; 2nd tertile: 0.0012-0.2456 (J,g/L; 3rd
tertile: > 0.2456 [ig/L). Symptoms were examined in relation to recreational and drinking water
exposure to cyanobacteria and microcystin. Only GI symptoms were associated with recreational
contact. The highest concentration of microcystin at which an episode of GI symptoms was
reported was 7.65 (J,g/L. There was no significant increase in adjusted relative risk of GI
symptoms with recreational exposure to more than 1 |ig/L microcystin. Adjusted relative risks
(adjusted for gender, gastrointestinal symptoms reported in the two weeks prior to data
collection, residence's source of drinking water) for GI illness without fever and GI illness with
fever were 1.06 (95% CI=0.32-3.52) and 1.48 (95% CI = 0.41-5.23), respectively. There were
significant increases in adjusted relative risk of several symptoms in participants who received
their drinking water from a source contaminated by cyanobacteria (muscle pain, GI illness, skin,
and ear symptoms).
Levesque et al. (2014) found that the geometric mean of E. coli at the three lakes ranged
from 0 to 145 CFU per 100 mL, and there was no association between GI illness and E.coli
levels. The authors noted that GI symptoms could have other causes, such as Aeromonas
infections; however, the symptoms were not related to fecal contamination as measured by
culturable E. coli. They also noted that people avoided full recreational contact during blooms
and more people engaged in limited contact recreation at higher cell counts. This observation
explains the counterintuitive finding that participants with limited contact exposure (fishing,
watercraft without direct water contact) had higher likelihood of symptom reporting compared to
participants with full contact.
A follow-up analysis (Levesque et al. 2016) characterized the same health data as
Levesque et al. (2014) to evaluate the relationship of bacterial endotoxin (lipopolysaccharides or
LPS) concentration to GI symptoms. Endotoxin concentrations were slightly correlated with
cyanobacterial counts (polychoric correlation coefficient = 0.57). The highest tertile of
endotoxin concentration (> 48 endotoxin units/mL) was significantly associated with GI illness
both with and without fever (GI illness without fever relative risk (RR) = 2.87, CI: 1.62-5.08; GI
illness with fever RR = 3.11, CI: 1.56-6.22). Adjustment to the level of cyanobacteria did not
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alter the relationship between endotoxin and GI illness and authors hypothesize that other gram
negative bacteria might play a role in the relationship between endotoxin levels and GI illness as
has been suggested in a previous study (Berg et al. 2011). Authors note that they stored filtered
water samples at -80 °C for several months prior to conducting endotoxin testing and that
another study (O'Toole et al. 2009) showed a 44 percent mean decline in the concentration of
endotoxins in samples stored at -80 °C for several weeks compared to samples stored at 4 °C for
24 hours. Levesque et al. (2016) caution that concentrations reported could be underestimated
and should be interpreted on an ordinal basis. Two other studies conducting endotoxin testing on
frozen samples found concentrations of a similar magnitude as this study (Berg et al. 2011;
Rapala et al. (2002).
Lin et al. (2015) conducted a prospective study based on data collected in 2009 at
Boqueron, Puerto Rico for 26 study days involving 15,726 enrollees to examine the association
between phytoplankton cell counts and illness among beachgoers. Three categories of
phytoplankton were evaluated: picocyanobacteria, cyanobacteria other than picocyanobacteria,
and total phytoplankton. The analysis compared people exposed at phytoplankton cell count
levels > 25th percentile (e.g., 25th to 75th percentile, > 75th percentile) to people exposed at
levels < 25th percentile (range of cyanobacteria other than picocyanobacteria: < 37-1461
cells/mL).
The study reported significant associations between recreational exposure to
cyanobacteria other than picocyanobacteria and respiratory symptoms, rash, and earache. For the
other symptoms measured, including eye irritation, no significant associations were observed.
More specifically, cyanobacterial (other than picocyanobacterial) densities of 37 to 237 cells/mL
(> 25th to < 75th percentile) and densities > 237 cells/mL (> 75th percentile) were associated
with increased respiratory symptoms (> 25th to < 75th percentile, odds ratio (OR) = 1.30, 95%
CI = 1.08-1.56; > 75th percentile, OR = 1.37, 95% CI = 1.12-1.67) in study participants who
reported body immersion. Respiratory symptom occurrence was defined as any two of the
following: sore throat, cough, runny nose, cold, or fever. Cyanobacterial (other than
picocyanobacterial) densities >237 cells/mL were associated with rash (OR = 1.32, 95%
CI = 1.05-1.66) and earache (OR = 1.75, 95% CI = 1.09-2.82). Study participants who reported
head submersion or swallowing of water showed no relationship between recreational exposures
to cyanobacteria (other than picocyanobacteria) and respiratory symptoms. There was no
association between recreational exposures to cyanobacteria (other than picocyanobacteria) and
respiratory symptoms in study participants who reported head submersion or swallowing of
water. A statistically significant association between cyanobacterial cell exposure (other than
picocyanobacterial cell exposure) and all health effects combined was also observed.
Lin et al. (2015) measured the dermatotoxins, debromoaplysiatoxin and lyngbyatoxin,
using high performance liquid chromatography-mass spectrometry and did not detect levels
above the limit of detection of 1.0 ppb. Authors reported that debromoaplysiatoxin and
lyngbyatoxin-a are photolabile and are unlikely to persist in the water column (Moikeha & Chu
1971). They noted that the health effects identified in this study were consistent with previous
blooms of Lyngbya majuscula, which can produce these toxins, though Lyngbya only comprised
3 percent of total planktonic cyanobacteria (other than picocyanobacteria). It is also possible that
the cyanobacterial cells could be having direct health effects as cyanotoxins levels were below
the limit of detection.
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To evaluate possible co-exposures, some studies measured cyanotoxins and fecal
indicators. Lin et al. (2015), Levesque et al. (2014), Pilotto et al. (1997), and Stewart et al.
(2006d) measured one or more cyanotoxins or total hepatotoxins. In some cases, cyanotoxin
levels were below the limit of detection. To determine if study participants possibly were
exposed to fecal contamination, three of the studies (Levesque et al. 2014; Lin et al. 2015;
Stewart et al. 2006d) measured bacterial fecal indicators at some study locations and times. Of
the studies that measured bacterial fecal indicators, none found an association between bacterial
fecal indicators and health effects. Of these studies, the only one with data available for viral
fecal indicators or concentrations of waterborne pathogens was Lin et al. (2015) provided in
Wade et al. (2010) and Soller et al. (2016).
In summary, although four studies identified significant associations between
cyanobacteria exposure and health effects, the type of health effect identified varied. One study
reported a significant association between GI illness and exposure to cyanobacteria (Levesque et
al. 2014). Stewart et al. (2006d) and Lin et al. (2015) identified statistically significant
associations between cyanobacterial cell exposure and respiratory effects. Lin et al. (2015) also
found a statistically significant association between earache and cyanobacterial densities (other
than picocyanobacteria). Both Pilotto et al. (1997) and Stewart et al. (2006d) found statistically
significant associations between cyanobacterial cell exposure and all symptoms combined. The
three cross-sectional studies conducted in the United Kingdom in 1990 found no statistically
significant associations, although some minor elevated morbidity was observed in exposed
individuals (Philipp 1992; Philipp & Bates 1992; Philipp et al. 1992). Another 1992 case-control
epidemiological study in Australia found no statistically significant symptoms for exposed
recreators (El Saadi et al. 1995).
The Centers for Disease Control and Prevention (CDC) has collected information on
illness outbreaks associated with HABs, which commonly involve cyanobacteria. This
information includes human health effects and water-sampling results voluntarily reported to the
Waterborne Disease Outbreak Surveillance System via the National Outbreak Reporting System
and the Harmful Algal Bloom Related Illness Surveillance System. CDC published summary
information on HAB-associated outbreaks from recreational exposures focusing on 2009-2010
with limited additional information available for outbreaks that occurred in 2001, 2004, and
2011-2012 (Dziuban et al. 2006; Hilborn et al. 2014; Hlavsa et al. 2014; Yoder et al. 2004).
CDC defines a recreational water-associated outbreak as the occurrence of similar illnesses in
two or more persons, epidemiologically linked by location and time of exposure to recreational
water or recreational water-associated chemicals volatilized into the air surrounding the water.
The 2009-2010 reporting cycle was notable, as almost half (46 percent) the recreational
water outbreaks reported to CDC were associated with HABs (Hilborn et al. 2014). Three of the
outbreaks confirmed the presence of cyanobacteria, and four confirmed the presence of
cyanotoxins. Gastrointestinal and dermatologic symptoms were the most commonly reported
symptom categories associated with HAB-related outbreaks in freshwater (Dziuban et al. 2006;
Hilborn et al. 2014; Hlavsa et al. 2014; Yoder et al. 2004). For the cyanobacteria-associated
outbreaks with reported symptom counts, the most common symptoms reported were GI related,
including vomiting, diarrhea, and nausea (estimated to be > 40 percent). The second most
frequent outbreak symptom reported was skin rash (> 27 percent cases reported). Fever, earache,
skin irritation, and headache were the next most frequently reported symptoms (11 percent, 9
percent, and 9 percent of cases reported, respectively).
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Hilborn et al. (2014) analyzed the HAB outbreak data from 2009-2010 and found
66 percent of case patients were individuals aged 1-19 years (n = 38 of 58 total) and 35 percent
were aged 9 years or younger (n = 20). In addition, in a cyanobacteria-associated outbreak in
2001, 42 children were affected. These data are limited and might be underreported, but they
suggest that children could be at increased risk for cyanobacteria-associated illness via
recreational exposure.
In addition to reports related to freshwater exposure, health effects including dermal,
eye/ear, and respiratory effects have been reported following exposure to marine cyanobacteria
and/or cyanotoxins including Lyngbya majuscula which can produce the cyanotoxins
lyngbyatoxin A and debromoaplysiatoxin (Osborne & Shaw 2008).
D.2 Mode of Action
Few mechanistic investigations have been completed on how exposure to cyanobacterial
cells might lead to inflammatory response. Torokne et al. (2001) evaluated the sensitization and
irritation potential of Microcystis, Anabaena, Cylindrospermopsis, and Aphanizomenon bloom
and strain samples and found no correlation between the cyanotoxin content and allergenicity.
For example, the nontoxic Aphanizomenon was the most allergenic sample, more allergenic than
the most toxic cyanobacterial cells they studied, Microcystis aeruginosa. Stewart et al. (2006e)
concluded that cutaneous effects strongly suggest allergic reactions, and symptoms such as
rhinitis, conjunctivitis, asthma, and urticaria (or hives) also indicate immediate hypersensitivity
responses, which are probably explained by a cascade action of pro-inflammatory cytokines.
Bernstein et al. (2011) suggested that the allergenic structure of cyanobacteria might be
associated with a nontoxin-producing part of the organism. Building on this conclusion, Geh et
al. (2015) conducted a series of experiments to identify the cyanobacteria allergen(s) responsible
for sensitization. Study participants were given skin-prick tests with extracts from nontoxic M.
aeruginosa strains. Serum from these individuals was collected from a subset of 15 patients who
elicited strong skin test responses toM aeruginosa and from 3 healthy control subjects. The
lysate from nontoxic M. aeruginosa strains was significantly (p < 0.01) more immunoreactive
than the lysate from the toxin-producing strains, which suggests that the nontoxic strain was
more allergenic than the toxic strain. They found, however, that IgE binds to M. aeruginosa
peptides present in lysates of both the toxic and nontoxic strains. Geh et al. (2015) also
performed a P-hexosaminidase release assay, as a surrogate assay for measuring histamine
release, to identify functional activity of theM aeruginosa extracts using rat basophil leukemia
cells. The authors concluded that the same allergen is present in toxic and nontoxic M.
aeruginosa lysates, but suggest the toxic M. aeruginosa lysate might contain an endogenous
inhibitor that prevents IgE from effectively binding to the specific allergen. The further analysis
by Geh et al. (2015) of the sera of individuals exposed to nontoxicM aeruginosa lysate
indicated that either linker core-membrane peptide or phycocyanin, or both, are potentially
responsible for M. aeruginosa allergenicity.
Epidemiological studies and case reports suggest respiratory effects that could be
consistent with an allergic or hay fever type reaction (Giannuzzi et al. 2011; Stewart et al.
2006e). Inhalation exposure to bacterial endotoxins (i.e., a toxin that is part of the cyanobacterial
cell as opposed to exotoxins such as microcystins and cylindrospermopsin) has been found to be
associated with pulmonary disease, including asthma, chronic obstructive airway disease, and
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emphysema (Stewart et al. 2006b). A recent review of the structure and effects of cyanobacterial
lipopolysaccharide suggested that it could act as an antagonist of the TLR4 receptor and inhibit
the inflammatory response pathway (Durai et al. 2015).
Stewart et al. (2006e) also noted that, although symptoms and time to onset can be
disparate, several reports described:
"a collective group of symptoms resembling immediate or Type-I hypersensitivity
reactions. Immediate hypersensitivity reactions are commonly associated with atopy,
which is the familial tendency to react to naturally occurring antigens, mostly proteins,
through an IgE-mediated process. Atopy frequently manifests as a spectrum of diseases,
e.g., seasonal rhinitis, conjunctivitis, asthma, and urticaria."
Documentation of this type of respiratory response is consistent with results from Geh et al.
(2015) and further supports that immune system response follows exposure to cyanobacteria.
In older literature, cyanobacterial lipopolysaccharide was suspected as being a cause of
inflammatory response because this cell structure, also found in many gram-negative bacterial
species, has been observed to initiate acute inflammatory responses in mammals that are typical
of a host reaction to tissue injury or infection (Stewart et al. 2006b). The Stewart et al. (2006e)
review, however, found evidence to support this mechanism lacking. Although all cyanobacteria
contain the pigment phycocyanin, not all species of cyanobacteria have shown dermal reactions.
Also, some species of cyanobacteria produce toxins that are known dermal irritants (e.g.,
lyngbyatoxin-a). Pilotto et al. (2004), however, found that 20-24 percent of the study
participants exposed to cyanobacterial cells via skin patches for 24 hours showed dermal
reactions to cyanobacteria species, both whole and lysed cells.
Stewart et al. (2006b) noted that the effects of microcystin- and cylindrospermopsin-
producing bacteria on the GI tract could suggest that cyanotoxins and lipopolysaccharide from
the cyanobacteria or other bacteria residing in the gut might cross a gut mucosal barrier that has
been disrupted and enhance the adverse effects of cyanotoxins.
An aquatic invertebrate study using brine shrimp (Artemia salina, Daphnia magna and
Daphnia galeata) to determine the toxicity of microcystin and cylindrospermopsin in
combination with cyanobacterial lipopolysaccharide found that pre-exposure to LPS increased
the lethal concentration (LCso) of cylindrospermopsin 8-fold (Lindsay et al. 2006). The authors
concluded that the decrease in susceptibility to cylindrospermopsin was due to the effects of
lipopolysaccharide on detoxification enzyme pathways; lipopolysaccharide decreased toxic
metabolites of cylindrospermopsin by suppressing the invertebrate cytochrome P450 system,
thus decreasing toxicity.
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D.3 References
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Hilborn ED, Roberts VA, Backer LC, DeConno E, Egan JS, Hyde JB, Nichohlas DC, Weigert EJ, Billing LM,
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associated disease outbreaks — United States, 2009-2010. Morbidity and Mortality Weekly Report
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& J. Bartram (Eds.), Toxic cyanobacteria in water -A guide to their public health consequences, monitoring
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Levesque B, Gervais MC, Chevalier P, GauvinD, Anassour-Laouan-Sidi E, Gingras S, FortinN, Brisson G, Greer
C, & Bird D (2014). Prospective study of acute health effects in relation to exposure to cyanobacteria. Sci
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Lin CJ, Wade TJ, Sams EA, Dufour AP, Chapman AD, & Hilborn ED (2015). A prospective study of marine
phytoplankton and reported illness among recreational beachgoers in Puerto Rico, 2009. Environ Health
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Lindsay J, Metcalf JS, & Codd GA (2006). Protection against the toxicity of microcystin-LR and
cylindrospermopsin in Artemia salina and Daphnia spp. by pre-treatment with cyanobacterial
lipopolysaccharide (LPS). Toxicon, 48(8), 995-1001. http://www.ncbi.nlm.nih.gov/pnbmed/16982077.
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Moikeha SN, & Chu GW (1971). Dermatitis-producing alga Lynbya majuscula gomont in Hawaii. II. Biological
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8817.1971. tbO .1.471.x.
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O'Toole J, Sinclair M, Jeavons T, & Leder K (2009). Influence of sample preservation on endotoxin measurement in
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Rapala J, Lahti K, Rasanen LA, Esala AL, Niemela SI, & Sivonen K (2002). Endotoxins associated with
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Soller JA, Eftim S, Wade TJ, Ichida AM, Clancy JL, Johnson TB, Schwab K, Ramirez-Toro G, Nappier S, &
Ravenscroft JE (2016). Use of quantitative microbial risk assessment to improve interpretation of a
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Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
D-14
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WHO (World Health Organization) (2003). Guidelines for Safe Recreational Water Environments: Volume 1:
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Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
D
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Human Health Recreational Ambient Water Quality Criteria or D-16
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
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APPENDIX E. INCIDENTAL INGESTION EXPOSURE FACTOR COMBINED
DISTRIBUTION ANALYSIS
EPA combined the distributions of incidental ingestion rate per hour and recreational
exposure duration to generate a hybrid distributions using R version 3.3.1. Table E-l presents the
parameters used to fit the distributions, and Table E-2 provides summary statistics for the
combined distributions. The R code follows these tables.
Table E-l. Parameters Used to Fit Distributions
Parameter
Ingestion rate
(L/hr)
Exposure
duration (hr/d)
Mean
0.0501
2.737
Standard deviation
0.0401
1.733
Minimum
0.417
Maximum
7.5
|i (In transformed)
-3.241
o2 (In transformed)
0.704
Minimum (In transformed)
-1010
Maximum (In transformed)
-1.59
Table E-2. Summary Statistics of Hybrid Distribution
Combined
Distribution
#
Ingestion
rate
(L/hr)*
Exposure
Duration
(hr/d)"
Summary Statistics for Ingestion (L/d)
Min
Q1
Median
Mean
Q3
Max
Percentile
at 0.33
L/d
Percentile
at 0.60
L/d
1
Normal
Normal
0.0000
0.0702
0.1424
0.1768
0.2465
1.2850
0.86
0.99
2
LN
Normal
0.0023
0.0564
0.1051
0.1448
0.1872
1.5340
0.91
0.99
3
LN
LN
0.0023
0.0479
0.0873
0.1241
0.1580
1.4730
0.94
0.99
4
LN
Gamma
0.0018
0.0467
0.0888
0.1281
0.1646
1.4580
0.93
0.99
5
Gamma
Gamma
0.0000
0.0396
0.0871
0.1307
0.1736
1.4020
0.92
0.99
* All input distributions were truncated to reflect observed minimum and maximum values
R Code
#Cyanotoxin recAWQC WA
#This script is to combine two distributions and generate histogram using five different
distribution combinations
rm(list=ls()); # Remove all current R objects from memory
library(truncnorm) #import library for truncated normal distribution
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
E-l
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# Convert exposure data from min/day to hr/day
mean_dur_min<-164.2 #mean exposure duration min/day
sd_dur_min<-103.97 #sd exposure duration min/day
med_dur_min<-140 #median exposure duration min/day
min_dur_min<-25 #minimum exposure duratrion min/day
max_dur_min<-450 #maximum exposure duration min/day
mean_dur<-mean_dur_min/60 #mean exposure duration hr/day
sd_dur<-sd_dur_min/60 #sd exposure duration hr/day
med_dur<-med_dur_min/60 #median exposure duration hr/day
min_dur<-min_dur_min/60 #minimum exposure duration hr/day
max_dur<-max_dur_min/60 #maximum exposure duration hr/day
#(1) Truncated normal ingestion and normal exposure duration distribution
mean_ing <- 0.05 #mean ingestion rate L/hr
sd_ing <- 0.04 #sd ingestion rate L/hr
min_ing<- 0 #minimum ingestion rate L/hr
max_ing<-0.205 #maximum ingestion rate L/hr
n = 100000 #number of samples
ingperhr_trunc<-rtruncnorm(n, a=min_ing, b=max_ing, mean ing, sd ing)
duration_hr_trunc<-rtruncnorm(n, a=min_dur, b=max_dur, mean dur, sd dur)
ingperday_trunc<-ingperhr_trunc*duration_hr_trunc
summary(ingperdaytrunc)
hist(ingperday_trunc,xlab="Ingestion rate (L/day)",ylab="Frequency", main ="Normal
distribution fit, truncated", xlim=c(0, 1.0), ylim=c(0, 400))
h=hi st(ingperdaytrunc)
h$density=h$counts/sum(h$counts)
plot(h,xlab="Ingestion rate (L/day)",ylab="Probability", main ="Truncated Normal
distribution fit", xlim=c(0, 1), ylim=c(0, 0.6), xaxp=c(0,1.5,15), freq=FALSE)
#Determine percentiles in combined normal distribution
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
E-2
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#(a) 97th ingestion rate =0.12 L/hr and mean exposure duration of 2.74 hr/day = 0.33
L/day
ecdf(ingperday_trunc)(0.33)
#(b) 97th ingestion rate =0.12 L/hr and 90th exposure duration of 5.0 hr/day = 0.60
L/day
ecdf(ingperday_trunc)(0.60)
#(2) Truncated Log-normal ingestion and normal exposure duration distribution
transform mean and std of ingestion rate
sd_ing_ln<-sqrt(log((sd_ing/mean_ing)A2+1))
mean_ing_ln<-log(mean_ing)-((sd_ing_lnA2)/2)
min_ing_ln<- -10A10
max_ing_ln<-log(max_ing)
ingperhr_ln_trunc<-exp(rtruncnorm(n, a=min_ing_ln, b=max_ing_ln,
mean=mean_ing_ln, sd=sd_ing_ln)) #truncated log normal distribution
duration_hr_trunc<-rtruncnorm(n, a=min_dur, b=max_dur, mean=mean_dur, sd=sd_dur)
#truncated normal distribution
ingperday_ln_trunc<-ingperhr_ln_trunc*duration_hr_trunc #combine distributions
summary(ingperday ln trunc) #summary statistics about the combined distribution
#Generate histogram
hist(ingperday_ln_trunc,xlab="Ingestion rate (L/day)",ylab="Probability", main
-'Truncated hybrid distribution fit", xlim=c(0, 2.0), ylim=c(0, 1))
h=hi st(ingperdaylntrunc)
h$density=h$counts/sum(h$counts)
plot(h,xlab="Ingestion rate (L/day)",ylab="Probability", main ="Truncated LN-Normal
hybrid distribution fit", xlim=c(0, 1), ylim=c(0, 0.6), xaxp=c(0,1.5,15), freq=FALSE)
#Generate empirical cumulative distribution function
plot(ecdf(ingperday_ln_trunc), main="")
#Determine percentiles in combined distribution
#(a) 97th ingestion rate =0.12 L/hr and mean exposure duration of 2.74 hr/day = 0.33
L/day
ecdf(ingperday_ln_trunc)(0.33)
#(b) 97th ingestion rate =0.12 L/hr and 90th exposure duration of 5.0 hr/day = 0.60
L/day
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
E-3
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ecdf(ingperday_ln_trunc)(0.60)
#(3) Truncated Log-normal ingestion and log-normal duration
sd_dur_ln<-sqrt(log((sd_dur/mean_dur)A2+1))
mean_dur_ln<-log(mean_dur)-((sd_dur_lnA2)/2)
min_dur_ln<-log(min_dur)
max_dur_ln<-log(max_dur)
ingperhr_ln_trunc<-exp(rtruncnorm(n=n, a=min_ing_ln, b=max_ing_ln,
mean=mean_ing_ln, sd=sd_ing_ln)) #truncated log normal distribution
duration_hr_ln_trunc<-exp(rtruncnorm(n=n, a=min_dur_ln, b=max_dur_ln,
mean=mean_dur_ln, sd=sd_dur_ln))
ingperday_ln2_trunc<-ingperhr_ln_trunc*duration_hr_ln_trunc #combine distributions
summary(ingperday_ln2_trunc) #summary statistics about the combined distribution
#Generate histogram
hist(ingperday_ln2_trunc,xlab="Ingestion rate (L/day)",ylab="Probability", main
-'Truncated hybrid distribution fit", xlim=c(0, 2.0), ylim=c(0, 1))
h=hi st(ingperday_ln2_trunc)
h$density=h$counts/sum(h$counts)
plot(h,xlab="Ingestion rate (L/day)",ylab="Probability", main ="Truncated log-normal
distribution fit", xlim=c(0, 1), ylim=c(0, 0.6), xaxp=c(0,1.5,15), freq=FALSE)
#Generate emperical cumulative distribution function
plot(ecdf(ingperday_ln2_trunc), main-'")
#Determine percentiles in combined distribution
#(a) 97th ingestion rate =0.12 L/hr and mean exposure duration of 2.74 hr/day = 0.33
L/day
ecdf(ingperday_ln2_trunc)(0.33)
#(b) 97th ingestion rate =0.12 L/hr and 90th exposure duration of 5.0 hr/day = 0.60
L/day
ecdf(ingperday_ln2_trunc)(0.60)
# (4) Truncated log-normal ingestion distribution and gamma duration distribution (beta
distribution for duration added by arun)
vr_dur<- sd_durA2 # variance of the duration distribution
theta_dur<-vr_dur/mean_dur # scale parameter of the gamma distribution
k_dur<-mean_dur/theta_dur # shape parameter of the gamma distribution
rgamma_trunc<-function(n,k,theta,min,max){
Human Health Recreational Ambient Water Quality Criteria or E-4
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
-------
i<-l
gv<-matrix(,n,l)
while(i<=n) {
a<-rgamma( 1, shape=k, scale=theta)
if (a>min & a
-------
duration_hr_gm<-rgamma_trunc(n,k_dur,theta_dur,min_dur,max_dur) #truncated
gamma distribution
ingperhr_gm<-rgamma_trunc(n,k_ing,theta_ing,min_ing,max_ing) #truncated gamma
distribution
ingperday_gm<-ingperhr_gm*duration_hr_gm #combine In and gm distributions
summary(ingperday gm) #summary statistics about the combined distribution
#Generate histogram
hist(ingperday_gm,xlab="Ingestion rate (L/day)",ylab="Probability", main ="Truncated
hybrid distribution fit", xlim=c(0, 2.0), ylim=c(0, 1))
h=hi st(ingperday_gm)
h$density=h$counts/sum(h$counts)
plot(h,xlab="Ingestion rate (L/day)",ylab="Probability", main ="Truncated Gamma
distribution fit", xlim=c(0, 1), ylim=c(0, 0.6), xaxp=c(0,1.5,15), freq=FALSE)
#Generate emperical cumulative distribution function
plot(ecdf(ingperday_gm), main="")
#Determine percentiles in combined distribution
#(a) 97th ingestion rate =0.12 L/hr and mean exposure duration of 2.74 hr/day = 0.33
L/day
ecdf(ingperday_gm)(0.33)
#(b) 97th ingestion rate =0.12 L/hr and 90th exposure duration of 5.0 hr/day = 0.60
L/day
ecdf(ingperday_gm)(0.60)
Human Health Recreational Ambient Water Quality Criteria or E-6
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
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APPENDIX F. INFORMATION ON CELLULAR CYANOTOXIN AMOUNTS AND
CONVERSION FACTORS
The information in the tables below was generated from a brief survey of the peer-reviewed and
published scientific literature. This survey was not a formal systematic literature search and was
conducted to evaluate the availability of data needed to calculate a cyanobacterial cell density
potentially associated with a specific cyanotoxin concentration.
Human Health Recreational Ambient Water Quality Criteria or F-l
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
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Table F-l. Cell Quotas for Cyanotoxins Available from a Spot Check of the Literature
Toxin
Species
Site/Clone
Toxin Quota
Reference
Notes
Cylindrospermopsin
Cylindrospermopsis
raciborskii
16 sites in 3
reservoirs in
Queensland,
Australia
4.5-55.8 fga cell1;
10.0 - 49.4 fg cell1
Orretal. (2010)
has values for
cylindrospermopsin and
d-cylindrospermopsin as
cell versus
cylindrospermopsin
concentration in table
Cylindrospermopsis
raciborskii
New South Wales,
Australia
31 (12-52) fg cell"1
Hawkins et al. (2001)
also has biomass
conversions (Table 2)
Cylindrospermopsis
raciborskii
Saudi Arabia lake
0.6 - 14.6 pgb cell"1
Mohamed and Al-
Shehri (2013)
Cylindrospermopsis
raciborskii
Queensland,
Australia
13.4 (± 2.6) - 14.9 (± 3.4) fg cell1
Davis et al. (2014)
range is for two strains
Cylindrospermopsis
raciborskii
12.1 (5.6) - 24.7 (9.5) ng° 10~6 cells;
3.2 (0.67) - 5.7 (1.4) ng 10"6 cells;
0.049 (0.002)-0.094 (0.001)
cylindrospermopsin chlorophyll a'1;
0.016 (0.005)-0.11 (0.003)
cylindrospermopsin chlorophyll a'1
Carneiro et al. (2013)
Cylindrospermopsis
raciborskii
Queensland,
Australia
0.28 x 10"2 (0.2 x 10"2) - 1.8 x 10"2
(0.4 x 10"2) pg cell"1
Willis et al. (2015)
Cylindrospermopsis
raciborskii
Queensland,
Australia
19 (3)-26 (4) fg cell"';
416 (67) - 447 (69) 103 fg fim"3
Pierangelini et al.
(2015)
breaks out
cylindrospermopsin and
d-cylindrospermopsin
Microcystin (MC)
Planktothrix agardhii
Paris, France lake
1.5 - 19 fg MC-LR cell"1
Briand et al. (2008)
Planktothrix agardhii
England; Turkey
0.7 - 1.9 fg (j.m 3;
75.6-91.2 fg cell"1
Akcaalan et al. (2006)
Planktothrix rubescens England; Turkey
1.4 - 2.9 fg (j.m"3;
103.9-235.6 fg cell1
Akcaalan et al. (2006)
Planktothrix rubescens
Italy lake
1.0 - 3.9 (j.g mm"3;
Salmaso et al. (2014)
Planktothrix rubescens
France lake
0.13 (0.16) - 0.16 (0.27) pg cell"1
Briand et al. (2008)
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
F-2
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Toxin Species Site/Clone Toxin Quota Reference Notes
Microcystin
(continued)
Model Cyanobacteria
91.5 fg cell-1
Jahnichen et al. (2001)
Cites Long et al. (2001),
Orr and Jones (1998),
Jahnichen et al. (2001),
and Watanabe et al.
(1989) for quotas
Microcystis
aeruginosa
MASH01-A19
50 - 170 fg cell1
Orr and Jones (1998)
estimated from Figure 5
Microcystis
aeruginosa
Lake Huron,
United States
140 fg cell"1
Fahnenstiel et al. (2008)
Microcystis
aeruginosa
Portugal lake
0.06 - 0.22 pg cell"1
Vasconcelos et al.
(2011)
Microcystis
aeruginosa
18 (0.95) - 23.7 (0.96) fg cell1
Jahnichen et al. (2007)
Microcystis
aeruginosa
PCC 7806
34.5-81.4 fg cell"1
Wiedner et al. (2003)
Microcystis
aeruginosa
France
0.05 - 3.8 pg cell"1
Sabart et al. (2010)
estimated from Figure 3
Microcystis
aeruginosa
New Zealand
0.1 - 1.55 pg cell"1
Wood et al. (2012)
estimated from Figure 1
" fg = femtogram
b pg = picogram
0 ng = nanogram
Human Health Recreational Ambient Water Quality Criteria or F-3
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
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Table F-2. A Brief Summary of Cell Concentration - Cyanotoxin Conversions Available from a Spot Check of the Literature
Toxin Species Site/Clone Conversion Reference Notes
Cylindrospermopsin Cylindrospermopsis 16 sites in 3 has cell versus cylindrospermopsin Orretal. (2010) additional data available
raciborskii reservoirs in concentration in table but need to be digitized
Queensland,
Australia
Cylindrospermopsis New South Wales, 0.13% (0.06 - 0.35%) diy weight; Hawkins et al. (2001)
raciborskii Australia 0.57 (0.18 - 1.52 %) fg (im3biovolume
Cylindrospermopsis Queensland, l%w/w (10 mg cylindrospermopsin per g Eaglesham et al. has cylindrospermopsin
raciborskii Australia of diy weight) to 1 (ig/g of dry weight (1999) versus Trichomes mg"1
(x 10"3) conversion in
Figure 4; data need to be
digitized
Microcystin (MC) Planktothrix agardhii England; Turkey 29.4 (2.3) - 34.9 (3.7) pg filament-1; Akcaalan et al. (2006)
0.2 (0.06) -1.1 (0.6) fg |im 'biovolume
Planktothrix agardhii
55 German lakes
1,500 - 2,200 (j.g g_1 dry weight;
0.25 - 0.5 (j.g MC (j.g chlorophyll a'1
Fastner et al. (1999)
derived from Figure 2
Planktothrix
rubescens
England; Turkey
28.2 (7.1) - 53.6 (20.6) pg filament"1;
0.9 - 3.4 (1) fg (j.m3
Akcaalan et al. (2006)
Planktothrix
rubescens
55 German lakes
1,600 - 4,000 (j.g g"1 diy weight;
0.22 - 0.5 (j.g MC (j.g chlorophyll a'1
Fastner et al. (1999)
derived from Figure 2
Planktothrix
rubescens
Italy lake
Salmaso et al. (2014)
has regression formulas
for both MC versus
chlorophyll and MC
versus biovolume
Planktothrix spp.
0.38 - 6.01 jig mg dry weight"1
0.17-4.5 (j.g mm ' biovolume
Kurmayer et al.
(2016)
low and high MC
producing strains of P.
rubescens and P. agardhii
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
F-4
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Toxin Species Site/Clone Conversion Reference Notes
Microcystin Planktothrix spp. UTEX2388 87.9 (5.2) - 339 (igMC-LR g"1 dry Oh et al. (2000) has values for MC-LR
(continued) weight; and MC-RR
0.56 (0.03) - 2.47 (0.03) mg MC-LR g"1
protein;
467 (8.1) - 773.5 g MC-RR g"1 dry
weight;
3.00 (0.05) - 5.63 (0.03) mg MC-LR g"1
protein
Microcystis
aeruginosa
Portugal lake
Vasconcelos et al.
(2011)
has cell versus MC
concentration in Figure 4
Microcystis
aeruginosa
UTEX 2388
y = 0.661 x -38.9 (r = 0.569);
y: (MC |ig g1), x: (chlorophyll a |ig L1)
Lee et al. (2000)
Microcystis
aeruginosa
France lake
Sabart et al. (2010)
has MC versus cell
concentrations in Figures
2 and 3
Microcystis
aeruginosa
Lake Biwam,
Japan
Ozawa et al. (2005)
has MC versus cell
concentration,
chlorophyll a in Figure 2
Microcystis
aeruginosa
MASH01-A19
1.2 - 9.3 mg g"1 dry weight
Orr and Jones (1998)
estimated from Figure 4
Microcystis spp.
San Francisco
estuary
0 - 1 (j.g g_1 dry weight
Lehman et al. (2008)
derived from Figure 6
Microcystis spp.
55 German lakes
0 - 1000 (j.g g"1 diy weight
0.08 - 0.31 (j.g MC (j.g chlorophyll aA
Fastner et al. (1999)
derived from Figure 2;
derived from Figure 7
Microcystis spp.
Lake Suwa, Japan
1.20 - 136 (j.g MC-RR 100 mg-1 dry
weight;
4 - 89.8 jig MC-RR 100 mg"1 dry
Park et al. (1998)
estimated from Figure 7
Unknown
Quebec, Canada
weight;
Giani et al. (2005)
has biomass (g C L1)
versus MC (mg g"1) in
Figure 4
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
F-5
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Appendix F References
Akcaalan R, Young FM, Metcalf JS, Morrison LF, Albay M, & Codd GA (2006). Microcystin analysis in single
filaments of Planktothrix spp. in laboratory cultures and environmental blooms. Water Research, 40(8),
1583-1590.
Briand E, Gugger M, Francois J-C, Bernard C, Humbert J-F, & Quiblier C (2008). Temporal variations in the
dynamics of potentially microcystin-producing strains in a bloom-forming Planktothrix agardhii
(cyanobacterium) population. Applied and Environmental Microbiology, 74(12), 3839-3848.
Carneiro RL, Pereira Ribeiro da Silva A, & Freitas de Magalhaes V (2013). Use of the cell quota and chlorophyll
content for normalization of cylindropermopsin produced by two Cylindrospermopsis raciborskii strains
grown under different light intensities. Ecotoxicology and Environmental Contamination, 8(1), 93-100.
Davis TW, Orr PT, Boyer GL, Burford MA (2014). Investigating the production and release of cylindrospermopsin
and deoxy-cylindrospermopsin by Cylindrospermopsis raciborskii over a natural growth cycle. Harmful
Algae, 31, 18-25.
Eaglesham GK, Norris RL, Shaw GR, Smith MJ, Chiswell RK, Davis BC, Neville GR, Seawright AA, & Moore
MR (1999). Use of HPLC-MS/MS to monitor cylindrospermopsin, a blue-green algal toxin, for public
health purposes. Environmental Toxicology, 14(1), 151-154.
Fahnenstiel GL, Millie DF, Dyble J, Litaker RW, Tester PA, McCormick MJ, Rediske R, & Klarer D (2008).
Microcystin concentrations and cell quotas in Saginaw Bay, Lake Huron. Aquatic Ecosystem Health &
Management, 11(2), 190-195.
Fastner J, Neumann U, WirsingB, Weckesser J, Wiedner C, Nixdorf B, Chorus I (1999). Microcystins (hepatotoxic
heptapeptides) in German fresh water bodies. Environmental Toxicology, 14(1), 13-22.
Giani A, Bird DF, Prairie YT, & Lawrence JF (2005). Empirical study of cyanobacterial toxicity along a trophic
gradient of lakes. Canadian Journal of Fisheries and Aquatic Sciences, 62(9), 2100-2109.
Hawkins PR, Putt E, Falconer I, Humpage A (2001). Phenotypical variation in a toxic strain of the phytoplankter,
Cylindrospermopsis raciborskii (Nostocales, Cyanophyceae) during batch culture. Environ Toxicol, 16,
460-467.
Jahnichen S, Ihle T, Petzoldt T, & Benndorf J (2007). Impact of inorganic carbon availability on microcystin
production by Microcystis aeruginosa PCC 7806. Applied and Environmental Microbiology, 73(21), 6994-
7002.
Jahnichen S, Petzoldt T, & Benndorf J (2001). Evidence for control of microcystin dynamics in Bautzen Reservoir
(Germany) by cyanobacterial population growth rates and dissolved inorganic carbon. Arch Hydrobiol,
150(2), 177-196.
Kurmayer R, Deng L, & Entfellner E (2016). Role of toxic and bioactive secondary metabolites in colonization and
bloom formation by filamentous cvanobacteria Planktothrix. Harmful Algae, 54, 69-86.
Lee SJ, Jang M-H, Kim H-S, Yoon B-D, & Oh HM (2000). Variation of microcystin content of Microcystis
aeruginosa relative to medium N: P ratio and growth stage. Journal of Applied Microbiology, 89(2), 323-
329.
Lehman PW, Boyer G, Satchwell M, Waller S (2008). The influence of environmental conditions on the seasonal
variation of Microcystis cell density and microcystins concentration in San Francisco Estuary.
Hydrobiologia, 600(1), 187-204.
Long BM, Jones GJ, & Orr PT (2001). Cellular microcystin content in N-limited Microcystis aeruginosa can be
predicted from growth rate. Appl Environ Microbiol, 67(1), 278-283.
Mohamed ZA, Al-Shehri AM (2013). Assessment of cylindrospermopsin toxin in an arid Saudi lake containing
dense cyanobacterial bloom. Environmental Monitoring and Assessment, 185(3), 2157-2166.
Oh H-M, June Lee S, Jang M-H, & Dae Yoon B (2000). Microcystin production by Microcystis aeruginosa in a
phosphorus-limited chemostat. Applied and Environmental Microbiology, 66(1), 176-179.
Orr PT, Jones GJ (1998). Relationship between microcystin production and cell division rates in nitrogen-limited
Microcystis aeruginosa cultures. Limnol Oceanogr, 43(1), 1604-1614.
Orr PT, Rasmussen JP, Burford MA, Eaglesham GK, Lennox SM (2010). Evaluation of quantitative real-time PCR
to characterise spatial and temporal variations in cyanobacteria, Cylindrospermopsis raciborskii
(Woloszynska) Seenaya et Subba Raju and cylindrospermopsin concentrations in three subtropical
Australian reservoirs. Harmful Algae, 9, 243-254.
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
F-6
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Ozawa K, Fujioka H, Muranaka M, Yokoyama A, Katagami Y, Homma T, Ishikawa K, Tsujimura S, Kumagai M,
Watanabe MF, & Park HD (2005). Spatial distribution and temporal variation of Microcystis species
composition and microcystin concentration in Lake Biwa. Environmental Toxicology, 20(3), 270-276.
Park HD, Iwami C, Watanabe M-F, Harada K-I, Okino T, & Hayashi H (1998). Temporal variabilities of the
concentrations of intra-and extracellular microcystin and toxic Microcystis species in a hypertrophic lake,
Lake Suwa, Japan (1991-1994). Environmental Toxicology and Water Quality, 13( 1), 61-72.
Pierangelini M, Sinha R, Willis A, Burford MA, Orr PT, Beardall J, & NeilanBA (2015). Constitutive
cylindrospermopsin pool size in Cylindrospermopsis raciborskii under different light and CO2 partial
pressure conditions. Applied and Environmental Microbiology, 81(9), 3069-3076.
Sabart M, Pobel D, Briand E, Combourieu B, Salcncon MJ, Humbert J-F, Latour D (2010). Spatiotemporal
variations in microcystin concentrations and in the proportions of microcystin-producing cells in several
Microcystis aeruginosa populations. Applied and Environmental Microbiology, 76(14), 4750-4759.
Salmaso N, Copetti D, Cerasino L, Shams S, Capelli C, Boscaini A, Valsecchi L, Pozzoni F, & Guzzella L (2014).
Variability of microcystin cell quota in metapopulations of Planktothrix rubescens: Causes and
implications for water management. Toxicon, 90, 82-96.
Vasconcelos V, Morais J, & Vale M (2011). Microcystins and cyanobacteria trends in a 14 year monitoring of a
temperate eutrophic reservoir (Aguieira, Portugal). Journal of Environmental Monitoring, 13(3), 668-672.
Watanabe MF, Harada KI, Matsuura K, Watanabe M, Suzuki M (1989). Heptapeptide toxin production during the
batch culture of two Microcystis species (cyanobacteria/ JAppl Phycol, 1, 161-165.
Wiedner C, Visser PM, Fastner J, Metcalf JS, Codd GA, & Mur LR (2003). Effects of light on the microcystin
content of Microcystis strain PCC 7806. Applied and Environmental Microbiology, 69(3), 1475-1481.
Willis A, Adams MP, Chuang AW, Orr PT, O'Brien KR, Burford MA (2015). Constitutive toxin production under
various nitrogen and phosphorus regimes of three ecotypes of Cylindrospermopsis raciborskii
((Woloszynska) Seenayya et SubbaRaju). Harmful Algae, 47, 27-34.
Wood SA, Dietrich DR, Cary SC, & Hamilton DP (2012). Increasing Microcystis cell density enhances microcystin
synthesis: A mesocosm study. Inland Waters, 2(1), 17-22.
Human Health Recreational Ambient Water Quality Criteria or F-7
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
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Human Health Recreational Ambient Water Quality Criteria or F-8
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
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APPENDIX G. TABLES OF STATE-ISSUED GUIDELINE SPECIFIC TO ANIMAL
CYANOTOXIN POISONING
G.l California
Table G-l. California Environmental Protection Agency (2012) Action levels for Selected
Pet and Livestock Scenarios
Microcystins"
Cylindrospermopsin
Media (units)
Subchronic water intake, dog1'
2
10
water (ng/L)
Subchronic crust and mat intake,
dog
0.01
0.04
crusts and mats (mg/kg dw)°
Acute water intake, dog1'
100
200
water (|ig/L)
Acute crust and mat intake, dog
0.5
0.5
crusts and mats (mg/kg dw)°
Subchronic water intake, cattle"
0.9
5
water (|ig/L)
Subchronic crust and mat intake,
cattle"
0.1
0.4
crusts and mats (mg/kg dw)°
Acute water intake, cattle"
50
60
water (|ig/L)
Acute crust and mat intake, cattle"
5
5
crusts and mats (mg/kg dw)°
aMicrocystins LA, LR, RR, and YR all had the same RfD so the action levels are the same.
b Subchronic refers to exposures over multiple days.
0 Based on sample dry weight (dw).
d Acute refers to exposures in a single day.
e Based on small breed dairy cows because their potential exposure to cyanotoxins is greatest.
Table G-2. California Environmental Protection Agency (2012) Reference Doses
and Acute and Subchronic Action Levels for Canine Exposure to Cyanotoxins in
Drinking Water
Microcystin
Cylindrospermopsin
Water consumption L/kg-d
0.085
0.085
Uncertainly factor (unitless)
3
3
Acute RID'1 mg/kg/d
0.037
0.04
Acute action level [ig/L
100
200
Subchronic RfD mg/kg/d
0.00064
0.0033
Subchronic action level |ig/L
2
10
Reference:
Butler N, Carlisle J, Kaley KB, & Linville R (2012). Toxicological Summary and Suggested Action Levels to
Reduce Potential Adverse Health Effects of Six Cyanotoxins.
http://www.waterboards.ca.gov/water issues/programs/peer review/docs/calif cvanotoxins/cvanotoxins05
pdf. Last Accessed: 08/03/2016.
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
G-l
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G.2 Oregon
Table G-3. Oregon Dog-specific Guideline Values for Cyanotoxins in
Recreational Waters (jig/L)
Microcystin
Cylindrospermopsin
Dog Guidance Value
0.2
0.4
Note: All dog-specific guideline values have been changed in this revision because California
EPA's estimate of the amount of water an exercising dog consumes per kilogram body weight
was updated in 2012 (from 0.168 to 0.255 L/kg-day). Current dog-specific guideline values are
now consistent with the California EPA update. The dog-specific value for saxitoxins was
further modified by application of an uncertainty factor to the dog-specific TDI for interspecies
differences in sensitivity between humans (the species in the critical study) and dogs.
Reference:
Oregon Health Authority (2016). Oregon Harmful Algae Bloom Surveillance (HABS) Program Public Health
Advisory Guidelines: Harmful Algae Blooms in Freshwater Bodies.
https://pnblic.health.oregon.gOv/.HealthvEnviron.ments/Recreation/.H.armfiilAlgaeBlooms/.Docnmenfs
PublicHeaithAdvisorvGuideiines.pdf.
G.3 Grayson County, Texas
Table G-4. Grayson County Texas Microcystin Guidelines for Dogs
Quantity of Lake Water Ingested to Receive a Potentially Lethal Dose of Microcystin, Assuming
that Mouse and Dog Toxic Responses are Equivalent
10 pound dog
80 pound dog
Quantity of Lake Water Ingested to Receive a Potentially Lethal Dose of Microcystin, Assuming
that Mouse and Dog Toxic Responses are Equivalent (at actual concentrations found in Grand
Lake, Oklahoma, in June 2011)
Highest measured concentration of Microcystin was 358 ppb.
10 pound dog
80 pound dog
*This is not including additional dose amounts that could be ingested from a dog self grooming
algae scum off its fur.
**LD50 for Microcystin- mouse used in Calculations = 45 mcg/kg
***20 ppb Microcystin is algal toxin threshold for BGA Warning (condition red)
Gallons of Water Pounds of Water
2.70
22.50
21.57
180.00
Gallons of Water Pounds of Water
0.15 (19.3 ounces)
1.26
1.21
10.06
Human Health Recreational Ambient Water Quality Criteria or
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
G-2
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Quantity of Lake Water Ingested to Receive a Potentially Lethal Dose of Cylindrospermopsin,
Assuming that Mouse and Dog Toxic Responses are Equivalent
20 ppb Cylindrospermopsin in Lake Water
10 pound dog
80 pound dog
*This is not including additional dose amounts that could be ingested from a dog self grooming
algae scum off its fur.
**LD50 for Cylindrospermopsin- mouse used in Calculations = 4400 mcg/kg
***20 ppb Cylindrospermopsin is algal toxin threshold for BGA Warning (condition red)
Reference:
Lillis J, Ortez A, & Teel JH (2012). Blue-Green Algae Response Strategy. Sherman, Texas.
http://www.co.grayson.tx.us/users/Health_Dept/Docs/Blue-Green_Algae_Response_Strategy.pdf
Gallons of Water Pounds of Water
263
2200
2109
17601
Human Health Recreational Ambient Water Quality Criteria or G-3
Swimming Advisories for Microcystins and Cylindrospermopsin - Draft
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