Assessment of Sediments from the Upper Mississippi River, Final Report, June 1997


             United States
             Environmental Protection
             Agency
            U.S. Department Of The
            Interior
EPA 823-R-9T-005
June i 997
             Office Of Water (4305)
 vvEPA
kf?
;
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An Assessment Of Sediments
From The Upper Mississippi
River

Final Report - June 1997  , it

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                                      NOTICE
This is the final report of research fUnded under USEPA Project No. DW14935486-01-0.
Mention of trade names or commercial products does not constitute endorsement or
recommendation for use.  Data from this report can be obtained electronically from:
      anonymous ftp - ftp://ftp.msc.nbs.gov/pub/umr/umr.zip
      world wide web - http://www.msc.nbs.gov/pubs/umr.html
For problems with access to the above addresses please e-mail the Webmaster, Chris Henke, at
chenke@msc.nbs.gov or call 573-875-5399.

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ABSTRACT
The U.S. Geological Survey (USGS) has been monitoring the Upper Mississippi River
(UMR) since 1987 to document the fate and transport of contaminats associated with sediments.
The UMR is that part of the river upstream of the confluence with the Ohio River at Cairo, IL and
consists of a series of 26 navigational pools created by a lock and dam system extending from
Minneapolis, MN to St. Louis, MO. The navigational pools are shallow, lake-like areas which
trap and store large quantities of fine-grained sediments during normal river flows. Concern with
the redistribution of the river sediments arose after the flood of 1993. This project was designed
to evaluate the current status of sediments in the UMR by: (1) measuring the concentrations of
contaminants in sediments of the UMR, (2) evaluating the toxicity of sediments collected from the
river, (3) determining the bioaccumulation of contaminants from UMR sediments using field-
collected and laboratory exposed oligochaetes, and (4) determining the benthic community
structure in fine-grain sediments within the river.
To conduct these assessments, sediment samples and benthic organisms were collected
from 24 of the 26 navigational pools in the river and from one pool in the Saint Croix River.
Two types of sediment samples were collected from the pools. One sediment sample was a
composite of 15 to 20 sediment grabs along one to five transects across the downstream one-third
of each pool (B samples). The other sediment sample was a composite of grabs from one station
on one transect within each pool (C samples). The latter stations were selected based on
historical chemistry data and the potential to collect oligochaetes. Samples were not collected
from the main navigation channels. Chapter 1 of this report describes whole-sediment toxicity
tests which were conducted for 28 days with the amphipod Hyalella azteca. Survival, growth and
sexual maturation were the measurement endpoints. Toxicity tests were conducted with both the
B and C sediment samples. Chapter 2 describes the bioaccumulation of contaminants from
sediments using field-collected oligochaetes and 28-day bioaccumulation studies conducted in the
laboratory with the oligochaete Lumbriculus variegatus. Bioaccumulation tests were conducted
with 13 of the 24 C sediment samples. Chapter 3 assesses the benthic community in all 24 C
samples. Using the Sediment Quality Triad approach, the status of UMR sediments was assessed
by integrating sediment chemistry, laboratory toxicity tests and benthic community measurements.
In the toxicity tests, Hyalella azteca survival was significantly reduced in only one
sediment sample (13B) relative to both a control and reference sediment. Growth of amphipods
was also reduced in only one sediment sample (26C). Sexual maturation was not significantly
reduced in any treatments. No correlations were observed between survival, growth or sexual
maturation and any of the physical or chemical sediment characteristics. Using sediment
chemistry and the Effect Range Median (ERM), 96% of the samples were classified as non-toxic
(i.e. measured chemical concentrations rarely exceeded ERMs). Classifications using ERMs and
sediment chemistry were consistent with the biological results from the H. azteca toxicity tests.
In the bioaccumulation tests, concentrations of contaminants were relatively low in native
oligochaetes collected from the pools as well as in oligochaetes exposed to the sediments in the
laboratory. Organochlorine pesticides were generally below detection in sediment and tissue
samples. Only aliphatic and polycyclic aromatic hydrocarbons (PAHs) and total polychlorinated
biphenyls were frequently measured above detection limits in oligochaete tissue and sediment
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samples. Concentrations for a specific contaminant in laboratory-exposed and field-collected
oligochaetes were similar within a station. About 90% of the paired PAH concentrations in
laboratory-exposed and field-collected oligochaetes were within a factor of three of one another.
With the detection limits used to analyze samples, contaminants were detected in tissue samples
more often than in sediment samples. Concentrations of PAHs in oligochaetes collected from the
pools or exposed in the laboratory to sediments from the UMR were up to 1000 times less than
tissue concentrations measured in oligochaetes from highly-contaminated sites within the U.S.
that our laboratory has previously studied.
The benthic community was dominated by oligochaetes and chironomids in 14 of the 23
sediment samples from the UMR and the one sediment sample from Saint Croix River. Fingernail
clams comprised a large portion of the community in 3 of the samples and exceeded 1,000/m2 in 5
of the samples. Total abundance values of invertebrates ranged from 250/m2 ( station 1C) to
22,3 89/m2 (station 19C) and were comparable to previously reported values for the UMR. The
frequency of chironomid mouthpart deformities was only 3% which is consistent with the
incidence of mouthpart deformities from uncontaminated sediments. Correlations between
benthic measures, sediment chemistry or other abiotic parameters exhibited few strong or
significant correlations indicating benthic communities are most likely controlled by factors
independent of contaminant concentrations.
The Sediment Quality Triad (Triad) is a weight-of-evidence approach used to assess the
contamination of sediments by integrating sediment chemistry, laboratory toxicity testing and
benthic community measures. Results from the Triad analysis indicated 88% of the samples were
classified as not impacted based on sediment chemistry, laboratory toxicity and benthic measures.
These results are consistent with the bioaccumulation study in which concentrations of
contaminants in tissue were less than other U.S. sites that our laboratory has previously studied.
In addition, pools in about the lower third of the river had lower sediment contaminant
concentrations, less accumulation of contaminants in tissue, and greater taxa richness.
Sediments are often both a sink for water-borne contaminants and a source of
contaminants to the overlying water. In addition, sediments may accumulate significant
concentrations of contaminants even when water quality criteria are not exceeded. The results
from the present study indicate that the UMR is not severely contaminated relative to other sites
that have been studied in the U.S. Perturbations that may occur could be attributed to
channelization, sedimentation from surface runoff or long term changes in the natural flow
conditions of the river due to lock and dam construction. This study only conducted a partial
assessment of the UMR sediments and included no assessment of river water. Further, this study
was a one-time assessment that was conducted after a major flood event and does not evaluate
temporal or spatial variability of sediment contamination within the pools. Future research on, or
management of, the Upper Mississippi River should evaluate the limitations of this study.
IV

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                                TABLE OF CONTENTS

List of Figures  	  iv

List of Tables	vii

List of Appendices	 viii

Chapter 1:    Evaluation of Contamination in Sediments Collected From
             Navigational Pools of the Upper Mississippi River Using a
             28-day Hyalella azteca Test.  Kemble, N.E.,
             E.L. Brunson, T.J. Canfield, F.J. Dwyer, and C.G. Ingersoll   	 1.1


Chapter 2:    An Evaluation of Bioaccumulation of Contaminants from
             Sediments from the Upper Mississippi River Using
             Field-collected Oligochaetes and Laboratory-exposed
             Lumbriculus variegatus.  Branson, E.L.,  T.J. Canfield,
             F.J. Dwyer, C.G. Ingersoll, andN.E. Kemble	2.1

Chapter 3:    Assessing Sediment Toxicity From Upper Mississippi
             River Navigational Pools Using a Benthic Invertebrate
             Community Evaluation and the Sediment Quality Triad
             Approach. Canfield, T.J., Brunson, E.L., F.J. Dwyer,
             C.G. Ingersoll, andN.E. Kemble 	 3.1

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                                  LIST OF FIGURES
Figure 1.1
Figure 1.2
Figure 1.3
Figure 1.4
Map of Upper Mississippi River from Minneapolis, MN to St. Louis, MO
Concentrations of simultaneously extracted metal (SEM) cadmium in
sediment samples compared to effect range median (ERM) for cadmium . .
Concentrations of SEM Pb in sediment samples compared to ERM for Pb
Concentrations of chrysene in sediment samples compared to ERM for
chrvsene 	
. . 1.15
. . 1.16
. . 1.17
1.18
Figure 1.5     Concentrations of benzo(a)pyrene in sediment samples compared to ERM
              for benzo(a)pyrene	  1.19

Figure 1.6     Number of ERM exceedances compared to sum ERM quotient  	  1.20

Figure 1.7     Survival versus sum ERM quotient samples compared to other known
              contaminated sites  	  1.21

Figure 2.1     Diagram of in-line flow splitter used to deliver water  	  2.12

Figure 2.2     Total accumulation of polycyclic aromatic hydrocarbons by
              laboratory-exposed Lumbriculus variegatus	  2.13

Figure 2.3     Total accumulation of polycyclic aromatic hydrocarbons by field-collected
              oligochaetes	  2.14

Figure 2.4     Comparison of tissue concentrations in laboratory-exposed Lumbriculus
              variegatus versus field-collected oligochaetes	  2.15

Figure 2.5     Ratio of tissue concentrations in laboratory-exposed Lumbriculus
              variegatus or field-collected oligochaetes for select PAHS	  2.16

Figure 2.6     Biota-sediment accumulation factors for laboratory-exposed Lumbriculus
              variegatus and field-collected oligochaetes for PAHs from the current
              study and calculated from PCB homolog data reported in the literature	2.17

Figure 3.1     Quadrant analysis for Hyalella azteca and sum ERM quotient  	3.19

Figure 3.2     Percent chironomid deformities for samples from the Upper
              Mississippi River  	  3.20
                                           VI

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                                   LIST OF TABLES
Table 1.1

Table 1.2

Table 1.3


Table 1.4


Table 1.5


Table 2.1



Table 2.2


Table 3.1

Table 3.2



Table 3.3
Results of sediment tests with Hyalella azteca .  .

Physical and chemical characteristics of sediments
Spearman rank correlations for simultaneously extracted metals and
physical sediment characteristics	•	
Regression data for amphipod survival, length and sexual maturation
with sediment physical and chemical characteristics	
Regression data normalized to total organic carbon for amphipod
survival, length and sexual maturation	
List of molecular weights and log Kow for contaminants used to
compare laboratory-exposed Lumbriculus variegatus and
field-collected oligochaetes	
Biota-sediment accumulation factors for the current study and reported
in the literature	

Percent contribution of each taxa to the overall total abundance estimates  .

Spearman rank correlation for select invertebrate metrics with naphthalene,
chrysene, benzo(a)pyrene, Cd, Ni, Pb, Zn, percent water, percent sand,
percent silt, percent clay and total organic carbon	
 Summary of quadrant analysis for scores of individual and combined benthic
 measures with the sum effect range median quotient	
Table 3.4     Summary of measures used for the Sediment Quality Triad
1.22

1.24


1.26


1.27


1.29



2.18


2.19

3.21



3.22


3.23

3.24,
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                               LIST OF APPENDICES

Appendix 1.1 Pore-water quality for whole-sediment tests	  1.30

Appendix 1.2 Mean measured overlying water quality for whole-sediment toxicity tests ...  1.32

Appendix 1.3 List of polycyclic aromatic hydrocarbons (PAHs) and organochlorines
             analyzed for in sediment	  1.34

Appendk 1.4 Amphipod length data from first study 	  1.35

Appendix 1.5 Amphipod length data from second study	  1.44

Appendix 1.6 Amphipod maturation and survival data from first study	  1.53

Appendix 1.7 Amphipdd maturation and survival data from second study	  1.54

Appendk 1.8 Concentrations of simultaneously extracted metals (SEM), acid volatile
             sulfides (AVS) and SEM/AVS 	  1.56

Appendix 1.9 Concentrations of organochlorines in sediment samples  	  1.58

Appendix 1.10 Concentrations of PAHs in sediment samples	  1.60

Appendix 2.1 Mean measured overlying water quality for bioaccumulation tests	2.20

Appendk 2.2 Tissue concentrations of organochlorine compounds measured in laboratory-
             exposed and field-collected oligochaetes	  2.21

Appendix 2.3 Tissue concentrations of PAHs measured in laboratory-exposed and
             field-collected oligochaetes	  2.22

Appendix 2.4 Total accumulation of PAHs in laboratory-exposed and field-collected
             oligochaetes	'.	  2.23

Appendk 2.5 Ratio of laboratory to field and field to laboratory tissue concentrations  ....  2.28

Appendix 3.1 Mean abundance data for Oligochaeta taxa	  3.25

Appendix 3.2 Mean abundance data for Chironomidae taxa	  3.26

Appendix 3.3 Mean abundance data for benthic invertebrate taxa excluding oligochaetes
             and chironomids	  3.27
                                          Vlll

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Chapter 1:  Evaluation of Contamination in  Sediments Collected from Navigational
 Pools of the Upper Mississippi River Using a 28 Day Hyalella azteca Test

Kemble, N.E., Brunson, E.L., Canfield, T.J., Dwyer, F.J., and Ingersoll, C.G.

Introduction

The Mississippi River is the largest river system in the United States.  Because of its location, the
river receives contaminant inputs from a variety of industrialized and agricultural sources. The
Upper Mississippi River (UMR), the stretch of river upstream from the confluence with the Ohio
River at Cairo, EL, contains a series of 26 navigational pools created by a lock and dam system
from St. Louis, MO to Minneapolis, MN (Rada et al 1990; Figure 1.1). These navigational pools
are shallow lake-like areas which trap and store large quantities (1 to 4 cm/yr) of primarily fine-
grained sediments during normal river flows (McHenry et al 1984; Nielsen et al 1984). Dredging
activities, commercial navigation, recreational boating and natural resuspension processes can
result in the remobilization of these sediments. Concern about the resuspension and transport of
these sediments and the contaminants associated with them arose after the flood of 1993 (Moody
and Meade 1995; Moody et al 1996).
   The United States Geological Survey (USGS) has been monitoring the transport and
degradation of pollutants in the UMR since the fall of 1987 (Moody and Meade 1995). Studies
have monitored concentrations of contaminants in fish (Hora 1984; Wiener et al 1984),
invertebrates (Beauvais etal 1995; Steingraeber and Wiener 1995), sediments (Bailey and Rada
1984; Wiener et al 1984; Rada et al 1990; Frazier et al. 1996; Ingersoll et al 1997) or a
combination of the three (Peddicord et al 1980; Boyer 1984) in select pools  in the UMR.
However, little information was available on contaminant concentrations and toxicity in sediment
samples throughout the entire pool system of the UMR.
   Four studies were conducted to assess the nature and extent of sediment contamination in the
navigational pools of the UMR:  (1) contaminant concentrations were measured in sediments
before and after the flood of 1993 (Moody et al 1996); (2) whole-sediment toxicity tests were
conducted (this chapter); (3) whole-sediment bioaccumulation tests were conducted (i.e.; Chapter
2); and (4) benthic-community structure were evaluated (i.e.; Chapter 3).  Sediment samples were
collected from June 11th to July 5th, 1994 from pool 1 (near Minneapolis, MN) to pool 26 (near
St. Louis, MO) of the UMR system (Figure 1.1). The objective of the study presented in this
chapter was to assess the toxicity of sediments from navigational pools of the UMR system using
28-day toxicity tests with the amphipod Hyalella azteca, measuring for potential effects on
survival, growth or sexual maturation.

Materials and Methods

Sample Collection, Handling, and Storage

Differential Global Positioning System (GPS) using a local reference was used to locate sampling
stations in the upper pools (1-14) and the Saint Croix River.  A differential GPS using the
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navigational beacon near St. Louis, MO. was the reference to locate sampling stations in the
lower pools (15-26).  A 3.5 composite sediment sample was collected from each of the 26
navigational pools (pool samples designated as "B" samples; Moody et al 1996). These
composite samples of surface (upper 10 cm) sediments were collected using a van Veen grab from
15 to 20 stations along one to five transects (typically 3 to 5 stations/transect) from the
downstream one-third of each navigation pool (except pool 17) in the UMR and from one site in
the Saint Croix River (SC) just upstream from its confluence with the Mississippi River in
Wisconsin (Figure 1.1; Moody etal 1996).  Samples were not collected from the main navigation
channel which was assumed to contain coarser sediment that had been deposited for a short
period of time. A 2-L subsample of the 3.5 L samples for toxicity testing and physical and
chemical characterization were removed and placed in a 2-L high density polyethylene (HDPE)
screw topped container.  Samples were stored in a cooler at 4°C for 7 to 14 days on the research
ship Acadiana, then shipped on ice to the Environmental and Contaminants Research Center
(ECRC - formerly the Midwest Science Center) in Columbia, MO. Two 125-mL subsamples
from each B sample were collected at the start of the toxicity tests for physical (grain size and
TOC) and chemical (organic and metal) characterization.
   A second composite sediment sample was also collected from each pool at one station on one
of the transects (station samples designated as "C" samples).  The individual stations (C samples)
were selected based on historical chemistry data and the potential for the collection of large
numbers of oligochaetes for bioaccumulation evaluations (Chapter 2). Station sediment samples
(C samples) for toxicity and bioaccumulation (Chapter 2) testing were collected with a Ponar grab
(S29 cm2 area). Each C sample was a composite sample collected from the upper 6 to 10 cm of
the sediment surface within  a 5-m radius area. A total of 35 to 80 L of sediment was collected
from each C station.  The sediment was then placed into a 120-L HDPE drum and homogenized
on ship with a stainless steel auger on a hand-held power drill. Subsamples of these C samples
were taken for (1) laboratory toxicity and laboratory bioaccumulation testing (10 L), (2) physical
characterization (250 mL) and chemical characterization (250 mL for organics and 250 mL for
metals) and (3) benthic invertebrate assessment (2 L).  The remaining C sample was then sieved
and native oligochaetes were collected for bioaccumulation analyses (Chapter 2).  Sediment
samples were stored in a cooler on the ship at 4°C for 7 to 14 days, then shipped on ice to the
ECRC in Columbia, MO. Once at the ECRC, sediment samples were stored in the dark at 4°C
until the start of the study. The control sediment (FLOR) used in the toxicity tests was a fine silt-
and clay-particle size soil collected near St. Louis MO.  This control sediment has been used in
previous studies (Kemble et al 1994).

Culturing of Test Organisms

Amphipods were mass cultured at 23°C with a luminance of about 800 lux according to
procedures outlined in Tomasovic et al (1995) using 80-L glass aquaria containing 50 L of ECRC
well water (hardness 283 mg/L as CaCO3, alkalinity 255 as CaCO3, pH 7.8).  Artificial substrates
were also placed in the amphipod culture aquaria (six 20-cm diameter sections/aquarium of
"coiled web material"; 3M Corp., Saint Paul, MN). Known-age amphipods were obtained by
isolating mixed-aged adults in a 5-mm mesh (#35 US Standard size sieve) sieve in a pan
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containing about 2 cm of well water.  After 24 h, well water was sprinkled through the sieve,
flushing <24-h-old amphipods into the pan below. These <24-h old  amphipods were then placed
into flow-through glass chambers for 10 d before the exposure began. Isolated amphipods were
fed maple leaves and ground Tetramin® ad lib until the start of the test.

Toxicity Tests

Sediment Preparation: Sediment samples were re-homogenized in the laboratory using either a
plastic spoon (for the B samples) or a hand-held power drill with a stainless steel auger (for the C
samples).  Subsamples were then collected for: (1) pore-water preparation, (2) physical and
chemical characterizations, (3) toxicity testing, and (4) bioaccumulation testing © samples only;
i.e., Chapter 2).

Water Quality:  About 170 mL of pore water was isolated from each sample by centrifugation at
4°C for 15 min at 5200 rpm (7000 x G). A 50-mL subsample for total sulfide determination was
removed from each sample and preserved with 0.1 mL of 2N zinc acetate solution (APHA 1985).
Total dissolved sulfide was determined with an Orion EA940 Expandable ionAnalyzer, Orion 94-
16 silver/sulfide electrode, and a Orion 90-02 double junction reference  electrode. Dissolved
oxygen (mg/L, with a YSI Model 54A oxygen meter and a YSI5739 probe), temperature (°C)
and conductivity (us/cm @ 25°C with a Orion 140 S-C-T meter and a 014010 conductivity cell)
were determined on the remaining volume. Subsamples of pore water were then removed for the
following determinations: total ammonia (mg/L) with an Orion EA940, and Orion 95-12 ammonia
electrode, alkalinity (mg/L, as CaCO3) and pH with an Orion EA940 Expandable ionAnalyzer,
Orion 917001 ATC probe, and Orion 8165BN combination pH probe, and total hardness as
(mg/L, as CaCO3) by EDTA titration. Unionized ammonia concentrations (mg/L, as NH3) were
calculated by adjusting total ammonia concentrations to pH and temperature using the formula
presented in Thurston et al (1979). Hydrogen sulfide concentrations (mg/L) were calculated by
adjusting the total dissolved sulfide concentrations to pH and temperature using the relationship
presented in Broderius and Smith (1977).
   Mean characteristics of porewater water quality (ranges in parentheses) are as follows: pH
7.45 (6.69 to 8.17); alkalinity 505 (244 to 852) mg/L; hardness 504  (148 to 852) mg/L; dissolved
oxygen 5.04 (1.50 to 9.35) mg/L; conductivity 906 (380 to 1680) us/cm @ 25°C; total ammonia
5.320 (1.210 to 22.700) mg/L; unionized ammonia 0.007 (0.000 to 0.025) mg/L; total sulfide
0.055 (0.000 to 0.569) mg/L; and hydrogen sulfide 0.023 (0.000 to 0.569) mg/L (Appendix 1.1).
   The following parameters were measured in overlying test water on Day -1 (the day before
amphipods were placed into the beakers) and at the end of each toxicity test: dissolved oxygen,
temperature, conductivity, pH, alkalinity, total hardness, and total ammonia. Methods used to
characterize overlying water quality in the whole-sediment tests were similar to the methods
described for characterization of pore water.  Dissolved oxygen, pH, and conductivity were also
measured weekly. Temperature in the water baths holding the exposure beakers was measured
daily. Overlying water pH, alkalinity, total hardness, conductivity and total ammonia
measurements were similar among all stations, the control, and the in flowing test water
(Appendix 1.2). Dissolved oxygen measurements were at or above  acceptable levels (>40% of
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saturation; ASTM 1995) in all treatments throughout the study (Appendix 1.2). Means (ranges in
parentheses) of overlying water quality of each parameter are as follows: pH 8.07 (7.58 to 8.72);
alkalinity 87 (59 to 151) mg/L; hardness 128 (111 to 160) mg/L; dissolved oxygen 6.70 (5.84 to
7.53) mg/L; conductivity 392 (359 to 428)  us/cm @25°C; total ammonia 0.416 (0.090 to 1.520)
mg/L; and unionized ammonia 0.003 (0.000 to 0.012) mg/L (Appendix 1.2).

Toxicity Tests: All sediment tests were started within three months of sample collection from the
field. Due to the number of samples collected, half of the samples (i.e., half of the sites) were
randomly selected for the initial testing. The second set of sediment samples was tested after
completion of testing of the first set of samples. Sediment samples for the toxicity tests were
homogenized the day before animals were added to exposure beakers (Day -1), using procedures
previously described.
   Toxicity tests were conducted with Hyalella azteca for 28 days. Effects of exposure to
sediments on survival, length, and sexual maturation of amphipods were measured (USEPA 1994;
ASTM 1995).  Each 300-mL beaker contained 100 mL of sediment and 150 mL of overlying
water.  The photoperiod was 16:8 h (lightdark) at a light intensity of about 500 lux. Four
replicate beakers/sample were placed in a ventilated water bath maintained at 23°C. Each beaker
received 1.0 volume additions/d of overlying water starting on Day -1 (Zumwalt et al 1994).
The overlying water used in the sediment toxicity exposures was a reconstituted moderately hard
water (hardness 95 mg/L as CaCO3, alkalinity 65-70 mg/L as CaCO3, pH 8.0-8.3; USEPA 1994).
One diluter cycle delivered 50 mL of water to each beaker (diluters cycled every 8 h ± 15 min).
Amphipods were acclimated to  the test water over 6 h before exposures began by sequentially
transferring animals at 2 h intervals into 50:50 and 25:75 mixtures of well watertest water, and
then into 100% test water. Tests were started on Day 0 by placing 10 amphipods (10- to 11-d
old) into each beaker. The water surface in each beaker was checked 15 min after organisms
were placed in the beaker for floating organisms. Amphipods in each beaker were fed 3 mg of
Purina Rabbit PelletsR in a water suspension three times a week for the first 7 days of the
exposure, and 6 mg three times a week for the last 21 days of the exposure. If excessive mold
(a60% sediment surface) was observed on the sediment surface of any of the beakers in a
treatment, feeding was withheld from all of the beakers for that treatment (the number of feedings
withheld ranged from 0 to 5 depending on the treatment; USEPA 1994; ASTM 1996).  Beakers
were observed daily for the presence of animals, signs of animal activity (i.e., burrowing), and to
monitor test conditions (i.e.; water clarity).
   Amphipods were retrieved from each beaker at the end of exposures using procedures
described in Kemble et al (1994).  Surviving organism were combined into a scintillation vial  and
preserved in 8% sugar formalin for later measurement of length, and sexual maturation. A Zeiss®
Interactive Digital Analysis System in combination with a Zeiss SV8 stereomicroscope at a
magnification of 25x was used to measure amphipods following methods described in Kemble et
al (1994). Amphipods were classified as either "mature male" or "not male" based on the
presence of an enlarged second gnathopod (Kemble et al 1994).  An enlarged second gnathopod
of male amphipods was a consistent measure, of sexual maturation (it is difficult to distinguish
immature males from females at this age).
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Chemical and Physical Characterization of Sediments

Acid-volatile Sulfides (AVS) and Simultaneously Extractable Metals (SEM): Subsamples of
sediments were measured for acid-volatile sulfides (AVS) and simultaneously extractable metals
(SEM) immediately after homogenization. Station samples (C samples) were collected on the
boat and stored at 4°C until shipment to the laboratory. Pool samples (B samples) were collected
in the laboratory immediately after sediment homogenization before the start of toxicity tests.
Concentrations of AVS in sediment samples were determined using a silver/sulfide electrode
following methods described in Brumbaugh et al (1994). Concentrations of SEM were
determined using atomic spectroscopy following methods described in Brumbaugh et al (1994).
Percentage recoveries for inorganics from both blank and sediment extracts averaged 96%.
The average range was from a low of 78% for antimony (spiked as sodium sulfide) in the
sediment extract to a high of 110% for Zn in the sediment extract. The average duplicate
coefficient of variation was 1.7% (6 compounds, n=2). Average duplicate coefficient of variation
ranged from 0.2% for both Pb samples to 5.1% for S in one of the duplicate samples.

Organochlorine Pesticides (OCPs), Poly chlorinated Biphenyls (PCBs), and Aliphatic and
Polycyclic Aromatic Hydrocarbons (PAHs): Sediment samples (C samples) were prepared for
the analyses of organochlorine pesticides (OCPs), polychlorinated biphenyls (PCBs), and aliphatic
and polycyclic aromatic hydrocarbons (PAHs) by extracting twenty grams of sediment with
acetone, followed by petroleum ether. A final acetone/petroleum ether extraction was done and
the extracts combined, centrifuged and transferred to a separatory funnel containing sufficient
water to facilitate partitioning of residues into petroleum ether portion. The petroleum ether was
washed twice with water and concentrated by Kuderna-Danish to appropriate volume.
Organochlorine determination was conducted by transferring an aliquot of concentrated extract
to a 1.6 g Florisil mini-column topped with 1.6 g sodium sulfate. Residues were eluted from the
column in two elution fractions. The first fraction consisted of 12 mL of hexane followed by 12
mL of 1% methanol in hexane; the-second fraction consisted of an additional 24 mL of 1%
methanol in hexane. Quantification of residues in the two Florisil fractions and three silicic acid
fractions was performed using a packed or megabore column and electron capture gas
chromatography.
Hydrocarbon determination was conducted by transferring a second aliquot of the
concentrated extract to a 20 g 1% deactivated silica gel column, topped with 5-g neutral alumina.
Aliphatic and polynuclear aromatic hydrocarbon residues were fractioned by eluting aliphatics
from the column with 100-mL petroleum ether (Fraction 1) followed by elution of aromatics
using, 100-mL 40% methylene chloride/60% petroleum ether, followed by 50-mL methylene
chloride (combined eluates, Fraction 2). .Quantification of fraction 1 by capillary column, flame
ionization gas chromatography was performed once the fraction was concentrated to appropriate
volume. The silica gel (fraction 2) containing aromatic hydrocarbons was concentrated,
reconstituted in methylene chloride and quantified by gas chromatography and mass spectrometry.
Average percent spike recovery for eighteen OCPs was 103% (n=2). The smallest average
spike recovery was 68% for HCB while o,p'-DDE had the greatest average spike recovery
(120%). Individual OCP concentrations were below minimum detection limits so duplicate
1.5

-------
analyses were not evaluated.  Average percent spike recovery for PAH compounds was 98% (29
compounds, n=2). Naphthalene (84%) had the smallest average percent recovery while
fluoranthene had the greatest average spike recovery (110%).  The average duplicate coefficient
of variation was 12.6% (13 compounds, n=2). Average duplicate coefficient,of variation ranged
from 0% for multiple PAHs in both duplicate samples to 61% for benzo(a)pyrene in one of the
samples.
   Methods  for the analyses of the B samples, detection limits and quality control are described in
Moody et al (1996). Quality control of B sediment samples analyzed for PAHs included: (1)
estimates of accuracy determined from the standard deviation of the percent recovery of
deuterated compounds added to the extracts and calculated based on absolute area counts and
external calibration, and (2) precision,  based on the relative standard deviation of the absolute
area of multiple analyses of a surrogate compound (Moody et al 1996). A list of all the PAHs
and OCPs analyzed for in both sets of sediment samples (B and C) are listed in Appendix 1.3.

Physical Characterization of Sediments

Physical characterization of sediments  included: (1) percentage water (Kemble et al 1993), (2)
particle size using a hydrometer (Forth et al 1982; Gee and Bauder 1986; Kemble et al 1993), and
(3) total organic carbon using a coulometric titration (Cahill et al 1987; Kemble et al 1993). All
physical characterizations  included analysis of duplicate samples. Differences in percentage water
for duplicate samples ranged from 0%  in treatments 2B, 7B, 13C, 14B and 18B to 7% in
treatment IOC.  Duplicate samples of control sediment, sucrose standards and blanks were
analyzed when determining sediment total organic. Precision and accuracy of the coulometric
technique used was tested against National Bureau of Standards and Standard Reference
Materials (NBS-SRM) with an error of less than 0.03% of the excepted values (Cahill et al 1987).
Differences between duplicates ranged from 0% in treatments 3B, 11B, 12B, 13C, 14C,  15C,
18C, 20C, 22C, 22B, 24C and 26C to  0.9% in treatments 5C, 9C and 26B.

Data Analysis and Statistics

Toxicity Tests: Before statistical analyses were performed, data for survival and maturation were
arcsin transformed. Comparisons of mean  survival and percentage sexual maturation were made
using a one-way nested analysis of variance (ANOVA) with mean separation by Fisher's protected
least significant difference test at alpha = 0.05 (Snedecor and Cochran 1982). Data for length had
a normal distribution and were not transformed before statistical analysis.  Comparison of mean
body length was made using a one-way ANOVA with  mean separation by Fisher's protected least
significant difference test at alpha = 0.05 (Snedecor and Cochran 1982).  A sample was
designated as toxic when survival, growth, or sexual maturation were significantly reduced
relative to the control and reference sediments.  Sediments from pools 6 and 11 were chosen as
reference sediment based on low concentrations of contaminants.  Simple linear regression was
used to compare physical and chemical sediment characteristics to amphipod survival, length or
sexual maturation. All statistical analyses were performed with Statistical Analysis System (SAS)
programs (SAS 1994).
                                          1.6

-------
Effects Range Median:  Chemistry concentrations and toxicity endpoints were evaluated using
28-day Hyalella azteca Effect Range Medians (ERMs) reported by Ingersoll et al (1996) and
Smith et al (1996). An ERM is defined as the concentration of a chemical in sediment above
which effects are frequently or always observed or predicted for most species (Long et al 1995).
The total number of individual ERMs exceeded with each sample was plotted against the sum
ERM quotient (SERM-Q; where Q is equal to the concentration of each chemical in the sediment
sample divided by the ERM for that chemical), similar to the toxic unit described by Canfield et al
(1996), Ingersoll et al (1996) and Swartz et al (1997). We chose to evaluate sediment toxicity
relative to nine ERMs which correctly classified >70% of the samples in Ingersoll et al (1996).
These 9 individual ERMs tended to minimize Type I (false positive) and Type II (false negative)
errors relative to other SECs reported by Ingersoll et al (1996). Due to insufficient chemistry
data for chromium and total PCBs, only 7 of the 9 individual ERMs were used in this evaluation.
These ERMs included: cadmium, lead, nickel, zinc, chrysene, benzo(a)pyrene, and
benzo(g,h,i)perylene.

Results and Discussion

Toxicity Tests

Survival of amphipods was significantly reduced relative to the control and reference sediments
only in the 13B treatment (Table 1.1).  Body length of amphipods was significantly reduced
relative to the control and reference sediments in only the 26C treatment (Table 1.1; Appendices
1.4 and 1.5).  Sexual maturation was not significantly reduced in any treatments when compared
to the control and reference sediments (Table  1.1; Appendices 1.6 and 1.7).
   Indigenous organisms recovered at the end of amphipod exposures included oligochaetes,
ostracods, clams, and a snail. Clam shells were present in many of the sediments; however, only a
few live clams were retrieved at the end of the exposure.  Pairs of amphipods were observed in
amplexus in the control,  1-B, 2-B, 5-B, 6-C, 8-B, 8-C, 9-B, 10-B, 11-B, 14-C, 15-B, 18-C, 24-B,
24-C, and 26-B treatments, and gravid females were observed in the control, 11-B, 16-C, and 24-
B treatments.
   Although significant differences in survival of amphipods relative to the control and reference
sediments were only observed in sample 13B, there was a relatively wide range in survival  among
the treatments.  For example survival was below 70% in 13 of the 51 treatments (Table 1.1).
Survival of amphipods in the control was acceptable (^80%), however, survival in two of the four
reference treatments (11C and 6B) was below 80%.  Subsequent studies have found that the
reconstituted water described in USEPA (1994) that was used to conduct this study does not
consistently support adequate survival and growth of Hyalella azteca in 28-day exposures
(McNulty 1995; Kemble et al 1996). Ingersoll et al (1997) retested sediment samples 4C, 11C,
14C, and 24C using well water as an overlying water and observed a mean survival of >90% in all
of the samples with no substantial effects on growth, or reproduction ofH. azteca. Survival of
amphipods in these same sediments ranged from 48% to 63% in the present chapter (Table 1.1).
Similarly, Benoit etal (1997) tested Station samples (7C, 9C, 13C, 22C,  arid 24C) in chronic
toxicity tests with midge Chironomus tentans using a natural overlying water and did not observe
                                           1.7

-------
effects on survival, growth, emergence, or reproduction.  Additional studies are ongoing to
evaluate 28-day Hyalella azteca exposures using reconstituted waters.

Physical and Chemical Characteristics of Sediments

Physical and chemical characteristics of sediment samples are listed in Table 1.2. Sediment
organic carbon content ranged from 0.2% for the sediment samples from Stations 6B and 20B to
5.2% for Station IOC.  Organic carbon content in the control sediment was 1.2%.  Percentage
solids ranged from 21% in the sediment sample from stations 4C and IOC to 84% for the
sediment sample from Station 20B. Classification of the sediment samples for grain size varied
from pool to pool (i.e., loam (11C), sandy-loam (8B), silly-clay-loam (25 C and 22C)) while the
control sediment was a silty-clay-loam (Table 1.2).  Acid volatile sulfide levels ranged from 0.005
umoles/g in the 1C sample to 63.0 umoles/g in the IOC sample (Table 1.2).
   Concentrations of simultaneously extracted metals in sediment samples are listed in Appendix
1.8.  Sediment from sample 4C had the highest concentrations of extractable SEM Cd, Cu, Ni,
and Pb. Sample 12C had the highest concentration of SEM Zn (Appendix 1.8). The sum
SEM/AVS molar ratio in the present study was typically less than 1 (except the two samples from
pool 1). This indicates the concentration of divalent metals listed in Appendix 1.8 were probably
not high enough to result in toxicity of the samples (DiToro et al 1990). Concentrations of SEM
Cd, Cu, Ni and Pb were highest hi sediment samples from treatment 4C (Appendix 1.8).
However, concentrations of SEM Cu and Pb were still below the ERMs reported by Ingersoll et
al (1996; Figures 1.2 and 1.3).
   Significant positive correlations were observed between SEM metals vs. TOC (Cu > Zn >
Cd>Pb>Ni), SEM metals vs. percentage clay (Zn>Ni>Pb>Cu>Cd) and between SEM metals vs.
percentage silt (Ni>Cu>Pb>Zn>Cd) when tested by Spearman's rho coefficient of rank correlation
(Table 1.3). The significant negative correlation with sand and the positive correlation with clay
and silt indicates that metals were concentrated in the finer sediment particles.
   Concentrations of organochlorine pesticides (OCPs) hi sediment samples are listed in Appendix
1.9. Concentrations of OCPs were below detection limits (0.01 ug/g) in all of the C samples
except the 2C and SCC samples which had detectable concentrations of DDE and ODD
(Appendix 1.9). Amphipod survival hi the 2C sediment sample was 75%. However, despite
having concentrations which were similar for both chemicals, survival of amphipods hi the SCC
sample was 90%.  This indicates that the levels of DDE and DDD detected in these samples was
not the sole cause of the lower survival observed in the 2C sediment sample.  Concentrations of
OCPs hi the B samples were at or below detection limits for 10 of the  15 individual pesticides
evaluated (Appendix 1.9).  Concentrations for all 5 OCPs detected hi the B samples were <; 0.079
ug/g dry weight and were below calculated ERMs (Smith et al 1996; Appendix 1.9).
   Concentrations of polycyclic aromatic hydrocarbons (PAHs) hi sediment samples are listed in
Appendix  1.10.  The highest concentrations were observed at Pool 1 and were generally lower in
the downstream pools.  Concentrations of PAHs in river sediments exceeded the Method Lower
Limit of Quantitation (MLLQ; 0.03 ug/g) hi at least one sediment sample for every PAH
evaluated (except for 1-methylnaphthalene; Appendix 1.10).  Concentrations of 4 of the 11 PAHs
measured exceed at least one calculated ERM (Ingersoll etal 1996; Figures 1.4 and 1.5).
                                          1.8

-------
Elevated PAH concentrations in sediment samples were, associated with sediment collected from
pools near Minneapolis, MN. Concentrations of PAHs below pool 4 were similar in the remaining
pools. Concentrations of fluoranthene exceeded the calculated ERM (0.175 ug/g) in 9 of the
sediment samples from the Upper Mississippi River. Amphipod survival in these samples was
above 75% in all but one of the samples (sample 4C which had a survival of 63%; Table 1.1).
This would indicate that concentrations of fluoranthene in these samples had little or no effect on
amphipod survival.

Comparisons of Sediment Characteristics to Toxicity Responses

Relationships of physical or chemical characteristics of sediments to toxicity were evaluated using
rank correlation (Table 1.4). No significant correlations were observed between survival, growth
or maturation and the measured physical or chemical characteristics of the sediment samples
(Table 1.4). Additionally, no significant correlation was observed between the toxicity endpoints
and concentrations of PAHs or OCPs normalized to total organic carbon concentrations (Table
1.5). Sediments from Pool 1 had the highest percent sand (>88%), but amphipod length and
maturation were not reduced with exposure to IB or 1C sediments relative to the control and
reference sediments (Table 1.1). Similarly, the control sediment had the highest percent silt and
clay relative to the other samples. Ingersoll and Nelson (1990), Kemble et al (1994), and
Ingersoll et al (1997) also reported sediment particle size did not affect the response ofHyalella
azteca in 28-d sediment exposures.
None of the 49 sediment samples exceeded any of the 7 individual ERMs. Use of these 7
ERMs correctly classified 47 of the 49 (96%) sediment samples from the UMR as non-toxic. The
two samples incorrectly classified were both type II errors (false negative; toxic sample that does
not exceed an ERM). This again may indicate something other than contaminants or
contaminants not measured were the cause of the relatively wide range in survival among the
treatments.
Additional ERMs for individual chemicals listed in Ingersoll et al (1996) and Smith et al
(1996) were also evaluated. About 20% of the sediment samples exceeded at least one of these
ERMs. However, use of these additional ERMs to classify samples as toxic or non-toxic resulted
in increased Type I error (false positive; non-toxic sample that exceeds an ERM). As was the
case when using only the seven ERMs, chemical concentrations from the two samples classified as
toxic did not exceed any of the additional ERMs.
The prediction of sediment toxicity was also evaluated using a toxic quotient approach. A
toxic quotient was calculated for each sample by first dividing the concentration of individual
chemicals by their respective ERM and then summing each of the individual values (Canfield et al
1996; Ingersoll et al. 1996). In the present study, quotients for the seven chemicals listed above
were used to calculate a toxic quotient for each sample (Table 1.2). Figure 1.6 plots the
relationship between the frequency of ERM exceedances and the sum of the ERM toxic quotient.
In the present study, the ERM toxic quotient was <• 2.6 and individual ERMs were not exceeded
indicating the sediment samples from the UMR were relatively non-contaminated compared to
sediments from areas of known contamination in the United States (Kemble et al 1994; Ingersoll
et al 1996). A toxic quotient approach was also used in Chapter 3 using a quadrant frequency
1.9

-------
analysis to evaluate the benthic community of the pools in the UMR system.

Summary

Toxicity tests using amphipods identified only two of the 49 sediment samples from the Upper
Mississippi River system as toxic (a significant reduction hi survival, growth or sexual maturation
compared to the control and reference sediments). However, there was a relatively wide range in
survival among the treatments. The overlying water used in this test was the reconstituted water
described in USEPA (1994), which McNulty (1995) and Kemble et al (1996) have reported does
not consistently support adequate survival ofHyalella azteca in 28-d sediment exposures.
Survival of amphipods and midge was >90% hi subsequent studies with sediments from the
present study when natural water was used as the overlying test water (Benoit et al 1997;
Ingersoll et al 1997). This would indicate that the reconstituted test water was a significant factor
in the wide range of survival observed in the present study.
Effect Range Medians (ERMs) were used to evaluate the toxicity of contaminants associated
with field collected sediments. ERMs correctly classified 96% of the UMR sediment samples as
non-toxic. The two samples incorrectly classified were type II errors (false negatives). Again this
indicates that factors other than contaminants or unmeasured contaminants may have been
responsible for the variation hi amphipod survival that was observed.
Concentrations of contaminants hi sediments from the UMR were typically 10 to 100 times
less than concentrations of contaminants hi sediments previously associated with toxicity (Kemble
et al 1994; Ingersoll et al 1996; Figure 1.7). This would indicate that the sediment samples from
the UMR were relatively non-contaminated compared to other areas of known contamination
across the United States.

Acknmvledgment. We would like to thank Linda Sappington for assistance on Quality Control
and Quality Assurance and the following individuals for their input on the project: Eugene Gfreer,
Doug Hardesty, Pam Haverland, Chris Henke, Ed Henry, Phil Lovely, John Moody, Shane
Reussler, Julie Soltvedt, Jeff Stevens, Ron Walton, Dave Whites, Dave Zumwalt, the Crew of the
Acadiana (Craig LeBoeuf and Pat Marmande), and the laboratories providing chemical analysis of
the sediment samples. Thanks to Dave Mount, James Fairchild and Parley Winger and two
anonymous reviewers for their helpful comments on the manuscript.
1.10

-------
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    1314
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Table 1.1.  Results of the Upper Mississippi River sediment tests with Hyalella azteca.  Means (Standard error of
the means in parentheses) within a column and within a set of sample are significantly different (p <0.05; n=4)
from the control and reference sediment and are designated with an asterix.
Sample
Survival (%)
Length (mm)1
Mature Males (%)
1st set of samples
Control
IB
1C
3B
5B
5C
SB
8C
10B
IOC
11B (reference)
11C (reference)
12B
12C
15B
15C
16B
16C
21B
21C
25B
25C
26B
26C
2nd Set of samples
Control
2B
2C
4B
4C
6B (reference)
6C (reference)
7B
7C
9B
9C
13B
13C
14B

80.0 (4.08)
92.5 (4.79)
65.0 (5.00)
95.0 (5.00)
80.0 (7.07)
80.0 (7.07)
97.5 (2.50)
92.5 (2.50)
92.5 (7.50)
72.5 (13.15)
87.5 (2.50)
57.5 (8.54)
72.5 (9.46)
85.0 (6.45)
90.0 (4.08)
72.5 (2.50)
70.0(9.13)
90.0 (7.07)
95.0 (2.89)
87.5 (4.79)
62.5 (13.15)
62.5 (15.48)
92.5 (4.79)
90.0 (7.07)

97.5 (2.50)
75.0 (8.66)
75.0 (10.41)
85.0 (6.45)
62.5 (21.75)
67.5 (17.02)
82.5 (2.50)
100.0 (0.00)
95.0 (2.89)
75.0 (10.41)
67.5 (13.77)
32.5 (7.50) *
47.5 (10.31)
65.0 (5.00)

3.39(0.16)
3.66(0.11)
3.17(0.11)
4.27 (0.08)
4.23 (0.06)
4.06 (0.10)
3.69 (0.09)
4.09(0.11)
4.28 (0.09)
3.86 (0.08)
4.31 (0.07)
3.61 (0.07)
3.48 (0.07)
3.78 (0.07)
3.74 (0.08)
3.59 (0.09)
3.72 (0.08)
3.83 (0.07)
3.46 (0.06)
3.87 (0.09)
3.60(0.11)
3.63 (0.08)
3.51 (0.09)
2.88 (0.01) *

2.59 (0.08)
4.07(0.11)
3.47(0.10)
3.39(0.10)
3.35 (0.09)
3.53 (0.09)
4.08(0.10)
3.66 (0.06)
3.70 (0.07)
3.72 (0.09)
3.65 (0.08)
3.87(0.19)
3.56 (0.11)
3.85 (0.12)

36.7 (8.91)
39.1 (5.71)
16.9 (6.90)
44.9 (8.43)
44.8 (10.30)
21.6 (4.23)
40.5 (7.72)
32.3 (7.68)
39.5 (18.49)
34.4 (6.88)
43.3 (11.57)
32.8 (15.79)
34.5 (3.00)
32.4 (5.85)
51.3 (11.46)
34.0 (8.64)
40.6 (6.56)
30.0(10.13)
52.2 (6.08)
51.4(5.29)
23.8 (10.51)
29.6 (8.34)
42.0 (6.82)
48.8(11.30)

5.9 (3.42)
31.3 (6.25)
43.8 (8.08)
36.7 (13.72)
12.1 (5.22)
26.9 (9.21)
54.5 (2.97)
42.5(10.31)
35.5 (3.41)
43.6 (6.47)
32.8 (11.24)
18.8(11.97)
50.0 (9.64)
31.6(7.36)
                                                 1.22
-------
Table 1.1. (continued)
Sample
Survival (%)
Length (mm)1
Mature Males (%)
14C
18B
18C
19B
19C
20B
20C
22B
22C
24B
24C
SCB
sec
47.5 (7.50)
77.5 (7.50)
72.5 (17.97)
85.0 (6.45)
72.5 (7.50)
82.5 (8.54)
95.0 (2.89)
85.0 (6.45)
52.5 (10.31)
87.5 (2.50)
60.0(8.16)
75.0 (10.41)
90.0 f4.08^
3.50(0.12)
3.57(0.12)
3.52 (0.09)
3.31 (0.07)
3.44 (0.07)
3.43 (0.08)
3.30 (0.06)
3.79(0.10)
3.64(0.11)
3.61 (0.08)
3.78 (0.12)
3.42 (0.10)
3.03 (0.06)
43.8 (15.72)
50.0 (18.89)
20.8 (7.50)
40.2 (7.50)
32.3 (15.91)
11.9(5.14)
27.2 (10.74)
24.4 (3.00)
39.9 (14.20)
34.4 (4.65)
66.9(14.19)
11.9(7.89)
31.7 (5.60)
'starting body length of amphipods in the 1st set of samples was 1.05 mm (0.02 SE, n=l 1) and was 1.17 mm (0.04
SE, n=10) in the 2nd set of samples.
                                                1.23
-------
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-------
Table 1.3. Spearman rank correlation for SEM Cd, Cu, Ni, Pb, and Zn with TOC, percent Sand,
percent Silt, and percent Clay for Upper Mississippi River sediments (excluding the control
sediment). All of the correlations listed below were significant (p <. 0.05).
Element
Cd
Cu
Ni
Pb
Zn
TOC%
0.826
0.868
0.808
0.823
0.854
%Sand
-0.449
-0.556
-0.634
-0.583
-0.589
%Silt
0.341
0.563
0.594
0.434
0.385
%Clay
0.394
0.468
0.553
0.549
0.570
                                          1.26
-------
Table 1.4. Linear regression (r2) of amphipod survival, length, or sexual maturation to sediment
physical and chemical characteristics. None of the regression were significant (p<0.05).

PW Total Ammonia
PW Unionized Ammonia
PW Total Sulfide
PW Hydrogen Sulfide
PW Alkalinity
PW Hardness
PWpH
PWDO
PW conductivity
AVS
Total Organic Carbon
Percent Sand
Percent Clay
Percent Silt
Percent Fines1
Percent Water
SEMCd
SEMCu
SEMNi
SEMPb
SEMZn
Toxaphene
Mirex
DDD
DDT
DDE
Endrin
Dieldrin
Heptachlor epoxide
Lindane
Naphthalene
Acenaphthalene
Acenaphthene
Phenanthrene
Anthracene
Survival
0.11
0.01
0.05
<0.01
<0.01
0.01
0.02
<0.01
0.01
0.11
0.02
0.02
0.01
0.05
0.02
<0.01
0.03
0.01
<0.01
0.02
0.02
0.01
0.05
<0.01
0.05
<0.01
0.05
0.05
0.05
0.05
0.04
0.04
<0.00
0.02
<0.01
Length
0.07
0.06
0.05
0.03
0.09
0.16
<0.01
0.04
0.13
<0.01
<0.01
<0.01
<0.01
<0.01
<0.01
0.02
0.04
0.03
<0.01
0.05
0.01
0.02
0.04
0.10
0.04
0.12
0.04
0.04
0.04
0.04
0.04
<0.01
0.02
0.01
<0.01
Sexual
Maturation
0.17
0.03
0.04
0.09
0.08
0.13
<0.01
0.01
0.03
0.02
0.02
<0.01
0.01
0.05
<0.01
0.02
0.11
0.03
<0.01
0.05
0.02
<0.01
<0.01
<0.01
<0.01
0.00
<0.01
<0.01
<0.01
<0.01
0.01
<0.01
0.07
0.01
<0.01
                                            1.27
-------
Table 1.4.  (Continued)


Fluorene
Fluoranthene
Chrysene
Pyrene
Benzo(b)fluoranthene
Benzo(k)fluoranthene
Benzo(a)pyrene
Indeo(l,2,3,-cd)pyrene
Benzofgfhjjftpervlene

Survival
0.01
0.04
0.02
0.03
0.04
0.01
0.01
<0.01
<0.01

Length
0.01
<0.01
<0.01
<0.01
<0.01
0.07
0.05
<0.01
0.01
Sexual
Maturation
<0.01
<0.01
<0.01
<0.01
<0.01
0.03
0.04
0.01
001
ISilt and Clay combined
                                         1.28
-------
Table 1.5. Linear regression (r2) of amphipod survival; length, sexual maturation to sediment
chemical characteristics normalized to organic carbon.  None of the regressions were significant
(p<0.05).

Toxaphene
Mirex
ODD
DDT
DDE
Endrin
Dieldrin
Heptachlor epoxide
Lindane
Naphthalene
Acenaphthalene
Acenaphthene
Phenanthrene
Anthracene
Fluorene
Fluoranthene
Chrysene
Pyrene
Benzo(b)fluoranthene
Benzo(k)fluoranthene
Benzo(a)pyrene
Indeo(l ,2,3 ,-cd)pyrene
Benzofgjh.j.ilpervlene
Survival
0.03
0.02
0.01
0.01
<0.01
0.01
<0.01
0.01
0.02
0.03
<0.01
0.02
0.01
<0.01
<0.01
0.04
0.14
0.05
0.06
0..01
0.04
<0.01
<0.01
Length
0.01
0.04
0.10
0.02
0.03
0.05
0.07
0.07
0.05
<0.01
<0.01
0.02
<0.01
0.06
<0.01
<0.01
0.07
0.06
0.02
0.03
0.07
0.05
0.05
Sexual
Maturation
0.05
<0.01
0.01
0.01
<0.01
<0.01
<0.01
<0.01
<0.01
<0.01
<0.01
0.02
<0.01
<0.01
0.01
<0.01
<0.01
<0.01
<0.01
<0.01
<0.01
0.01
0.01
                                            1.29
-------
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                                                                    ts  vo — i <— i
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                                                                    oo  >n n  I-H
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-------
Appendix 1.3. List of polycyclic aromatic hydrocarbons (PAHs) and organochlorines (OCs) analyzed for in the
                                              23.     2-methylnaphthalene
                                              24.     Biphenyl
                                              25.     Acenaphthalene
                                              26.     2,3,5-trimethylnaphthalene
                                              27.     Dibenzothiophene
                                              28.     Anthracene
                                              29.     Fluoranthene
                                              30.     Benzo(b)fluoranthene
                                              31.     Benzo(k)fluoranthene
                                              32.     Benzo(e)pyrene
                                              33.     Benzo(a)pyrene
                                              34.     1,2,5,6-dibenzanthracene
                                              35.     Cl-naphthalenes
                                              36.     C2-naphthalenes
                                              37.     C3-naphthalenes
                                              38.     C4-naphthalenes
                                              39.     Cl-dibenzothiophenes
                                              40.     C3-dibenzothiophenes
                                              41.     Cl-chrysenes
                                              42.     C2-chrysenes
                                              43.     C3-chrysenes
                                              44.     C4-chrysenes
                                              15.     HCB
                                              16.     alpha BHC
                                              17.     beta BHC
                                              18.     delta BHC
                                              19.     Oxychlordane
                                              20.     gamma Chlordane
                                              21.     trans-nonachlor
                                              22.     PCB 1242
                                              23.     PCB 1248
                                              24.     PCB 1254
                                              25.     PCB 1260
                                              26.     alpha Chlordane
                                              27.     o.p1 ODD
                                              28.     cis-nonchlor
                                              29.     o,p' DDT
sediment samples from the Upper Mississippi Ri
Polvcvclic aromatic hydrocarbons
1.
2.
3.
4.
5.
6.
7.
8.
9.
10.
11.
12.
13.
14.
15.
16.
17.
18.
19.
20.
21.
22.
Naphthalene
1-methylnaphthalene
2,6-dimethylnaphthalene
Acenaphthene
Fluorene
Phenathrene
1,-methylphenanthrene
Pyrene
Chrysene
1,2-Benzanthracene
Perylene
Indeno(l,2,3-cd)pyrene
Benzo(g,h,i)perylene
Cl-fluorenes
C2-fluorenes
C3-fluorenes
Cl-phenanthrenes
C2-phenanthrenes
C3-phenanthrenes
C4-phenanthrenes
Cl-fluoranthenes+Cl-pyrene
C2-dibenzothiophenes
Organochlorines
1.
2.
3.
4.
5.
6.
7.
8.
9.
10.
11.
12.
13.
14.
15.
16.
Lindane
Heptachlor
Aldrin
Heptachlor epoxide
Chlordane
Endo
Dieldrin
DDE
Endrin
Perthane
ODD
DDT
Methoxychlor
Mirex
Toxaphene
o.p1 DDE
                                                1.34
-------
Appendix 1.4. Amphipod length data for the 1st set of sediment samples. Replication (Rep), Animal (individual
animal number), and length (mean length for individual animal; n=2 measurements).
Sample
ARCH
ARCH
ARCH
ARCH
ARCH
ARCH
ARCH
ARCH
ARCH
ARCH
ARCH
IB
IB
IB
IB
IB
IB
IB
IB
IB
IB
IB
IB
IB
IB
IB
IB
IB
IB
IB
IB
IB
IB
IB
IB
IB
IB
IB
IB
IB
IB
IB
IB
IB
IB
IB
IB
1C
Animal
1
2
3
4
5
6
7
8
9
10
11
1
2
3
4
5
6
7
8
1
2
3
4
5
6
7
8
9
10
1
2
3
4
5
6
7
8
9
1
2
3
4
5
6
7
8
9
1
Rep
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
A
A
A
A
A
A
A
A
B
B
B
B
B
B
B
B
B
B
C
C
C
C
C
C
C
C
C
D
D
D
D
D
D
D
D
D
A
Length
0.946
1.077
1.134
1.092
0.973
1.024
1.122
1.086
1.000
1.051
1.086
3.373
2.522
3.048
3.084
3.610
3.090
3.655
3.265
3.843
3.666
4.348
3.630
3.783
3.765
4.207
3.556
3.846
3.332
4.398
4.889
3.864
4.646
3.409
4.942
4.073
4.883
4.222
3.173
2.925
2.474
4.282
2.752
3.179
3.164
3.472
3.215
3.170
Sample
1C
1C
1C
1C
1C
1C
1C
1C
1C
1C
1C
1C
1C
1C
1C
1C
1C
1C
1C
1C
1C
1C
1C
3B
3B
3B
3B
3B
3B
3B
3B
3B
3B
3B
3B
3B
3B
3B
3B
3B
3B
3B
3B
3B
3B
3B
3B
3B
Animal
2
3
4
5
1
2
3
4
5
6
1
2
3
4
5
6
7
1
2
3
4
5
6
1
2
3
4
1
2
3
4
5
6
7
8
1
2
3
4
5
6
7
8
9
10
11
1
2
Rep
A
A
A
A
B
B
B
B
B
B
C
C
C
C
C
C
C
D
D
D
D
•D
D
A
A
A
A
B
B
B
B
B
B
B
B
C
C
C
C
C
C
C
C
C
C
C
D
D
Length
3.610
3.475
3.783
3.669
2.794
3.616
3.102
4.112
3.078
2.946
2.157
2.656
2.943
2.271
3.445
2.695
2.531
2.725
3.433
4.076
3.616
3.454
2.656
4.595
4.456
4.441
3.690
5.497
4.119
4.885
4.985
4.388
4.077
4.607
4.030
4.296
4.118
4.335
4.036
4.935
4.068
4.027
3.879
3.891
3.891
3.923
4.089
4.476
                                                1.35
-------
Appendix 1.4. Amphipod length data for the 1st set of sediment samples (continued).
Sample
3B
3B
3B
3B
3B
3B
SB
5B
SB
SB
SB
SB
SB
SB
SB
SB
SB
SB
SB
SB
SB
SB
SB
SB
SB
SB
SB
SB
SB
SB
SB
SB
SB
SB
SB
SB
SB
5C
SC
5C
5C
5C
SC
5C
SC
5C
SC
SC
Animal
3
4
5
6
7
8
1
2
3
4
5
6
7
8
9
1
2
3
4
5
6
7
8
9
1
2
3
4
5
6
7
1
2
3
4
5
6
1
2
3
4
5
6
7
1
2
3
4
Rep
D
D
D
D
D
D
A
A
A
A
A
A
A
A
A
B
B
B
B
B
B
B
B
B
C
C
C
C
C
C
C
D
D
D
D
D
D
A
A
A
A
A
A
A
B
B
B
B
Length
4.053
4.181
3.805
3.778
4.867
3.920
3.974
3.825
4.037
4.103
4.754
4.249
4.357
3.669
4.112
4.467
3.965
4.198
3.834
4.524
4.404
4.216
4.070
3.971
4.682
4.088
4.387
4.987
4.216
5.265
3.732
4.422
4.088
4.159
4.261
4.091
4.162
3.251
4.216
4.073
3.230
4.512
5.157
3.765
4.192
3.696
3.974
3.672
Sample
5C
5C
5C
5C
5C
5C
5C
5C
5C
5C
5C
5C
5C
5C
5C
5C
5C
5C
5C
5C
8B
8B
8B
8B
8B
8B
8B
8B
8B
8B
8B
8B
8B
8B
8B
8B
8B
8B
8B
8B
8B
8B
8B
8B
8B
8B
8B
8B
Animal
5
6
1
2
3
4
5
6
7
8
9
1
2
3
4
5
6
7
8
9
1
2
3
4
5
6
7
8
9
10
1
2
3
4
5
6
7
8
9
10
1
2
3
4
5
6
7
8
Rep
B
B
C
C
C
C
C
C
C
C
C
D
D
D
D
D
D
D
D
D
A
A
A
A
A
A
A
A
A
A
B
B
B
B
B
B
B
B
B
B
C
C
C
C
C
C
C
C
Length
3.388
3.099
3.822
3.825
3.762
3.750
4.046
3.944
3.995
4.700
3.553
4.667
4.617
4.569
4.327
5.295
4.019
4.431
3.801
4.482
4.443
3.867
3.887
3.517
3.858
4.094
4.046
5.017
4.826
5.098
4.159
3.777
3.622
3.834
4.270
3.669
3.580
3.490
3.242
2.253
3.054
3.654
3.389
3.816
3.455
3.066
3.081
3.837
                                                1.36
-------
Appendix 1.4.  Amphipod length data for the 1st set of sediment samples (continued).
Sample
SB
SB
8B
8B
SB
8B
8B
8B
8B
8B
8B
8C
8C
8C
8C
8C
8C
8C
8C
8C
8C
8C
8C
8C
8C
8C
8C
8C
8C
8C
8C
8C
8C
8C
8C
8C
8C
8C
8C
8C
8C
8C
8C
8C
10B
10B
10B
10B
Animal
1
2
3
4
5
6
7
8
9
10
11
1
2
3
4
5
6
7
8
9
1
2
3
4
5
6
1
2
3
4
5
6
7
8
9
1
2
3
4
5
6
7
8
9
1
2
3
4
Rep
D
D
D
D
D
D
D
D
D
D
D
A
A
A
A
A
A
A
A
A
B
B
B
B
B
B
C
C
C
C
C
C
C
C
C
D
D
D
D
D
D
D
D
D
A
A
A
A
Length
3.352
3.675
3.151
4.045
3.557
3.374
3.560
2.940
3.922
3.075
3.373
4.283
4.238
3.581
4.027
3.916
2.807
3.367
2.458
4.142
4.241
4.253
3.858
5.099
3.831
3.678
4.136
4.184
4.127
3.467
4.524
3.876
3.461
3.587
4.425
4.460
4.346
4.322
3.654
4.747
4.383
5.024
5.575
5.015
4.594
4.104
3.959
3.632
Sample
10B
10B
10B
10B
10B
10B
10B
10B
10B
10B
10B
10B
10B
10B
10B
10B
10B
10B
10B
10B
10B
10B
10B
10B
10B
10B
10B
10B
10B
10B
10B
10B
IOC
IOC
IOC
IOC
IOC
IOC
IOC
IOC
IOC
IOC
IOC
IOC
IOC
IOC
IOC
IOC
Animal
5
6
7
8
9
10
1
2
3
4
5
6
7
8
9
1
2
3
4
5
6
7
1
2
3
4
5
6
7
8
9
10
1
2
3
4
5
6
1
2
3
4
5
6
1
2
3
4
Rep
A
A
A
A
A
A
B
B
B
B
B
B
B
B
B
C
C
C
C
C
C
C
D
D
D
D
D
D
D
D
D
D
A
A
A
A
A
A
B
B
B
B
B
B
C
C
C
C
Length
4.694
4.288
3.702
4.733
4.403
3.995
4.440
3.738
3.741
4.415
4.781
3.675
4.161
4.252
5.477
4.025
4.724
3.922
3.665
3.687
4.636
3.696
4.412
6.042
4.155
3.892
4.781
4.512
3.959
4.276
4.739
4.001
3.641
3.829
4.573
4.052
3.989
3.396
3.321
3.944
4.086
3.411
3.647
3.844
3.826
3.575
4.122
4.815
                                                1.37
-------
Appendix 1.4. Amphipod length data for the 1st set of sediment samples (continued).
Sample
IOC
IOC
IOC
IOC
IOC
IOC
IOC
IOC
IOC
IOC
IOC
IOC
IOC
11B
11B
11B
11B
11B
1IB
11B
HB
11B
11B
11B
11B
11B
11B
11B
11B
11B
11B
11B
11B
11B
11B
11B
11B
11B
HB
11B
11B
11B
11B
11B
11B
11B
11B
11B
Animal
5
6
7
8
9
1
2
3
4
5
6
7
8
1
2
3
4
5
6
7
8
1
2 ,
3
4
5
6
7
8
9
1
2
3
4
5
6
7
8
9
1
2
3
4
5
6
7
8
9
Rep
C
C
C
C
C
D
D
D
D
D
D
D
D
A
A
A
A
A
A
A
A
B
B
B
B
B
B
B
B
B
C
C
C
C
C
C
C
C
C
D
D
D
D
D
D
D
D
D
Length
4.691
3.947
3.620
3.638
4.270
3.807
4.255
3.650
3.811
2.858
3.623
4.180
3.547
4.589
4.051
4.333
4.164
4.152
4.262
4.066
3.950
3.828
4.220
5.000
3.768
4.119
4.244
3.408
3.661
4.235
4.357
4.057
4.878
4.351
5.122
4.408
4.351
5.006
5.116
3.479
4.116
4.661
4.432
4.577
4.360
5.027
4.137
4.217
Sample
11C
11C
11C
lie
lie
lie
nc
nc
iic
nc
nc
nc
nc
nc
nc
nc
nc
nc
iic
iic
iic
nc
nc
12B
12B
12B
12B
12B
12B
12B
12B
12B
12B
12B
12B
12B
12B
12B
12B
12B
12B
12B
12B
12B
12B
12B
12B
12B
Animal
1
2
3
4
5
6
7
1
2
3
4
5
1
2
3
4
5
6
1
2
3
4
5
1
2
3
4
5
6
7
8
9
1
2
3
4
5
6
7
8
9
1
2
3
4
5
6
7
Rep
A
A
A
A
A
A
A
B
B
B
B
B
C
C
C
C
C
C
D
D
D
D
D
A
A
A
A
A
A
A
A
A
B
B
B
B
B
B
B
B
B
C
C
C
C
C
C
C
Length
3.813
3.095
3.628
3.357
3.741
3.143
3.732
3.741
4.179
3.325
4.107
3.497
3.280
3.664
3.571
4.146
4.122
3.652
3.315
3.717
3.688
3.574
3.057
4.003
4.275
4.095
3.586
3.414
3.837
2.515
3.154
3.870
3.210
2.669
3.447
3.678
2.964
3.039
3.873
3.769
3.876
3.453
3.755
3.293
3.036
3.494
3.335
3.512
                                               1.38
-------
Appendix 1.4.  Amphipod length data for the 1st set of sediment samples (continued).
Sample
12B
12B
12B
12B
12B
12B
12B
12C
12C
12C
12C
12C
12C
12C
12C
12C
12C
12C
12C
12C
12C
12C
12C
12C
12C
12C
12C
12C
12C
12C
12C
12C
12C
12C
12C
12C
12C
12C
12C
12C
12C
12C
12C
12C
15B
15B
15B
15B
Animal
1
2
3
4
5
6
7
1
2
3
4
5
6
7
8
9
1
2
3
4
5
6
7
8
9
10
1
2
3
4
5
6
7
8
1
2
3
4
5
6
7
8
9
10
1
2
3
4
Rep
D
D
D
D
D
D
D
A
A
A
A
A
A
A
A
A
B
B
B
B
B
B
B
B
B
B
C
C
C
C
C
C
C
C
D
D
D
D
D
D
D
D
D
D
A
A
A
A
Length
3.663
3.352
2.808
3.565
3.988
3.293
3.518
4.236
4.233
4.209
3.249
4.518
4.155
3.051
3.738
4.224
4.026
3.477
3.813
4.353
3.669
4.242
3.711
4.506
3.033
3.594
3.960
3.615
4.062
3.798
3.972
4.110
2.748
4.062
3.639
3.705
3.408
3.327
3.249
3.831
3.240
4.083
3.444
3.681
3.801
4.273
3.870
3.054
Sample
15B
15B
15B
15B
15B
15B
15B
15B
15B
15B
15B
15B
15B
15B
15B
15B
15B
15B
15B
15B
15B
15B
15B
15B
15B
15B
15B
15B
15B
15B
15C
15C
15C
15C
15C
15C
15C
15C
15C
15C
15C
15C
15C
15C
15C
15C
15C
15C
Animal
5
6
7
8
9
1
2
3
4
5
6
7
8
1
2
3
4
5
6
7
8
9
1
2
3
4
5
6
7
8
1
2
3
4
1
2
3
4
5
6
7
1
2
3
4
5
6
7
Rep
A
A
A
A
A
B
B
B
B
B
B
B
B
C
C
C
C
C
C
C
C
C
D
D
D
D
D
D
D
D
A
A
A
A
B
B
B
B
B
B
B
C
C
C
C
C
C
C
Length
3.293
3.801
3.341
3.968
4.216
3.873
4.736
3.813
3.699
3.995
4.401
4.533
4.073
3.364
3.811
3.485
4.006
4.003
2.624
3.926
4.036
3.953
3.281
3.355
3.444
3.447
3.267
3.314
3.550
3.494
3.169
2.908
3.033
4.287
4.240
3.278
3.497
3.391
3.796
4.219
4.459
4.932
4.764
3.698
3.352
3.538
3.355
3.113
                                                1.39
-------
Appendix 1.4. Amphipod length data for the 1st set of sediment samples (continued).
Sample
15C
15C
15C
15C
15C
15C
15C
15C
15C
15C
16B
16B
16B
16B
16B
16B
16B
16B
16B
16B
16B
16B
16B
16B
16B
16B
16B
16B
16B
16B
16B
16B
16B
16B
16B
16B
16C
16C
16C
16C
16C
16C
16C
16C
16C
16C
16C
16C
Animal
8
1
2
3
4
5
6
7
8
9
1
2
3
4
5
6
7
8
9
1
2
3
4
5
1
2
3
4
5
6
1
2
3
4
5
6
1
2
3
4
5
6
1
2
3
4
5
6
Rep
C
D
D
D
D
D
D
D
D
D
A
A
A
A
A
A
A
A
A
B
B
B
B
B
C
C
C
C
C
C
D
D
D
D
D
D
A
A
A
A
A
A
B
B
B
B
B
B
Length
4.077
3.234
3.589
3.059
3.391
3.793
3.166
3.722
3.056
3.204
3.196
3.507
3.547
4.435
3.302
3.485
3.581
3.245
2.920
3.097
3.838
3.941
4.510
3.072
4.078
3.889
3.805
3.625
4.134
3.917
4.280
4.230
3.699
3.733
3.764
3.864
4.261
3.494
4.469
3.901
3.929
2.770
4.581
3.327
3.777
3.907
3.578
4.009
Sample
16C
16C
16C
16C
16C
16C
16C
16C
16C
16C
16C
16C
16C
16C
16C
16C
16C
16C
16C
16C
16C
16C
16C
16C
21B
21B
21B
21B
21B
21B
21B
21B
2 IB
21B
2 IB
21B
21B
21B
21B
21B
21B
21B
21B
21B
21B
21B
21B
21B
Animal
7
8
9
1
2
3
4
5
6
7
8
9
1
1
2
3
4
5
6
7
8
9
10
11
1
2
3
4
5
6
7
8
9
10
1
2
3
4
5
6
7
8
9
1
2
3
4
5
Rep
B
B
B
C
C
C
C
C
C
C
C
C
C
D
D
D
D
D
D
D
D
D
D
D
A
A
A
A
A
A
A
A
A
A
B
B
B
B
B
B
B
B
B
C
C
C
C
C
Length
3.612
3.755
3.590
3.497
3.503
3.308
4.084
3.330
3.637
3.991
3.376
3.951
3.376
3.851
3.680
3.730
3.541
4.438
4.364
4.659
4.165
4.308
4.395
3.826
3.280
3.224
3.301
3.916
3.200
3.999
3.107
3.483
2.872
3.265
3.283
3.319
3.030
3.781
4.160
4.178
2.809
3.775
3.856
3.480
3.161
3.808
3.579
4.056
                                               1.40
-------
Appendix 1.4. Amphipod length data for the 1st set of sediment samples (continued).
Sample
2 IB
2 IB
21B
2 IB
21B
2 IB
21B
2 IB
21B
2 IB
2 IB
2 IB
2 IB
21C
21C
21C
21C
21C
21C
21C
21C
21C
21C
21C
21C
21C
21C
21C
2 1C
21C
21C
21C
21C
21C
21C
21C
21C
21C
21C
21C
21C
21C
21C
21C
21C
21C
21C
21C
Animal
6
7
8
1
2
3
4
5
6
7
8
9
10
1
2
3
4
5
6
7
8
9
1
2
3
4
5
6
7
8
1
2
3
4
5
6
7
8
1
2
3
4
.5
6
7
8
9
10
Rep
C
C
C
D
D
D
D
D
D
D
D
D
D
A
A
A
A
A
A
A
A
A
B
B
B
B
B
B
B
B
C
C
C
C
C
C
C
C
D
D
D
D
D
D
D
D
D
D
Length
3.501
3.960
3.579
3.611
2.887
3.140
3.632
2.565
3.543
3.811
3.012
3.537
3.254
4.595
3.895
4.476
3.397
4.512
3.343
4.509
3.850
3.808
3.069
3.358
2.863
3.295
3.069
3.671
3.110
3.376
3.865
4.539
4.109
4.366
4.921
2.920
3.901
3.620
4.047
4.273
3.373
4.545
3.987
3.957
4.643
4.094
4.050
4.088
Sample
25B
25B
25B
25B
25B
25B
25B
25B
25B
25B
25B
25B
25B
25B
25B
25B
25B
25B
25B
25B
25B
25B
25B
25C
25C
25C
25C
25C
25C
25C
25C
25C
25C
25C
25C
25C
25C
25C
25C
25C
25C
25C
25C
25C
25C
25C
25C
25C
Animal
1
2
3
4
5
6
1
2
3
4
5
6
7
1
2
3
4
5
1
2
3
4
5
1
2
3
4
5
6
7
8
1
2
1
2
3
4
5
6
7
8
9
10
1
2
3
4
5
Rep
A
A
A
A
A
A
B
B
B
B
B
B
B
C
C
C
C
C
D
D
D
D
D
A
A
A
A
A
A
A
A
B
B
C
C
C
C
C
C
C
C
C
C
D
D
D
D
D
Length
2.579
4.374
3.433
3.347
3.735
2.955
3.777
3.726
3.840
3.735
4.225
4.237
4.129
3.816
2.854
3.675
4.094
3.923
3.837
2.561
3.565
3.430
2.976
2.952
3.120
3.953
3.831
4.052
3.834
3.729
3.093
5.053
3.920
3.547
3.487
3.469
3.299
3.215
3.681
3.356
3.108
3.571
3.448
3.645
4.494
3.344
3.538
3.729
                                                  1.41
-------
Appendix 1.4. Amphipod length data for the 1st set of sediment samples (continued).
Sample
25C
25C
25C
25C
26B
26B
26B
26B
26B
26B
26B
26B
26B
26B
26B
26B
26B
26B
26B
26B
26B
26B
26B
26B
26B
26B
26B
26B
26B
26B
26B
26B
26B
26B
26B
26B
26B
26B
26B
26C
26C
26C
26C
26C
26C
26C
26C
26C
Animal
6
7
8
9
1
2
3
4
5
6
7
8
9
10
1
2
3
4
5
6
7
8
1
2
3
4
5
6
7
8
9
1
2
3
4
5
6
7
8
1
2
3
4
5
6
7
8
9
Rep
D
D
D
D
A
A
A
A
A
A
A
A
A
A
B
B
B
B
B
B
B
B
C
C
C
C
C
C
C
C
C
D
D
D
D
D
D
D
D
A
A
A
A
A
A
A
A
A
Length
3.266
3.648
3.887
3.953
3.090
3.837
3.750
3.642
4.744
4.735
4.163
3.319
4.072
4.603
3.883
4.253
2.313
3.762
3.139
3.536
3.825
4.184
3.072
2.958
3.293
2.922
3.338
3.039
3.533
2.961
3.003
2.946
3.740
3.434
3.219
3.084
2.931
2.925
3.344
2.701
3.018
3.039
2.832
2.659
3.341
3.063
3.012
3.126
Sample
26C
26C
26C
26C
26C
26C
26C
26C
26C
26C
26C
26C
26C
26C
26C
26C
26C
26C
26C
26C
26C
26C
26C
26C
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
Animal
10
1
2
3
4
5
6
7
1
2
3
4
5
6
1
2
3
4
5
6
7
8
9
10
1
2
3
4
5
6
7
1
2
3
4
5
6
7
8
1
2
3
4
5
6
7
1
2
Rep
A
B
B
B
B
B
B
B
C
C
C
C
C
C
D
D
D
D
D
D
D
D
D
D
A
A
A
A
A
A
A
B
B
B
B
B
B
B
B
C
C
C
C
C
C
C
D
D
Length
2.829
2.764
3.267
3.084
2.713
2.875
3.171
3.006
2.761
2.686
2.731
2.883
2.632
2.680
3.204
3.012
2.665
2.620
2.593
2.764
2.964
3.054
2.958
2.255
2.734
4.479
3.796
3.362
4.323
3.401
3.826
4.156
3.087
3.237
3.278
4.587
2.455
3.332
3.434
2.731
1.338
1.976
3.683
3.668
2.973
1.868
4.341
3.790
                                               1.42
-------
Appendix 1.4. Amphipod length data for the 1st set of sediment samples (continued).
Sample Animal Rep    Length
Sample Animal Rep    Length
FLOR
FLOR
FLOR
FLOR
3
4
5
6
D
D
D
D
3.644
4.072
2.578
4.760
                                                   FLOR    7     D    3.976
                                                   FLOR    8     D    4.231
                                                   FLOR    9     D    1.967
                                              1.43
-------
Appendix 1.5. Amphipod length data for the 2nd set of samples.  Replication (Rep), Animal (individual animal
number), and length (mean length for individual animal).
Sample Animal
ARCH 1
ARCH 2
ARCH 3
ARCH 4
ARCH 5
ARCH 6
ARCH 7
ARCH 8
ARCH 9
ARCH 10
2B 1
2B 2
2B 3
2B 4
2B 5
2B 6
2B 7
2B 8
2B 1
2B 2
2B 3
2B 4
2B 5
2B 6
2B 7
2B 8
2B 1
2B 2
2B 3
2B 4
2B 1
2B 2
2B 3
2B 4
2B 5
2B 6
2B 7
2B 8
2C 1
2C 2
2C 3
2C 4
2C 5
2C 1
2C 2
2C 3
2C 4
2C 5
Rep
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
A
A
A
A
A
A
A
A
B
B
B
B
B
B
B
B
C
C
C
C
D
D
D
D
D
D
D
D
A
A
A
A
A
B
B
B
B
B
Length
1.339
1.369
1.193
1.178
1.101
1.107
1.021
1.056
1.134
1.196
4.177
3.556
3.890
4.572
4.001
3.920
4.467
4.195
4.461
4.491
3.729
4.246
4.622
3.502
4.718
3.421
5.184
3.980
5.558
4.213
2.874
4.548
3.027
3.711
3.568
4.040
3.777
3.436
3.108
2.898
2.737
2.880
3.102
3.096
3.442
3.359
3.403
2.764
Sample
2C
2C
2C
2C
2C
2C
2C
2C
2C
2C
2C
2C
2C
2C
2C
2C
2C
2C
2C
2C
2C
4B
4B
4B
4B
4B
4B
4B
4B
4B
4B
4B
4B
4B
4B
4B
4B
4B
4B
4B
4B
4B
4B
4B
4B
4B
4B
4B
Animal
6
7
8
9
10
1
2
3
4
5
6
7
1
2
3
4
5
6
7
8
9
1
2
3
4
1
2
3
4
5
6
7
8
1
2
3
4
5
6
7
8
9
1
2
3
4
5
6
Rep
B
B
B
B
B
C
C
C
C
C
C
C
D
D
D
D
D
D
D
D
D
A
A
A
A
B
B
B
B
B
B
B
B
C
C
C
C
C
C
C
C
C
D
D
D
D
D
D
Length
3.066
3.475
3.200
2.755
3.956
2.848
3.938
4.109
3.884
3.514
3.804
4.046
4.593
3.093
2.352
4.001
3.968
4.443
3.117
4.234
4.243
3.616
4.485
4.467
3.174
3.337
3.243
3.295
2.579
3.391
3.343
4.148
2.841
3.415
3.457
3.270
3.682
3.944
2.291
3.433
3.388
4.413
2.600
2.444
3.860
4.205
3.240
2.850
                                               1.44
-------
Appendix 1.5.  Amphipod length data for the 2nd set of sediment samples (continued).
Sample
4B
4B
4B
4B
4B
4B
4B
4B
4C
4C
4C
4C
4C
4C
4C
4C
4C
4C
4C
4C
4C
4C
4C
4C
4C
4C
4C
4C
4C
4C
4C
4C
4C
4C
6B
6B
6B
6B
6B
6B
6B
6B
6B
6B
6B
6B
6B
6B
Animal
7
8
9
10
11
12
13
14
1
2
3
4
5
6
7
8
9
10
11
1
2
3
4
5
6
7
8
1
2
3
4
5
6
7
1
2
3
4
5
6
7
1
2
1
2
3
4
5
Rep
D
D
D
D
D
D
D
D
B
B
B
B
B
B
B
B
B
B
B
C
C
C
C
C
C
C
C
D
D
D
D
D
D
D
A
A
A
A
A
A
A
B
B
C
C
C
C
C
Length
3.502
3.213
2.714
3.821
3.478
2.868
2.922
3.785
3.024
2.744
2.594
3.661
2.802
3.195
2.934
3.556
3.562
3.234
3.439
3.093
3.030
4.269
3.036
3.039
3.986
3.830
3.144
3.493
3.758
4.323
3.721
3.427
3.195
3.090
3.317
3.508
3.499
3.326
3.132
3.493
2.949
3.024
3.908
3.132
3.636
4.013
4.443
3.741
Sample
6B
6B
6B
6B
6B
6B
6B
6B
6B
6B
6B
6B
6B
6C
6C
6C
6C
6C
6C
6C
6C
6C
6C
6C
6C
6C
6C
6C
6C
6C
6C
6C
6C
6C
6C
6C
6C
6C
6C
6C
6C
6C
6C
6C
6C
6C
7B
7B
Animal
6
7
8
9
10
1
2
3
4
5
6
7
8
1
2
3
4
5
6
7
8
9
1
2
3
4
5
6
7
8
1
2
3
4
5
6
7
8
1
2
3
4
5
6
7
8
1
2
Rep
C
C
C
C
C
D
D
D
D
D
D
D
D
A
A
A
A
A
A
A
A
A
B
B
B
B
B
B
B
B
C
C
C
C
C
C
C
C
D
D
D
D
D
D
D
D
A
A
Length
4.360
3.045
3.200
3.242
3.831
2.943
3.311
3.935
4.634
3.553
2.994
3.335
3.777
3.568
3.819
2.943
4.204
4.336
5.462
3.027
4.608
4.019
4.790
4.775
4.183
3.870
4.688
4.760
4.617
3.855
3.583
4.682
3.439
3.317
4.969
3.601
3.589
4.115
3.765
3.911
3.622
3.941
4.318
4.464
4.085
3.657
3.886
3.844
                                                1.45
-------
Appendix 1.5.  Amphipod length data for the 2nd set of sediment samples (continued).
Sample
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7B
7C
7C
7C
7C
7C
7C
7C
7C
7C
7C
Animal
3
4
5
6
7
8
9
10
1
2
3
4
5
6
7
8
9
10
1
2
3
4
5
6
7
8
9
10
1
2
3
4
5
6
7
8
9
10
1
2
3
4
5
6
7
8
9
1
Rep
A
A
A
A
A
A
A
A
B
B
B
B
B
B
B
B
B
B
C
C
C
C
C
C
C
C
C
C
D
D
D
D
D
D
D
D
D
D
A
A
A
A
A
A
A
A
A
B
Length
3.605
3.725
3.635
3.695
3.361
4.294
3.537
4.127
3.263
3.185
3.835
3.447
3.263
3.587
4.262
3.832
3.158
3.641
3.549
3.531
3.531
3.140
3.087
3.084
3.486
3.110
4.411
3.570
4.470
4.136
4.238
4.050
3.543
3.707
3.638
3.948
3.543
3.590
4.405
4.160
3.283
3.659
3.468
3.090
4.005
3.361
3.987
4.178
Sample
7C
7C
7C
7C
7C
7C
7C
7C
7C
7C
7C
7C
7C
7C
7C
7C
7C
7C
7C
7C
7C
7C
7C
7C
7C
7C
7C
7C
7C
9B
9B
9B
9B
9B
9B
9B
9B
9B
9B
9B
9B
9B
9B
9B
9B
9B
9B
9B
Animal
2
3
4
5
6
7
8
9
10
1
2
3
4
5
6
7
8
9
1
2
3
4
5
6
7
8
9
10
11
1
2
3
4
5
6
7
1
2
3
4
5
1
2
3
4
5
6
7
Rep
B
B
B
B
B
B
B
B
B
C
C
C
C
C
C
C
C
C
D
D
D
D
D
D
D
D
D
D
D
A
A
A
A
A
A
A
B
B
B
B
B
C
C
C
C
C
C
C
Length
3.593
3.889
3.859
3.546
4.172
4.232
3.304
3.895
4.348
3.507
3.450
4.578
3.725
3.701
3.435
3.656
3.534
3.898
3.602
3.743
4.005
3.925
3.811
3.671
3.352
3.307
3.477
3.632
1.813
4.027
4.675
4.033
3.801
• 3.178
3.587
3.443
3.301
3.398
5.358
4.527
3.346
3.214
3.455
3.705
3.844
3.870
3.654
3.843
                                               1.46
-------
Appendix 1.5.  Amphipod length data for the 2nd set of sediment samples (continued).
Sample
9B
9B
9B
9B
9B
9B
9B
9B
9B
9B
9B
9B
9C
9C
9C
9C
9C
9C
9C
9C
9C
9C
9C
9C
9C
9C
9C
9C
9C
9C
9C
9C
9C
9C
9C
9C
9C
9C
9C
9C
9C
9C
9C
9C
13B
13B
13B
13B
Animal
8
9
10
11
1
2
3
4
5
6
7
8
1
2
3
4
1
2
3
4
5
6
7
8
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
1
2
3
4
1
2
1
2
Rep
C
C
C
C
D
D
D
D
D
D
D
D
A
A
A
A
B
B
B
B
B
B
B
B
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
D
D
D
D
A
A
B
B
Length
3.772
3.352
3.566
3.461
3.662
3.759
2.822
3.304
4.416
3.753
3.699
3.361
2.913
2.976
3.666
4.283
4.093
3.843
3.374
4.202
4.081
3.708
4.280
4.018
3.072
3.229
3.334
3.982
3.681
3.509
3.178
3.340
3.566
3.692
3.162
2.825
3.289
3.656
4.075
4.256
4.100
3.991
3.256
4.142
5.304
4.062
4.652
3.870
Sample
13B
13B
13B
13B
13B
13B
13B
13B
13B
13C
13C
13C
13C
13C
13C
13C
13C
13C
13C
13C
13C
13C
13C
13C
13C
13C
13C
14B
14B
14B
14B
14B
14B
14B
14B
14B
14B
14B
14B
14B
14B
14B
14B
14B
14B
14B
14B
14B
Animal
1
2
3
4
5
1
2
3
4
1
2
3
1
2
3
4
5
6
1
2
3
4
5
6
1
2
3
1
2
3
4
5
6
7
1
2
3
4
5
6
7
1
2
3
4
5
6
1
Rep
C
C
C
C
D
D
D
D
D
A
A
A
B
B
B
B
B
B
C
C
C
C
C
C
D
D
D
A
A
A
A
A
A
A
B
B
B
B
B
B
B
,C
C
C
C
C
C
D
Length
4.581
3.388
4.079
3.308
3.006
2.766
3.970
3.635
3.701
3.968
3.553
3.460
3.364
3.290
3.905
4.138
3.759
3.977
2.940
3.448
2.851
4.643
3.565
2.737
3.478
3.565
3.451
3.092
3.693
3.503
3.235
4.714
3.271
3.024
4.211
3.429
4.274
4.482
3.786
4.092
5.723
3.542
3.408
3.780
2.872
4.452
4.036
3.934
                                                1.47
-------
Appendix 1.5.  Amphipod length data for the 2nd set of sediment samples (continued).
Sample
14B
14B
14B
14B
14B
14B
14C
14C
14C
14C
14C
14C
14C
14C
14C
14C
14C
14C
14C
14C
14C
14C
14C
14C
14C
18B
18B
18B
18B
18B
18B
18B
18B
18B
18B
18B
18B
18B
18B
18B
18B
18B
18B
18B
18B
18B
18B
18B
Animal
2
3
4
5
6
7
1
2
3
4
5
6
1
2
3
4
5
6
1
2
3
4
1
2
3
1
2
3
4
5
6
7
1
2
3
4
5
6
7
1
2
3
4
5
6
7
1
2
Rep
D
D
D
D
D
D
A
A
A
A
A
A
B
B
B
B
B
B
C
C
C
C
D
D
D
A
A
A
A
A
A
A
B
B
B
B
B
B
B
C
C
C
C
C
C
C
D
D
Length
3.173
3.756
3.863
4.821
4.036
3.863
3.434
3.066
4.009
3.268
4.053
3.426
2.568
3.009
2.640
2.631
3.765
4.054
3.527
4.161
3.134
3.946
3.934
3.845
4.077
3.816
3.375
2.355
4.167
3.978
3.000
2.559
4.803
3.636
2.898
3.681
3.738
4.503
4.020
3.618
2.664
3.711
3.579
3.570
2.946
2.928
4.065
4.434
Sample
18B
18B
18B
18C
18C
18C
18C
18C
18C
18C
18C
18C
18C
18C
18C
18C
18C
18C
18C
18C
18C
18C
18C
18C
18C
18C
18C
18C
19B
19B
19B
19B
19B
19B
19B
19B
19B
19B
19B
19B
19B
19B
19B
19B
19B
19B
19B
19B
Animal
3
4
5
1
2
3
4
5
6
7
8
9
1
2
3
4
5
1
2
3
4
5
6
7
8
9
10
1
1
2
3
4
5
6
7
1
2
3
4
5
6
7
8
9
1
2
3
4
Rep
D
D
D
A
A
A
A
A
A
A
A
A
B
B
B
B
B
C
C
C
C
C
C
C
C
C
C
D
A
A
A
A
A
A
A
B
B
B
B
B
B
B
B
B
C
C
C
C
Length
3.534
3.618
3.693
3.608
3.310
3.841
2.696
3.412
4.169
3.793
3.552
2.767
2.857
3.823
4.601
3.626
3.304
3.444
3.578
4.178
2.994
3.414
3.868
2.896
3.728
3.155
3.364
3.904
4.054
3.348
3.872
3.339
3.304
3.167
3.146
3.348
4.214
2.450
3.455
2.938
3.923
3.506
3.726
3.173
3.774
3.851
3.378
3.295
                                               1.48
-------
Appendix 1.5.  Amphipod length data for the 2nd set of sediment samples (continued).
Sample
19B
19B
19B
19B
19B
19B
19B
19B
19B
19B
19B
19B
19B
19B
19C
19C
19C
19C
19C
19C
19C
19C
19C
19C
19C
19C
19C
19C
19C
19C
19C
19C
19C
19C
19C
19C
19C
19C
19C
19C
19C
19C
20B
20B
20B
20B
20B
20B
Animal
5
6
7
8
1
2
3
4
5
6
7
8
9
10
1
2
3
4
5
6
7
8
1
2
3
4
5
6
1
2
3
4
5
6
7
8
1
2
3
4
5
6
1
2
3
4
5
6
Rep
C
C
C
C
D
D
D
D
D
D
D
D
D
D
A
A
A
A
A
A
A
A
B
B
B
B
B
B
C
C
C
C
C
C
C
C
D
D
D
D
D
D
A
A
A
A
A
A
Length
2.818
3.711
2.970
3.205
3.009
3.304
3.319
3.015
2.774
3.089
3.545
2.616
2.836
2.917
3.893
3.792
3.238
2.929
3.581
2.515
3.512
4.137
3.631
3.777
2.997
3.176
3.875
2.979
3.646
3.256
3.435
3.955
3.438
3.307
3.149
3.720
2.920
3.845
3.366
3.494
3.173
3.485
3.630
3.580
4.004
3.711
3.834
3.884
Sample
20B
20B
20B
20B
20B
20B
20B
20B
20B
20B
20B
20B
20B
20B
20B
20B
20B
20B
20B
20B
20B
20B
20B
20B
20B
20B
20C
20C
20C
20C
20C
20C
20C
20C
20C
20C
20C
20C
20C
20C
20C
20C
20C
20C
20C
20C
20C
20C
Animal
7
8
1
2
3
4
5
6
7
8
9
10
1
2
3
4
5
6
7
8
1
2
3
4
5
6
1
2
3
4
5
6
7
8
9
1
2
3
4
5
6
7
8
9
1
2
3
4
Rep
A
A
B
B
B
B
B
B
B
B
B
B
C
C
C
C
C
C
C
C
D
D
D
D
D
D
A
A
A
A
A
A
A
A
A
B
B
B
B
B
B
B
B
B
C
C
C
C
Length
3.454
2.940
3.433
3.663
3.577
3.837
3.269
3.317
3.768
3.281
4.132
3.995
3.122
2.510
2.854
4.183
3.155
3.442
3.380
3.353
2.582
2.872
2.883
3.433
3.738
3.030
3.669
3.807
3.454
4.138
3.403
3.750
3.634
3.030
3.466
3.370
3.379
2.794
3.466
3.364
3.547
2.749
3.484
3.457
3.472
2.994
3.374
3.320
                                                1.49
-------
Appendix 1.5.  Amphipod length data for the 2nd set of sediment samples (continued).
Sample
20C
20C
20C
20C
20C
20C
20C
20C
20C
20C
20C
20C
20C
20C
20C
22B
22B
22B
22B
22B
22B
22B
22B
22B
22B
22B
22B
22B
22B
22B
22B
22B
22B
22B
22B
22B
22B
22B
22B
22B
22B
22B
22B
22B
22B
22B
22B
22B
Animal
5
6
7
8
9
1
2
3
4
5
6
7
8
9
10
1
2
3
4
5
6
7
8
9
10
1
2
3
4
5
6
7
8
9
1
2
3
4
5
6
7
8
9
1
2
3
4
5
Rep
C
C
C
C
C
D
D
D
D
D
D
D
D
D
D
A
A
A
A
A
A
A
A
A
A
B
B
B
B
B
B
B
B
B
C
C
C
C
C
C
C
C
C
D
D
D
D
D
Length
3.350
3.418
3.143
3.021
3.780
2.979
2.964
3.230
3.185
3.388
2.522
3.182
3.015
3.221
2.564
3.586
3.708
3.786
4.004
3.063
3.550
3.350
4.121
3.415
4.778
5.253
4.351
3.666
3.732
3.281
3.173
3.885
3.290
2.848
3.188
3.639
4.760
3.726
4.527
4.258
4.978
2.934
3.729
3.595
3.753
3.556
3.917
3.636
Sample
22B
22C
22C
22C
22C
22C
22C
22C
22C
22C
22C
22C
22C
22C
22C
22C
22C
22C
22C
22C
22C
22C
22C
22C
24B
24B
24B
24B
24B
24B
24B
24B
24B
24B
24B
24B
24B
24B
24B
24B
24B
24B
24B
24B
24B
24B
24B
24B
Animal
6
1
2
3
1
2
3
4
5
6
7
8
1
2
3
4
5
1
2
3
4
5
6
7
1
2
3
4
5
6.
7
8
1
2
3
4
5
6
7
8
9
1
2
3
4
5
6
7
Rep
D
A
A
A
B
B
B
B
B
B
B
B
C
C
C
C
C
D
D
D
D
D
D
D
A
A
A
A
A
A
A
A
B
B
B
B
B
B
B
B
B
C
C
C
C
C
C
C
Length
3.873
3.914
4.025
4.679
3.335
3.813
3.126
3.063
3.598
3.601
2.674
3.935
3.457
3.998
3.427
2.755
3.391
3.610
3.227
4.144
3.436
4.856
3.311
4.252
3.925
2.958
3.886
3.617
3.533
3.781
3.434
4.302
2.263
3.165
3.820
4.159
4.108
3.536
4.054
3.599
3.329
3.293
4.045
4.362
2.931
3.012
3.707
3.922
                                               1.50
-------
Appendix 1.5.  Amphipod length data for the 2nd set of sediment samples (continued).
Sample
24B
24B
24B
24B
24B
24B
24B
24B
24B
24B
24B
24C
24C
24C
24C
24C
24C
24C
24C
24C
24C
24C
24C
24C
24C
24C
24C
24C
24C
24C
24C
24C
24C
24C
24C
SCB
SCB
SCB
SCB
SCB
SCB
SCB
SCB
SCB
SCB
SCB
SCB
SCB
Animal
8
9
1
2
3
4
5
6
7
8
9
1
2
3
4
5
6
7
8
1
2
3
4
5
6
1
2
3
4
5
6
1
2
3
4
1
2
3
4
1
2
3
4
5
1
2
3
4
Rep
C
C
D
D
D
D
D
D
D
D
D
A
A
A
A
A
A
A
A
B
B
B
B
B
B
C
C
C
C
C
C
D
D
D
D
A
A
A
A
B
B
B
B
B
C
C
C
C
Length
3.189
3.257
3.862
3.877
3.563
3.931
3.054
3.060
3.653
3.722
4.311
3.877
3.750
4.007
3.490
2.985
3.414
3.481
3.980
2.822
3.820
3.841
4.542
3.611
4.836
3.157
3.859
3.505
3.832
3.756
3.139
4.001
5.571
3.387
4.083
3.794
3.490
3.758
3.171
3.748
3.736
2.994
3.239
3.393
3.325
2.933
3.319
3.583
Sample
SCB
SCB
SCB
SCB
SCB
SCB
SCB
SCB
SCB
sec
sec
sec
sec
sec
sec
sec
sec
sec
sec
sec
sec
sec
sec
sec
sec
sec
sec
sec
sec
sec
sec
sec
sec
sec
sec
sec
sec
sec
sec
sec
sec
sec
sec
sec
FLOR
FLOR
FLOR
FLOR
Animal
5
6
7
1
2
3
4
5
6
1
2
3
4
5
6
7
8
1
2
3
4
5
6
7
8
1
2
3
4
5
6
7
8
9
10
1
2
3
4
5
6
7
8
9
1
2
3
4
Rep
C
C
C
D
D
D
D
D
D
A
A
A
A
A
A
A
A
B
B
B
B
B
B
B
B
C
C
C
C
C
C
C
C
C
C
D
D
D
D
D
D
D
D
D
A
A
A
A
Length
2.784
2.651
3.661
3.605
3.708
4.795
3.343
3.352
2.955
3.373
2.286
2.506
2.973
2.678
3.051
3.331
3.238
3.259
2.955
3.346
3.563
3.518
2.952
3.111
2.961
2.789
2.584
2.732
2.753
2.937
2.861
2.855
2.922
3.352
2.404
2.725
3.275
3.359
3.741
3.648
2.537
2.523
3.143
3.648
2.334
2.522
2.256
2.561
                                                1.51
-------
Appendix 1.5.  Amphipod length data for the 2nd set of sediment samples (continued).
Sample
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
Animal
5
6
7
8
9
1
2
3
4
5
6
7
8
9
10
1
2
3
Rep
A
A
A
A
A
B
B
B
B
B
B
B
B
B
B
C
C
C
Length
2.624
2.546
1.743
1.960
4.070
2.394
1.978
2.913
2.949
2.650
2.850
2.901
3.436
2.268
2.531
3.320
2.328
3.230
Sample
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR
FLOR


Animal
4
5
6
7
8
1
2
3
4
5
6
7
8
9
10
11


Rep
C
C
C
C
C
D
D
D
D
D
D
D
D
D
D
D


Length
3.606
2.444
2.916
2.866
2.761
2.113
1.909
2.576
1.790
2.283
2.360
2.522
3.347
2.668
2.059
1.981


                                               1.52
-------
Appendix 1.6.  Amphipod maturation and survival data for the 1st set of samples. Replication (Rep), number of
amphipods recovered (Recov), and number of males recovered (Males).
Sample
01B
01B
01B
01B
QIC
QIC
01C
01C
03B
03B
03B
03B
05B
05B
05B
05B
05C
05C
05C
05C
08B
08B
08B
08B
08C
08C
08C
08C
10B
10B
10B
10B
IOC
IOC
IOC
IOC
11B
11B
11B
11B
11C
11C
11C
11C
12B
12B
12B
1'2B
Rep
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
Recov
8
10
9
9
5
6
7
6
4
8
11
8
9
9
7
6
7
6
9
9
10
10
8
11
9
6
8
9
10
9
7
10
6
6
9
8
7
9
9
9
7
5
6
5
9
9
7
7
Males
3
3
5
3
1
0
1
2
1
3
6
5
2
6
4
2
1
1
3
2
5
2
3
6
3
3
1
3
5
2
6
0
3
1
3
3
2
3
3
7
1
4
1
1
3
3
2
3
Sample
12C
12C
12C
12C
15B
15B
15B
15B
15C
15C
15C
15C
16B
16B
16B
16B
16C
16C
16C
16C
2 IB
2 IB
21B
21B
21C
21C
21C
21C
25B
25B
25B
25B
25C
25C
25C
25C
26B
26B
26B
26B
26C
26C
26C
26C
FLB
FLB
FLB
FLB
Rep
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
Recov
9
10
8
10
9
8
9
10
7
7
8
9
9
5
6
6
6
9
9
11
10
9
9
10
9
8
8
10
6
7
5
4
8
2
10
9
10
8
9
8
10
7
6
10
6
8
7
9
Males
4
2
2
4
3
6
6
3
1
2
3
5
2
2
3
3
1
1
5
4
5
4
4
7
5
3
5
5
1
2
0
2
2
1
1
3
5
2
5
3
7
2
4
3
2
5
2
2
                                                1.53
-------
Appendix 1.7. Amphipod maturation and survival data for the 2nd set of samples. Replication (Rep),  number of
amphipods recovered (Recov), and number of males recovered (Males).
Sample
02B
02B
02B
02B
02C
02C
02C
02C
04B
04B
04B
04B
04C
04C
04C
04C
06B
06B
06B
06B
06C
06C
06C
06C
07B
07B
07B
07B
07C
07C
07C
07G
09B
09B
09B
09B
09C
09C
09C
09C
13B
13B
13B
13B
13C
13C
13C
13C
Rep
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
Recov
8
8
4
8
5
10
7
9
4
8
9
14
0
11
8
7
7
2
10
6
9
8
8
8
10
10
10
10
9
10
9
11
7
5
11
8
4
8
16
4
2
2
5
4
3
6
6
3
Males
4
2
1
2
2
4
2
6
3
2
1
5
0
1
2
1
1
1
1
2
5
4
4
5
2
7
4
4
3
3
3
5
4
2
3
4
2
3
7
0
0
1
0
1
2
2
2
2
Sample
14B
14B
14B
14B
14C
14C
14C
14C
18B
18B
18B
18B
18C
18C
18C
18C
19B
19B
19B
19B
19C
19C
19C
19C
20B
20B
20B
20B
20C
20C
20C
20C
22B
22B
22B
22B
22C
22C
22C
22C
24B
24B
24B
24B
24C
24C
24C
24C
Rep
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
Recov
7
7
6
6
6
6
4
3
7
7
7
5
9
5
10
1
7
9
8
10
8
6
8
6
8
10
8
6
9
9
9
10
10
9
9
6
3
8
5
7
8
9
9
9
8
5
6
4
Males
1
2
3
2
1
1
3
2
2
4
1
5
3
1
3
0
3
7
0
4
3
3
2
1
2
1
1
0
4
0
4
2
2
2
2
2
2
4
0
3
3
2
4
3
3
4
6
2
                                               1.54
-------
Appendix 1.7. (Continued)
Sample
Rep    Recov   Males
Sample
Rep    Recov   Males
SCB
SCB
SCB
SCB
sec
sec
sec
sec
1
2
3
4
1
2
3
4
4
5
7
6
8
8
10
9
0
0
1
2
2
3
2
4
FLC
FLC
FLC
FLC




1
2
3
4




9
10
8
11




1
0
1
0




                                              1.55
-------
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-------
Appendix 1.9.  Concentrations (ng/g dry weight) of organochlorine pesticides (OCs) in Upper Mississippi River
sediments.
POOL
IB
1C
2B
2C
3B
4B
4C
SB
5C
6B
6C
7B
7C
8B
8C
9B
9C
10B
IOC
11B
11C
12B
12C
13B
13C
14B
14C
15B
15C
16B
16C
18B
18C
19B
19C
20B
20C
21B
21C
Chlordane
0.001
ND
0.001
ND
ND
0.002
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
0.001
ND
ND
ND
ND
ND
ND
ND
ND
ND
0.002
ND
Dieldrin
ND1
ND
0.0003
ND
0.0003
0.0005
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
0.0001
ND
0.0002
ND
0.0002
ND
0.0002
ND
0.0003
ND
ND
ND
0.0004
ND
DDE
0.0004
ND
ND
0.0520
0.0011
0.0010
ND
0.0001
ND
0.0001
ND
0.0003
ND
0.0002
ND
0.0003
ND
0.0002
ND
0.0002
ND
0.0003
ND
0.0002
ND
0.0001
ND
0.0004
ND
0.0004
ND
0.0003
ND
0.0001
ND
0.0001
ND
0.0004
ND
ODD
0.0005
ND
0.0016
0.0790
0.0038
0.00.19
ND
0.0001
ND
0.0003
ND
0.0010
ND
0.0004
ND
0.0010
ND
0.0001
ND
0.0004
ND
0.0006
ND
0.0004
ND
0.0002
ND
0.0005
ND
0.0004
ND
0.0006
ND
0.0002
ND
0.0002
ND
0.0008
ND
DDT
ND
ND
0.0002
ND
0.0002
ND
ND
ND
ND
ND
ND
0.0001
ND
ND
ND
0.0001
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
0.0018
ND
ND
ND
ND
ND
ND
ND
ND
ND
0.0003
ND
                                                1.58
-------
Appendix 1.9. Concentrations of organochlorine pesticides (OCs) in Upper Mississippi River sediments (cont.).
POOL
Chlordane
Dieldrin
DDE
ODD
DDT
22B
22C
24B
24C
25B
25C
26B
26C
SCB
sec
ND
ND
0.0010
ND
0.0010
ND
ND
ND
ND
ND
0.0003
ND
0.0004
ND
0.0006
ND
0.0007
ND
ND
ND
0.0001
ND
0.0001
ND
0.0005
ND
0.0005
ND
0.0007
0.0780
0.0001
ND
0.0001
ND
0.0005
ND
0.001
ND
0.0004
0.0780
ND
ND
ND
ND
ND
ND
ND
ND
0.0001
ND
ND = Not detected
                                                1.59
-------
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-------
 Chapter 2: An Evaluation of Bioaccumulation of Contaminants from Sediments from the Upper
 Mississippi River Using Field-collected Oligochaetes and Laboratory-exposed Lumbriculus
 variegatus

 Brunson, E.L., Canfield, T.J., Dwyer, F.J., Ingersoll, C.G., and Kemble, N.E.

 Introduction

 Over the past 10 years, a variety of methods have been described for evaluating the toxicity of
 sediment-associated contaminants with benthic invertebrates. However, only a limited number of
 methods are currently available for assessing bioaccumulation of contaminants from field-
 collected or laboratory spiked sediments (Ingersoll et al 1995).  Standard guides have been
 published for conducting 28-d bioaccumulation tests with the oligochaete Lumbriculus variegatus
 including determination of bioaccumulation kinetics  for different compound classes (USEPA,
 1994; ASTM 1996).   Lumbriculus variegatus was selected for use in sediment bioaccumulation
 testing in the present study of upper Mississippi River (UMR) for six reasons: (1) ease of culture
 and handling, (2) known chemical exposure history,  (3) adequate tissue mass for chemical
 analyses, (4) tolerance of a wide range of sediment physico-chemical characteristics, (5) low
 sensitivity to contaminants associated with sediment, and (6) amenability to long-term exposures
 without feeding. Other organisms do not meet many of these selection criteria including mollusks
 (valve closure), midges (short-life cycle), mayflies (difficult to culture), amphipods (small tissue
 mass, too sensitive), cladocerans and fish (not in direct contact with sediment).
    Several investigators have conducted bioaccumulation studies in the laboratory with L.
 variegatus using either field-collected or laboratory-spiked sediments (Schuytema et al  1988;
Nebekerefa/. 1989; Ankley et al. 1991; Called al.,  1991; Carlson et al.  1991; Ankley etal. 1993;
Kukkonen and Landrum 1994).  However, only one  previous study has compared results of
laboratory bioaccumulation  studies conducted with L. variegatus to residues from synoptically-
 collected field populations of oligochaetes (Ankley et al.  1992).  The author reported good
 agreement between concentrations of polychlorinated biphenyls in the laboratory and field
organisms,  particularly for PCB  congeners with K,,w  values <7. This suggests that laboratory
exposures longer than 28 d may be required to reach equilibrium for super-hydrophobic
chemicals.
    The United States Geological Survey (USGS) has been monitoring the Upper Mississippi
River since 1987 to document the fate and transport  of contaminated sediments (Moody and
Meade 1995). Concern with the redistribution of these contaminated sediments arose after the
flood of 1993. This project is designed to  evaluate the current status of sediments in the UMR
and is one chapter in a series designed to assess the extent of sediment contamination in
navigational pools of the river.  The overall project consists of the following assessments: (1)
measuring concentrations of contaminants  in sediments of the UMR (Moody et al.  1996), (2)
toxicity testing with sediments collected from the river (Chapter 1), (3) analysis of benthic
community structure (Chapter 3), and (4) bioaccumulation of sediment associated contaminants
(the present chapter).  The present study had two objectives: (1) to assess the bioaccumulation of
contaminants from UMR sediments using L. variegatus and (2) to compare bioaccumulation in
                                           2.1
-------
these laboratory-exposed oligochaetes to oligochaetes collected from the field.

Materials and Methods

Sample Collection

Sediment samples and native oligochaetes were collected from 23 navigational pools on the UMR
and from the Saint Croix River ("C" samples described in Chapter 1). Sample stations were
selected based on the potential of oligochaetes or fine grained sediment.   For each C sample, 35-
to 80-L of sediment (6 to 25 grabs) were collected with a stainless steel Ponar grab sampler
(Wildlife Supply Company, Saginaw, MI). All grabs from a station within a pool were collected
within a 5-meter radius and combined in a 114-L high-density polyethylene (HOPE) container.
The composited sample was homogenized on board the research ship Acadiana using an electric
drill and a stainless steel auger. Once homogenized, the following subsamples were removed: (1)
three separate 250 ml subsamples for organic chemistry, metals/acid-volatile sulfides, and total
organic carbon/particle size (Chapter 1), (2) one 2-L subsample for benthic invertebrates (Chapter
3), and (3) one  10-L subsample for laboratory toxicity (Chapter 1) and bioaccumulation testing.
Sediment samples were stored at 4° C until used in laboratory exposures or physical/chemical
analysis.
    The remainder of the composited C sample of sediment was rinsed on ship through a Wildco
wash bucket ( U.S. Standard sieve size #30, 600 (jm opening). The material captured by the
wash bucket was transferred to a HOPE tub along with river water.  After all the sediment was
sieved, native oligochaetes were isolated from the detritus.  These oligochaetes from each sample
were placed in a HDPE jar containing aerated river water and held for 24 hours to depurate gut
contents.  After the 24-hour elimination period, dead oligochaetes were discarded. The remaining
oligochaetes were rinsed, blotted dry, weighed, transferred to clean glass jars, and frozen at -22°C
until analyzed for chemical contaminants.  Weights of native oligochaete samples selected for
analysis ranged  from  0.34g (Pool 4) to 9.8g (Pool 9)

Laboratory  Testing

Lwnbriculus variegatus were exposed hi the laboratory to sediment following methods described
in USEPA (1994) and ASTM (1996).  Sediment from  13 of the 23  sampled pools were used in
these laboratory exposures.  Samples were chosen for testing on the basis of sufficient mass of
field-collected oligochaetes for chemical analyses (or the previously documented presence  of
PCBs for pool 4 in lower Lake Pepin; e.g. Rostad et al, 1996 ). Oligochaetes were mass cultured
in the laboratory following methods similar to those described in USEPA (1994) using 75-L glass
aquarium  containing  50 L of well water (hardness 290 mg/L as CaCO3, alkalinity 255 mg/L as
CaCO3, pH  7.8).  Each aquaria received about  27 volume, additions (about 1.5 L/minute) of well
water daily. The culture water was aerated and maintained at 23°C.  Pre-soaked, shredded brown
paper towels were used as substrate. Cultures were fed Tetramin flake fish food twice weekly ad
libitum.
     Exposures of oligochaetes in the laboratory were conducted for 28 days in 4-L glass Pyrex
                                           2.2
-------
beakers containing 1 L of sediment and 3 L of overlying water.  Four replicate chambers were
tested for each of the thirteen sediment samples. Reconstituted fresh water (hardness 90 to 96
mg/L as CaCO3, alkalinity 60 to 70 mg/L as CaCO3; USEPA 1994) was used as the overlying
water. Each beaker was calibrated to 4-L using a glass standpipe that exited through the beaker
wall and was held in place with a silicon stopper.  Test chambers received 2 volume additions (6 L
± 10%) of overlying water per day.  Water was delivered using a modified Mount and Brungs
diluter system (Ingersoll and Nelson  1990). An in-line flow splitter was attached to each delivery
line to split the water flow evenly to each of four test chambers. The splitters were constructed of
1/4 inch PVC pipe with four silicone stoppers and 14-gauge stainless steel hypodermic needles
with the points and connector ends cut off the needles (Figure 2.1). Glass stands were used to
support the splitters keeping them level to maintain a constant volume delivery to each exposure
chamber. Chambers were held in a temperature-controlled waterbath (23±1°C) on a 16:8
lightdark photoperiod at about 500 lux. Oligochaetes were not fed during the sediment
exposure.
    Sediment and overlying water were placed in the chambers the day before adding organisms
(Day -1). Sediments were first homogenized with a hand-held electric drill and stainless steel
auger before being placed into the test beakers. One-L of sediment was transferred into each
chamber using a plastic spoon.  Overlying water was poured into the beakers through a piece of
fine-mesh Nitex® material to minimize suspension of the sediment. Water delivery started after
chambers were placed in the waterbath.
    Twenty-four hours before stocking the test (Day -1) oligochaetes were removed from the
culture with  a fine-mesh nylon aquarium net,  placed in beakers containing well water, and rinsed
to remove excess toweling and debris. Beakers containing the oligochaetes were then placed in a
waterbath and aerated.  With substrate absent, the L. variegatus formed tight masses or clumps in
the beakers which was helpful during transfer of organisms into the exposure chambers.
    Oligochaetes were acclimated to the test water by removing half of the water in  each beaker
and replacing it with temperature-acclimated test water. Two hours later this process was
repeated. After another two hours, the L. variegatus were combined into a glass pan and rinsed
with well water to break up the masses of worms and remove any remaining debris.  With the
mass of worms disturbed, oligochaetes were grouped together with a stainless steel dental pick
and allowed  to form small clumps of about 1  g. The clumps of oligochaetes were removed from
the pan with the dental pick, touched against the rim of the pan to remove excess water, and
placed on a tared weigh boat. About 2.6 g unblotted oligochaetes were transferred  to each test
chamber containing sediment and overlying water .  Using this approach, the 2.6 g of unblotted
oligochaetes represents about 2 g of blotted oligochaetes or about 200 organisms.
    General conditions of the exposure system and behavior were evaluated daily. Dissolved
oxygen and conductivity of the overlying water were measured weekly in all chambers. Total
hardness (as  CaCO3), pH, alkalinity (as CaCO3), and total ammonia of overlying water were
measured at  the beginning and end of the test. Overlying water pH, alkalinity, total hardness,
conductivity and total ammonia measurements were similar among all stations and inflowing test
water (Appendix 2.1). Dissolved oxygen measurements were at or above acceptable levels
(>40% of saturation; ASTM 1996) in all treatments throughout the study (Appendix 2.1).
Ranges of mean water quality for each parameter were as follows: pH 7.7 to 7.9; alkalinity as
                                          2.3
-------
CaCO3 61 to 67 mg/L; total hardness as CaCO3104 to 110 mg/L; conductivity 342 to 350 |uS
@25°C; total ammonia 0.1 to 0.4 mg/L; and calculated unionized ammonia 0.0028 to 0.0094
mg/L.
    On Day 28 of the exposure, L. variegatus were isolated from each test chamber by washing
the sediment through No. 18 (1.0 mm opening) followed by No. 50 (300 urn opening) U.S.
standard stainless steel sieves. The contents of each sieve was rinsed into several clear glass pans
and all oligochaetes were removed. Lumbriculus variegatus were separated from native
oligochaetes based on behavior (native oligochaetes tended to form a tight, spring-like coil,
whereas L. variegatus would not (USEPA 1994)).  Once isolated, all L. variegatus from a
chamber were cleaned of any remaining debris and held for 24 h in 1-L water-only chambers to
allow them to clear their gut contents. The L. variegatus were then isolated, cleaned of any
remaining debris, and transferred to a tared weigh boat.  Samples were then blotted, weighed,
placed in glass jars, and stored at -22 °C pending chemical analysis for contaminants. Weights of
laboratory-exposed oligochaete samples ranged from 1.3g to 3.0g.

Chemical Analyses

Sediment physical characteristics included the following: (1) sediment particle size, (2) total
organic carbon, (3) inorganic carbon and (4) percent water.  Sediment chemical parameters
included: (1) organochlorine pesticides (OCs), (2) polychlorinated biphenyls (PCB), (3) select
aliphatic and polynuclear aromatic hydrocarbons (PAH), (4) simultaneously extracted metals
(SEM), (5) acid volatile sulfide (AVS), and  (6) total metals. See Chapter 1 for additional
information on methods and results of chemical and physical characterizations of the sediments.
    Concentrations of metals and organochlorines in sediment samples were low (Chapter 1).
Therefore, replicate tissue samples from the laboratory exposures were combined for
organochlorine pesticide/PCB analyses and metals were not analyzed because of limited sample
mass.  Tissues were analyzed by Geochemical and Environmental Research Group at Texas
A&M University, College Station, Texas for the following: (1) organochlorine pesticides (OCs),
(2) polychlorinated biphenyls (PCBs), (3) select aliphatic and polynuclear aromatic hydrocarbons
(PAHs), and (4) percent lipid. Prior to analysis, tissue samples were homogenized and extracted
using a Teckmar Tissumizer, sodium sulfate, and methylene chloride (MacLeod et al. 1985; Wade
et al 1988; Brooks et al. 1989).  Tissue extracts were split into two fractions: one fraction was
used to measure percent lipid and the second fraction was used for measuring PAHs, OCs, and
PCBs. Extracts for chemical analyses were purified using absorption chromatography to isolate
the aliphatic fraction and the PAH/OC/PCB fraction. Lipid interference in the PAH/OC/PCB
fraction was eliminated with further purification using HPLC.  The quantitative analyses were
performed by capillary gas chromatography (CGC)  with electron capture detector for OCs and
PCBs and a mass spectrometer detector in the SIM mode for PAHs (Wade et al., 1988).  Percent
lipids were calculated on a wet-weight basis. A 20-ml aliquot of the total extract was filtered,
concentrated to 1 ml, and weighed.  A 100-ul subsample was then removed, evaporated to
dryness, and weighed.  Percent lipid was calculated using the weight of the dried subsample and
the concentrated sample. Tissue residue data are presented in Appendix 2.2 and Appendix 2.3.
Sediment data are shown in Table  1.1, and Tables 1.3 to 1.5 in Chapter 1.
                                           2.4
-------
    Average percent spike recovery for twenty-two OCs and was 88% (n=4). Beta BHC had
the smallest average spike recovery (53%) while oxychlordane had the greatest average spike
recovery (104%).  Individual OC concentrations were often below minimum detectable limits so
duplicate analyses were evaluated only for total PCBs. The average duplicate coefficient of
variation was 26% (range 0.7 to 61%,  n=4).  Average percent spike recovery for PAH
compounds was 96% (25 compounds, n=4). L123(c,d)pyrene had the smallest average percent
recovery (81%) while 1-methylnaphthalene had the greatest average percent recovery (110%).
The average duplicate coefficient of variation was 21% (34 possible compounds, n=l-4).
Average duplicate coefficient of variation ranged from 1% for cl-phenanthracene to 79% for
benzo-a-pyrene.
    In addition to the laboratory-exposed and field-collected oligochaetes, three samples of
oligochaetes from laboratory cultures were collected at the beginning of the exposure for analysis
contaminants.  Two of the three samples had detectable concentrations of PAHs and total PCBs
however, the concentrations were generally less than those of oligochaetes exposed to or
collected from the UMR sediments.  For some unexplained reason, total PCB (1.3 ug/g dry wt)
and some PAH concentrations (up to 0.25 ug/g dry wt.) in one of those three samples was
similar to oligochaetes exposed during the test.

Results and Discussion

General Trends

Individual organochlorine pesticides (OC) were generally below the detection limits (ranging from
0.0007 to.0.0217 ng/g wet weight) for oligochaetes from both field-collection and laboratory-
exposed animals (Appendix 2.2). For the 13 field collected samples and 22 OCs measured,
individual OCs were identified a total of 6 times.  The greatest individual OC concentration was
0.009 ng/g (wet weight) for dieldrin from oligochaetes collected from Pool 22.  As was the case
with the field-collected oligochaetes, tissue concentrations of individual OCs were often below the
detection limit for many of the laboratory-exposed oligochaetes. All oligochaete samples had at
least one OC concentration above background (Pool 13 and Pool 16; 4,4'DDE); however, no
sample had more than 6 OCs detected (Pool 11 and 14; gamma-chlordane, alpha-chlordane,
aldrin, dieldrin, 4,4'DDE, 4,4T)DD). The greatest individual OC concentration was 0.013 ^ig/g
(wet weight) for 4,4 DDE for oligochaetes exposed in the laboratory to sediment collected from
Pool 4. Also, 4,4 DDE was the most frequently measured OC (12 samples) with concentrations
ranging from 0.0021 to 0.013 ug/g (wet weight).
    Total PCBs were the only chlorinated organic compound detected in all field-collected and
laboratory-exposed oligochaetes. Concentrations ranged from 0.045 jig/g (wet weight - pool 13)
to 0.697 ug/g (wet weight - Pool 4). The geometric mean for total PCBs measured in
oligochaetes exposed to the sediment samples was 0.129 jig/g
    Field-collected and laboratory-exposed oligochaete samples were analyzed for 44 PAH
isomers. Field collected oligochaetes from Pool 4 had the fewest number of PAHs (14) while
Pool 19 had the most (36).  Only 16 PAH isomers  (about 40% of those analyzed for) had
detectable concentrations (detection limits from 0.0217 to 0.0024 \ig/g wet weight) in 7 of the 13
                                          2.5
-------
Pools for both the field-collected and laboratory-exposed oligochaetes (for the laboratory
exposures, 2 of the 4 replicates had to exceed the detectable limit in order to be included in this
analysis).  Table 2.1 lists all compounds measured in tissues that met these selection criteria.
Figures 2.2 and 2.3 depict accumulation of total PAH in samples from laboratory-exposed or
field-collected oligochaetes for each UMR pool evaluated.  Concentrations of the 16 PAH
isomers were converted to  molar units, normalized to percent lipid, and summed.  Total PAH
from field-collected and laboratory-exposed oligochaetes, show a trend of decreasing
concentrations in the down river Pools (14 to 22). Field-collected oligochaetes from Pool 7 were
more contaminated than oligochaetes from the other pools. For the laboratory exposures,
oligochaetes exposed to sediments from Pool 4 were more  contaminated than oligochaetes
exposed to sediments from the other pools.  In general, perylene had the highest concentration of
any PAH from field-collected and laboratory-exposed oligochaetes.  This trend was greater for
laboratory exposed oligochaetes than for those collected from the field. Perylene concentrations
ranged from 0.056 to 0.53  ug/g (wet weight) in field collected oligochaetes and from 0.052 to
0.84 ug/g (wet weight) hi oligochaetes from laboratory exposures.
    Sediments and oligochaetes from the TJMR are relatively uncontaminated compared to other
locations we have evaluated using sediment toxicity tests (Ingersoll et al. 1996) or
bioaccumulation tests (sediments from Little Scioto River in Ohio, unpublished data).  Ingersoll et
al (1996) calculated sediment effect concentrations including Effects Range Medians (ERMs)
from 28-day sediment exposures with Hyalella azteca. An ERM is  defined as that concentration
of a material in sediment above which toxic effects are frequently or always observed or
predicted. In the current study, tissue concentrations of PAHs were generally greatest in samples
from Pool 4.  Two low molecular weight (LMW) PAHs (naphthalene and phenanthrene) and two
high molecular weight  (HMW) PAHs (pyrene and chrysene) were generally the PAHs of highest
concentration hi tissue samples from pool 4. The calculated sediment ERM concentrations (ug/g
dry weight) for those PAHs are; naphthalene - 0.097, phenanthrene - 0.345, pyrene - 0.347,  and
chrysene - 0.500.  The sediment concentrations (ug/g dry weight) from Pool 4 were; naphthalene
- 0.049, phenanthrene- 0.049, pyrene  - 0.245, and chrysene - 0.147. The sediment ERMs are
1.4 to 7 times greater than the highest concentrations of these PAHs in sediments from the current
study.  ERMs are not directly applicable to contaminant concentrations in tissues; however, tissue
concentrations hi UMR Pool 4 were more than two orders  of magnitude less than tissue
concentrations of oligochaetes exposed to sediments from the Little Scioto River. Collectively,
this information would indicate that  sediment and biota from the UMR is relatively
uncontaminated when compared to known contaminated sites previously evaluated by our
laboratory.

Detection of Compounds in Tissue vs. Sediment

Detection limits for tissue and sediment are usually different which creates difficulties in
interpreting bioaccumulation potential from relatively uncontaminated sediments.  In the UMR,
concentrations of PAHs and PCBs were detected hi both sediments and tissue samples 79% of the
time for the laboratory-exposed oligochaetes and 58% of the time for the field-collected
oligochaetes. PAHs and PCBs were not detected hi the sediments but were detected in
                                          2.6
-------
laboratory-exposed oligochaetes in 17% of the samples and in field-collected oligochaetes in 41%
of the  samples. PAHs and PCBs were detected in sediment samples but not in 3% of the samples
from laboratory-exposed oligochaetes and 1% of the samples of field-collected oligochaetes.
Although the detection limits for sediments and tissues met established guidelines (USEPA 1984),
detection limits for sediments may need to be decreased in order to better represent potentially
bioavailable compounds.

Laboratory to Field Comparisons

Tissue concentrations of naphthalenes were generally higher in field-collected oligochaetes than in
laboratory exposed oligochaetes (Figure 2.4). Naphthalenes are LMW PAHs with log Kow values
less than 4.5. PAHs with similar concentrations in both the laboratory-exposed and field-collected
oligochaetes included a similar number of HMW and LMW compounds (biphenyl, fluorene, 1-
methylphenanthrene, pyrene, fluoranthene, chrysene, and benzo(e)pyrene). Most of these
compounds are intermediate in molecular weight and log Kow (except for benzo(e)pyrene which
has the highest molecular weight and log Kow of all compounds included in Figure 2.4). PAHs
typically higher in the laboratory-exposed than in field-collected oligochaetes were primarily
HMW compounds ( benzo(a)anthracene, benzo[b(k)]fluoranthene, and perylene) with log Kows
greater than 5.1 (Figure 2.4 and 2.5).
     The ratio of tissue concentrations in laboratory-exposed oligochaetes to concentrations in
'field-collected oligochaetes were generally similar (Figure 2.5).  About 90%  of the corresponding
concentrations were within a factor of three between the laboratory-exposed and field collected
oligochaetes (represented by the crosshatched region in Figure 2.5).  However, there appears to
be a shift from field>lab to lab>field as the molecular weight of PAHs increases.  Concentrations
that differed by more than a factor of three were primarily LMW PAHs (naphthalene, 1-
methymaphthalene, 2-methylnaphthalene, 2,6-dimethylnaphthalene, fluorene, 1,6,7-
trimethylnaphthalene, phenanthrene, and 1-methylphenanthrene) and were usually elevated in the
field-collected oligochaetes compared to the laboratory-exposed oligochaetes. Ratios >3  in the
laboratory-exposed or field-collected oligochaetes were most frequently associated with a small
group of pools (Field > 3x lab in Pools 4,  12, 22; lab >3x field in Pool 7).
     Differences between tissue concentrations in the laboratory-exposed and field-collected
oligochaetes may have resulted from LMW PAHs being lost during the sampling of sediments.  A
second possibility for differences between the laboratory and field-exposed may be spatial
heterogeneity of contaminants in the sediments in the field. Other possible explanations could
include the rout of exposure. Exposure to contaminants in the field may occur through sediment,
food and overlying water while the route of exposure to oligochaetes in the laboratory was
sediment.  Species-specific differences in exposure between Lumbriculus variegatus and the
native oligochaetes may also contribute to the differential accumulation. For example,
concentrations of metals reportedly differ among taxa inhabiting the  same locations (Cain et al.
 1992).
                                            2.7
-------
 Biota-sediment Accumulation Factors

 Biota-sediment accumulation factors (BSAFs) were calculated by dividing the lipid-normalized
 tissue concentrations by the organic-carbon normalized sediment concentrations (USEPA 1994).
 Mean BSAFs for this study were only listed for compounds in which BSAF could be calculated
 for both laboratory-exposed and field-collected oligochaetes in at least seven of 13 pools (Table
 2.2). For laboratory-exposed oligochaetes, mean BSAFs ranged from 1.1 for benzo(a)anthracene
 to 5.3 for naphthalene. Mean BSAFs for field-collected oligochaetes, mean BSAFs ranged from
 0.5 for benzo(a)anthracene to 8.8 for naphthalene.  Individual sample BSAFs for naphthalene
 ranged from 1.6 to 10.1 in laboratory-exposed oligochaetes and 2.5 to 26.6 in field-collected
 oligochaetes. BSAFs for pyrene, benzo(a)anthracene, and benzo(b,k)fluoranthene were typically
 greater than BSAFs reported for marine organisms (Lee 1992).  BSAFs were also calculated
 using PCB homolog data reported in Ankley et al. (1992) for laboratory-exposed L. variegatus
 and field-collected oligochaetes (Figure 2.6).  BSAFs were similar between laboratory-exposed
 and field-collected oligochaetes in both Ankley et al (1992) and in the present study; however,
 BSAFs in the present study were  typically greater (0.5 to 8.8) than those from Ankley et al.
 (1992; 0.17 to 2.26).
     A theoretical value of 1.7 for BSAFs has been estimated based on partitioning of non-ionic
 organic compounds between sediment carbon and tissue lipids (McFarland and Clarke 1986).  A
 BSAF of less than 1.7 indicates less partitioning into lipids than predicted and a value greater than
 1.7 indicates more uptake than  can be explained by partitioning theory alone (Lee 1992). The
 majority of the BSAFs in Table 2.2 were within a range of about 0.5 to 2.6 suggesting the
 theoretical BSAF value of 1.7 could be used to predict these mean BSAFs with a fair amount of
 certainty.  However, mean BSAFs for naphthalene (8.8) and 2-methyl naphthalene (6.7) in the
 field-collected oligochaetes were elevated relative to a theoretical BSAF of 1.7.  Moreover,
 BSAFs for individual pools were as high as 10.1 for laboratory-exposed oligochaetes and 26.6 for
 field-collected oligochaetes. The  higher BSAFs in the field-collected oligochaetes may be the
 result of (1) exposure to contaminants in the overlying water, (2) spatial differences in sediment
 contamination (i.e., sediments were not sampled from a depth representative of the habitat of the
 oligochaetes), (3) increased error  in chemical determinations due to low concentration of
 contaminants in sediments, or (4)  taxonomic-specific differences in exposure. BSAFs
 substantially different from the theoretical value of 1.7 may also result when the system has not
 reached steady state (i.e., depletion or release of contaminants in pore water).

 Summary

 Contaminant concentrations were relatively low in sediments and tissues from the 13 UMR pools
 evaluated.  Only PAHs and total PCBs were frequently measured above detection limits. Most of
the concentrations of PAHs in UMR sedimgnt were similar to concentrations in  sediments
identified as non-toxic in amphipod toxicity tests from these previous studies. PAH
 concentrations in tissues of oligochaetes tested with highly contaminated samples from previous
 studies were up to 1000 times greater than tissue concentrations measured in the present study.
 Concentrations in laboratory exposed and field-collected oligochaetes for a compound from a
                                          2.8
-------
specific pool in the UMR were generally similar. About 90% of the paired PAH concentrations in
laboratory-exposed and field-collected oligochaetes were within a factor of three of one another.
With the detection limits used to analyze samples in the present study, contaminants were
detected in tissue samples more often than in the associated sediment samples.
Acknowledgments. We would like to thank the following individuals for their contributions to the
project: I.E. Greer, D.K. Hardesty, P.S. Haverland, C.E. Henke, E.K. Henry, P. A. Lovely, J.A.
Moody, D.S. Reussler, J.D. Soltvedt, J. Stevens, R.W. Walton, D.W. Whites, P.R. Heine, J.L.
Zajicek, D.C. Zumwalt, and the Crew of the Acadiana (Craig LeBoeuf and Pat Marmande).

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                                         2.11
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                              d    r-'
                                                                                                     5?
                                                                                                                 vo
                                                                                         in

                                                                                         es
                                                                                               p

                                                                                               55
                                                                                               m
                                                                                   es
                                                                                   ts
                                                                                                           cs
                                                                                         o
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                                                                                         o    gj
•<*•    ON

vo'    vo'
OO

in
**.      .
O    vo'
                                                                                   111
-------
Appendix 2.2. Tissue concentrations of organochlorine compounds measured in laboratory-
exposed and field-collected oligochaetes. All concentrations are on a wet-weight basis (ug/g).
Appendix 2.2 data can be obtained electronically from:
    anonymous ftp - ftp://ftp.msc.nbs.gov/pub/umr/umr.zip
    world wide web - http://www.msc.nbs.gov/pubs/umr.html
For problems with access to the above addresses please e-mail the Webmaster, Chris Henke, at
chenke@msc.nbs.gov or call 573-875-5399.
                                          2.21
-------
Appendix 2.3. Tissue concentrations of PAHs measured in laboratory-exposed and field-
collected oligochaetes. All concentrations are on a wet-weight basis (ug/g).  Appendix 2.3 data
can be obtained electronically from:
    anonymous ftp - ftp://ftp.msc.nbs.gov/pub/umr/umr.zip
    world wide web - http://www.msc.nbs.gov/pubs/umr.html
For problems with access to the above addresses please e-mail the Webmaster, Chris Henke, at
chenke@msc.nbs.gov or call 573-875-5399.
                                         2.22
-------
Appendix 2.4 Total accumulation (uMole/g lipid) of PAHs in Laboratory exposed (LMML) and
Field Collected (FMML) oligochaetes. Lipid-normalized concentrations (ug/g lipid) are given for
laboratory-exposed (LLCONC) and field-collected (FLCONC) oligochaetes.  Chemical numbers
(CHEM) correspond to those listed in Figure 2.3.
                  DBS
                        POOL
                               LLCONC
                                        FLCONC   HOLEWT
                                                       CHEM
                                                               LMML
                                                                        FMML
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
4
4
4
4
4
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
5
5
5
5
6
6
6
6
6
6
6
6
6
6
6
6
6
6
6
6
3.7278
1.3910
1 .3835
1.0671
0.0000
0.9915
0.0000
6.3817
7.0939
3.6973
3.0205
1.2794
2.5340
24.7897
4.4511
4.2986
2.2823
2.0964
1.5249
0.0000
0.0000
1 .4302
9.4287
1.1943
2.7115
2.7236
1.6731
48.6282
1.2731
3.6883
2.2485
2.0274
1.9501
3.1882
2.4097
3.9054
10.3210
2.1980
5.8612
5.0932
5.2747
2.5364
2.0751
13.9559
3.5580
26.0370
7.6111
12.0000
11.9630
13.5741
0.0000
9.4630
11.2222
10.9074
6.2778
4.9074
0.0000
4.4444
20.7963
10.4074
8.3673
3.3061
4.5918
2.8980
5.3061
2.6735
3.3469
3.9184
1.0204
2.6735
1 .3265
1.5306
21.6735
0.0000
5.6421
5.2421
5.0105
2.6632
3.4526
1 .8526
2.1579
3.6000
1.0526
5.3158
3.2842
3.0947
1.0737
0.8211
5.9053
1.9474
128.17
142.20
142.20
154.21
156.23
166.22
170.25
178.23
202.26
202.26
228.29
228.29
252.32
252.32
252.32
128.17
142.20
142.20
154.21
156.23
166.22
170.25
178.23
192.26
202.26
202.26
228.29
252.32
252.32
128.17
142.20
142.20
154.21
156.23
166.22
170.25
178.23
192.26
202.26
202.26
228.29
228.29
252.32
252.32
252.32
1
2
3
4
5
6
7
8
10
11
12
13
14
15
16
1
2
3
4
5
6
7
8
9
10
11
12
15
16
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
0.02908
0.00978
0.00973
0.00692
0.00000
0.00597
0.00000
0.03581
0.03507
0.01828
0.01323
0.00560
0.01004
0.09825
0.01764
0.03354
0.01605
0.01474
0.00989
0.00000
0.00000
0.00840
0.05290
0.00621
0.01341
0.01347
0.00733
0.19272
0.00505
0.02878
0.01581
0.01426
0.01265
0.02041
0.01450
0.02294
0.05791
0.01143
0.02898
0.02518
0.02311
0.01111
0.00822
0.05531
0.01410
0.20314
0.05352
0.08439
0.07758
0.08689
0.00000
0.05558
0.06296
0.05393
0.03104
0.02150
0.00000
0.01761
0.08242
0.04125
0.06528
0.02325
0.03229
0.01879
0.03396
0.01608
0.01966
0.02198
0.00531
0.01322
0.00656
0.00670
0.08590
0.00000
0.04402
0.03686
0.03524
0.01727
0.02210
0.01115
0.01267
0.02020
0.00548
0.02628
0.01624
0.01356
0.00470
0.00325
0.02340
0.00772
                                          2.23
-------
Appendix 2.4
                    DBS   POOL
                                   LLCONC
                                             FLCONC    MOLEUT
                                                               CHEM
                                                                        LMML
                                                                                   FMML
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
61
62
63
64
65
66
67
68
69
70
71
72
73
74
75
76
77
78
79
80
81
82
83
84
85
86
87
88
89
90
7
7
7
7
7
7
7
7
7
7
7
7
7
7
7
7
9
9
9
9
9
9
9
9
9
9
9
9
9
9
9
9
10
10
10
10
10
10
10
10
10
10
10
10
10
5.014
3.454
3.939
2.312
2.896
1.600
1.622
9.197
1.527
5.646
4.740
3.972
1.362
2.357
190.789
3.386
4.952
2.112
3.648
1.782
3.469
1.057
1.321
7.274
0.000
2.744
2.532
2.018
1.819
1.052
70.149
2.052
5.103
1 .794
1.835
1.360
1.261
0.971
1.175
1.916
1.466
2.581
1.785
1.412
0.000
5.9123
2.6316
3.5789
2.4737
3.4386
1.3158
1.8772
2.3509
1.1228
3.1053
2.5789
3.0175
0.6140
0.7193
62.2456
1 .4035
6.0877
6.2807
8.1579
3.5088
8.6491
2.2456
3.0000
4.6667
1 .8070
2.7719
1 .8947
1.2807
0.3333
0.6667
35.6667
1.1579
4.6900
1 .3400
2.6300
1.6200
2.6500
0.9100
1.2500
1 .6300
0.8700
1 .4000
1.2300
1 .2500
0.1800
128.17
142.20
142.20
154.21
156.23
166.22
170.25
178.23
192.26
202.26
202.26
228.29
228.29
252.32
252.32
252.32
128.17
142.20
142.20
154.21
156.23
166.22
170.25
178.23
192.26
202.26
202.26
228.29
228.29
252.32
252.32
252.32
128.17
142.20
142.20
154.21
156.23
166.22
170.25
178.23
192.26
202.26
202.26
228.29
228.29
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
1
2
3
4
5
6
7
8
9
10
11
12
13
0.03912
0.02429
0.02770
0.01500
0.01853
0.00962
0.00953
0.05160
0.00794
0.02791
0.02344
0.01740
0.00597
0.00934
0.75614
0.01342
0.03864
0.01485
0.02566
0.01156
0.02220
0.00636
0.00776
0.04081
0.00000
0.01356
0.01252
0.00884
0.00797
0.00417
0.27802
0.00813
0.03981
0.01261
0.01290
0.00882
0.00807
0.00584
0.00690
0.01075
0.00762
0.01276
0.00883
0.00618
0.00000
0.04613
0.01851
0.02517
0.01604
0.02201
0.00792
0.01103
0.01319
0.00584
0.01535
0.01275
0.01322
0.00269
0.00285
0.24669
0.00556
0.04750
0.04417
0.05737
0.02275
0.05536
0.01351
0.01762
0.02618
0.00940
0.01370
0.00937
0.00561
0.00146
0.00264
0.14135
0.00459
0.03659
0.00942
0.01850
0.01051
0.01696
0.00547
0.00734
0.00915
0.00453
0.00692
0.00608
0.00548
0.00079
                                               2.24
-------
Appendix 2.4
                     OBS
                           POOL
                                    LLCONC
                                              FLCONC
                                                       HOLEWT
                                                                CHEM
                                                                         LMHL
                                                                                    FHML
91
92
93
94
95
96
97
98
99
100
101
102
103
104
105
106
107
108
109
110
111
112
113
114
115
116
117
118
119
120
121
122
123
124
125
126
127
128
129
130
131
132
133
134
135
10
10
10
11
11
11
11
11
11
11
11
11
11
11
11
11
11
11
11
12
12
12
12
12
12
12
12
12
12
12
12
12
12
12
12
13
13
13
13
13
13
13
13
13
13
1.042
113.346
1.006
3.610
1.830
1.981
1.585
1.673
0.000
0.858
1.735
0.855
2.051
1.421
1.413
0.613
0.709
129.307
1.672
3.986
1.608
2.121
1.727
1.493
1.071
1.052
2.053
1.043
2.587
2.167
1.548
0.000
0.876
54.336
1.057
5.511
2.642
2.956
2.530
1.658
0.805
1.055
1.927
1.420
3.165
0.4700
52.9900
0.4900
7.2371
2.5155
4.7216
2.1031
4.2474
14.5464
1.7938
2.4639
1 .3299
2.4433
0.0000
2.2474
0.0000
0.0000
48.6082
0.0000
11.4694
5.9796
7.1429
3.6531
10.5918
3.7959
8.8980
19.2245
5.6939
4.5102
3.9184
1.6122
2.4490
0.0000
26.2653
0.0000
3.9242
3.6212
5.4242
1.4242
5.0303
0.8333
2.3788
1.7727
0.7121
2.0455
252.32
252.32
252.32
128.17
142.20
142.20
154.21
156.23
166.22
170.25
178.23
192.26
202.26
202.26
228.29
228.29
252.32
252.32
252.32
128.17
142.20
142.20
154.21
156.23
166.22
170.25
178.23
192.26
202.26
202.26
228.29
228.29
252.32
252.32
252.32
128.17
142.20
142.20
154.21
156.23
166.22
170.25
178.23
192.26
202.26
14
15
16
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
1
2
3
4
5
6
7
8
9
10
0.00413
0.44922
0.00399
0.02817
0.01287
0.01393
0.01028
0.01071
0.00000
0.00504
0.00974
0.00445
0.01014
0.00702
0.00619
0.00268
0.00281
0.51247
0.00663
0.03110
0.01131
0.01491
0.01120
0.00956
0.00645
0.00618
0.01152
0.00542
0.01279
0.01071
0.00678
0.00000
0.00347
0.21535
0.00419
0.04300
0.01858
0.02079
0.01641
0.01061
0.00484
0.00619
0.01081
0.00739
0.01565
0.00186
0.21001
0.00194
0.05646
0.01769
0.03320
0.01364
0.02719
0.08751
0.01054
0.01382
0.00692
0.01208
0.00000
0.00984
0.00000
0.00000
0.19265
0.00000
0.08949
0.04205
0.05023
0.02369
0.06780
0.02284
0.05226
0.10786
0.02962
0.02230
0.01937
0.00706
0.01073
0.00000
0.10410
0.00000
0.03062
0.02547
0.03815
0.00924
0.03220
0.00501
0.01397
0.00995
0.00370
0.01011
                                                2.25
-------
Appendix 2.4
                    DBS
                           POOL
                                   LLCONC
                                              FLCONC
                                                       HOLEWT
                                                                CHEM
                                                                         LMML
                                                                                   FHML
136
137
138
139
140
141
142
143
144
145
146
147
148
149
150
151
152
153
154
155
156
157
158
159
160
161
162
163
164
165
166
167
168
169
170
171
172
173
174
175
176
177
178
179
180
13
13
13
13
13
13
14
14
14
14
14
14
14
14
14
14
14
14
14
14
14
14
16
16
16
16
16
16
16
16
16
16
16
16
16
16
16
16
19
19
19
19
19
19
19
2.4696
2.6575
1.1395
1.0265
92.5926
1.4223
4.0636
2.2940
2.2365
.3810
.6780
.1280
.1299
.9380
.0663
2.3709
2.4181
2.0901
0.5499
0.7160
49.2031
1.2293
5.4776
2.2395
2.1533
1.6495
1.5549
1 .3456
1.5136
6.3437
1.0846
4.8371
4.3327
3.9894
1 .4460
1.8866
26.3365
1 .9889
5.0755
1.8034
1.9948
1.7565
1 .5249
1.1007
1.1964
1.8182
1.1364
0.4848
0.5606
37.9545
1 .3788
5.2432
5.2793
6.4685
2.5676
5.8739
1.2973
2.3243
3.2793
1.1622
1 .9459
1.6577
1.1351
0.0000
0.0000
15.3694
0.6036
7.3816
6.6974
6.0526
2.7237
7.5132
1.4868
4.1974
4.5263
1.157?
4.2237
3.8684
1.7500
1.8816
0.0000
14.3816
2.1974
3.4023
3.0805
4.1839
2.4023
5.5057
1.2529
3.4253
202.26
228.29
228.29
252.32
252.32
252.32
128.17
142.20
142.20
154.21
156.23
166.22
170.25
178.23
192.26
202.26
202.26
228.29
228.29
252.32
252.32
252.32
128.17
142.20
142.20
154.21
156.23
166.22
170.25
178.23
192.26
202.26
202.26
228.29
228.29
252.32
252.32
252.32
128.17
142.20
142.20
154.21
156.23
166.22
170.25
11
12
14
14
15
16
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
1
2
3
4
5
6
7
0.01221
0.01164
0.00499
0.00407
0.36696
0.00564
0.03170
0.01613
0.01573
0.00896
0.01074
0.00679
0.00664
0.01087
0.00555
0.01172
0.01196
0.00916
0.00241
0.00284
0.19500
0.00487
0.04274
0.01575
0.01514
0.01070
0.00995
0.00810
0.00889
0.03559
0.00564
0.02392
0.02142
0.01748
0.00633
0.00748
0.10438
0.00788
0.03960
0.01268
0.01403 .
0.01139
0.00976
0.00662
0.00703
0.00899
0.00498
0.00212
0.00222
0.15042
0.00546
0.04091
0.03713
0.04549
0.01665
0.03760
0.00780
0.01365
0.01840
0.00604
0.00962
0.00820
0.00497
0.00000
0.00000
0.06091
0.00239
0.05759
0.04710
0.04256
0.01766
0.04809
0.00895
0.02465
0.02540
0.00602
0.02088
0.01913
0.00767
0.00824
0.00000
0.05700
0.00871
0.02655
0.02166
0.02942
0.01558
0.03524
0.00754
0.02012
                                                2.26
-------
Appendix 2.4
                    OBS    POOL
                                   LLCONC
                                             FLCONC   MOLEWT
                                                                CHEM
                                                                         LMHL
                                                                                   FMML
181
182
183
184
185
186
187
188
189
190
191
192
193
194
195
196
197
198
199
200
201
202
203
204
205
19
19
19
19
19
19
19
19
19
22
22
22
22
22
22
22
22
22
22
22
22
22
22
22
22
5.2950
1.1791
3.5176
3.1881
1 .9302
1.0474
1.1345
41 .6354
1.2650
4.2526
2.0454
2.1461
1.2772
1.2666
1 .3352
0.9046
3.9079
1.1377
3.1845
2.5594
2.3395
0.8239
0.9537
25.0994
1.5719
4.3793
1.6897
2.6322
2.5747
0.9425
0.7816
0.5172
18.0115
0.8161
4.1146
2.0208
3.1354
1 .8646
4.8958
1.5521
6.4375
2.7708
0.8542
2.6979
2.4896
1.2188
0.6458
0.5625
18.6667
0.9583
178.23
192.26
202.26
202.26
228.29
228.29
252.32
252.32
252.32
128.17
142.20
142.20
154.21
156.23
166.22
170.25
178.23
192.26
202.26
202.26
228.29
228.29
252.32
252.32
252.32
8
9
10
11
12
13
14
15
16
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
0.02971
0.00613
0.01739
0.01576
0.00846
0.00459
0.00450
0.16501
0.00501
0.03318
0.01438
0.01509
0.00828
0.00811
0.00803
0.00531
0.02193
0.00592
0.01574
0.01265
0.01025
0.00361
0.00378
0.09947
0.00623
0.024571
0.008788
0.013014
0.012730
0.004129
0.003424
0.002050
0.071384
0.003234
0.032103
0.014211
0.022049
0.012091
0.031337
0.009338
0.037812
0.015546
0.004443
0.013339
0.012309
0.005339
0.002829
0.002229
0.073980
0.003798
                                                2.27
-------
Appendix 2.5. Ratios of laboratory to field (L:F) and field to laboratory (F:L) tissue
concentrations. Ratios were calculated using lipid-normalized tissue concentrations. Lipid-
normalized concentrations (ug/g lipid) are listed for laboratory-exposed (LLCONC) and field-
collected (FLCONC) oligochaetes.
              OBS   CHEMICAL
                                      CHEH #    POOL
                                                     LLCONC
                                                              FLCONC
                                                                       F:L
                                                                              L:F
1
z
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
1,6,7-TRIMETHNAPH
1,6,7-TRIHETHNAPH
1,6,7-TRIMETHNAPH
1,6,7-TRIMETHNAPH
1,6,7-TRIMETHNAPH
1,6,7-TRIMETHNAPH
1,6,7-TRIMETHNAPH
1,6,7-TRIMETHNAPH
1,6,7-TRIMETHNAPH
1,6,7-TRIMETHNAPH
1,6,7-TRIMETHNAPH
1,6,7-TRIMETHNAPH
1-METHYLNAPH
1-METHYLNAPH
1-METHYLNAPH
1-METHYLNAPH
1-METHYLNAPH
1-METHYLNAPH
1-METHYLNAPH
1-METHYLNAPH
1-METHYLNAPH
1-METHYLNAPH
1-METHYLNAPH
1-METHYLNAPH
1-METHYLNAPH
1-METHYLPHEN
1-METHYLPHEN
1-METHYLPHEN
1-METHYLPHEN
1-METHYLPHEN
1-METHYLPHEN
1-METHYLPHEN
1-METHYLPHEN
1-METHYLPHEN
1-METHYLPHEN
1-METHYLPHEN
2,6-DIMETHNAPH
2,6-DIMETHNAPH
2,6-DIMETHNAPH
2,6-DIMETHNAPH
2,6-DIMETHNAPH
2,6-DIMETHNAPH
2,6-DIMETHNAPH
2,6-DIMETHNAPH
2,6-DIMETHNAPH
7
7
7
7
7
7
7
7
7
7
7
7
2
2
2
2
2
2
2
2
2
2
2
2
2
9
9
9
9
9
9
9
9
9
9
9
5
5
5
5
5
5
5
5
5
5
6
7
9
10
11
12
13
14
16
19
22
4
5
6
7
9
10
11
12
13
14
16
19
22
5
6
7
10
11
12
13
14
16
19
22
6
7
9
10
11
12
13
14
16
1.43018
3.90541
1 .62179
1.32051
1.17534
0.85849
1.05160
1.05454
1.12992
1.51357
1.19635
0.90463
1.39105
2.28229
2.24855
3.45378
2.11157
1 .79360
1 .82974
1.60816
2.64189
2.29404
2.23951
1 .80344
2.04537
1.19430
2.19802
1.52692
1 .46592
0.85487
1.04266
1 .42040
1.06633
1.08460
1.17914
1.13769
3.18818
2.89561
3.46864
1.26107
1.67319
1.49311
1 .65836
1 .67797
1 .55492
3.3469
2.1579
1.8772
3.0000
1 .2500
1.7938
8.8980
2.3788
2.3243
4.1974
3.4253
6.4375
7.6111
3.3061
5.2421
2.6316
6.2807
1 .3400
2.5155
5.9796
3.6212
5.2793
6.6974
3.0805
2.0208
1.0204
1.0526
1.1228
0.8700
1 .3299
5.6939
0.7121
1.1622
1.1579
1 .6897
0.8542
3.4526
3.4386
8.6491
2.6500
4.2474
10.5918
5.0303
5.8739
7.5132
2.34022
0.55254
1.15748
2.27184
1.06352
2.08951
8.46136
2.25577
2.05706
2.77316
2.86311
7.11619
5.47149
1 .44860
2.33133
0.76194
2.97442
0.74710
1 .37477
3.71828
1 .37069
2.30131
2.99055
1.70810
0.98801
0.85440
0.47890
0.73534
0.59348
1.55566
5.46091
0.50135
1.08987
1 .06758
1 .43296
0.75079
1.08295
1.18752
2.49352
2.10139
2.53851
7.09381
3.03330
3.50059
4.83186
0.42731
1 .80982
0.86395
0.44017
0.94027
0.47858
0.11818
0.44331
0.48613
0.36060
0.34927
0.14052
0.18277
0.69032
0.42894
1.31244
0.33620
1 .33851
0.72740
0.26894
0.72956
0.43454
0.33439
0.58545
1.01214
1.17041
2.08812
1.35992
1 .68497
0.64281
0.18312
1 .99461
0.91754
0.93670
0.69786
1 .33193
0.92341
0.84209
0.40104
0.47588
0.39393
0.14097
0.32967
0.28567
0.20696
                                           2.28
-------
Appendix 2.5
               DBS
                      CHEMICAL
                                          CHEM #    POOL
                                                          LLCONC
                                                                     FLCONC
                                                                               F:L
                                                                                      L:F
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
61
62
63
64
65
66
67
68
69
70
71
72
73
74
75
76
77
78
79
80
81
82
83
84
85
86
87
88
89
90
2,6-DIMETHNAPH
2,6-DIMETHNAPH
2-METHYLNAPH
2-HETHYLNAPH
2-METHYLNAPH
2-METHYLNAPH
2-METHYLNAPH
2-METHYLNAPH
2-HETHYLNAPH
2-METHYLNAPH
2-METHYLNAPH
2-METHYLNAPH
2-METHYLNAPH
2-METHYLNAPH
2-METHYLNAPH
ACENAPHTHENE
ACENAPHTHENE
ACENAPHTHENE
ACENAPHTHENE
ACENAPHTHENE
BENaANTHRACENE
BENaANTHRACENE
BENaANTHRACENE
BENaANTHRACENE
BENaANTHRACENE
BENaANTHRACENE
BENaANTHRACENE
BENbkFLUORAN
BENbkFLUORAN
BENbkFLUORAN
BENbkFLUORAN
BENbkFLUORAN
BENbkFLUORAN
BENbkFLUORAN
BENbkFLUORAN
BENePYRENE
BENePYRENE
BENePYRENE
BENePYRENE
BENePYRENE
BENePYRENE
BENePYRENE
BENePYRENE
BENePYRENE
BENePYRENE
5
5
3
3
3
3
3
3
3
3
3
3
3
3
3





13
13
13
13
13
13
13
14
14
14
14
14
14
14
14
16
16
16
16
16
16
16
16
16
16
19
22
4
5
6
7
9
10
11
12
13
14
16
19
22
10
12
13
14
16
6
7
9
13
16
19
22
4
6
7
9
10
13
19
22
4
6
7
9
10
13
14
16
19
22
1.52495
1.26658
1 .38352
2.09642
2.02741
3.93896
3.64825
1 .83465
1 .98082
2.12090
2.95584
2.23649
2.15334
1 .99482
2.14610
0.80778
1.00739
0.69608
0.90012
1.13831
2.53638
1 .36242
1 .81888
1.13952
1 .44604
1.04739
0.82395
5.06801
4.15012
4.71436
2.10390
2.08379
2.05292
2.26898
1 .90736
4.45106
3.55795
3.38590
2.05173
1.00561
1.42232
1 .22928
1 .98890
1.26503
1.57189
5.5057
4.8958
12.0000
4.5918
5.0105
3.5789
8.1579
2.6300
4.7216
7.1429
5.4242
6.4685
6.0526
4.1839
3.1354
0.3800
1 .3265
0.5758
0.6667
1.0132
1.0737
0.6140
0.3333
0.4848
1.8816
0.7816
0.6458
8.8889
1.6421
1.4386
1 .3333
0.9400
1.1212
1.0345
1.1250
10.4074
1 .9474
1.4035
1.1579
0.4900
1 .3788
0.6036
2.1974
0.8161
0.9583
3.61045
3.86540
8.67355
2.19032
2.47140
0.90860
2.23611
1.43351
2.38369
3.36784
1 .83509
2.89224
2.81082
2.09739
1.46098
0.47042
1.31680
0.82714
0.74064
0.89005
0.42331
0.45069
0.18326
0.42549
1.30119
0.74624
0.78383
1.75392
0.39568
0.30515
0.63374
0.45110
0.54615
0.45593
0.58982
2.33819
0.54733
0.41452
0.56435
0.48727
0.96939
0.49102
1.10481
0.64512
0.60967
0.27697
0.25871
0.11529
0.45655
0.40463
1.10059
0.44720
0.69759
0.41952
0.29693
0.54493
0.34575
0.35577
0.47678
0.68447
2.12574
0.75942
1.20898
1.35018
1.12353
2.36231
2.21880
5.45664
2.35025
0.76853
1 .34005
1.27579
0.57015
2.52732
3.27705
1.57792
2.21680
1.83099
2.19334
1 .69543
0.42768
1 .82706
2.41246
1.77195
2.05226
1.03157
2.03656
0.90513
1.55010
1.64024
                                               2.29
-------
Appendix 2.5
               OBS
                     CHEMICAL
                                          CHEH #    POOL
LLCONC
                                                                    FLCONC
                                                                               F:L
                                                                                      L:F
91
92
93
94
95
96
97
98
99
100
101
102
103
104
105
106
107
108
109
110
111
112
113
114
115
116
117
118
119
120
121
122
123
124
125
126
127
128
129
130
131
132
133
134
135

BIPHENYL
BIPHENYL
BIPHENYL
BIPHENYL
BIPHENYL
BIPHENYL
BIPHENYL
BIPHENYL
BIPHENYL
BIPHENYL
BIPHENYL
BIPHENYL
BIPHENYL
C1 -NAPHTHALENES
C1 -NAPHTHALENES
C1 -NAPHTHALENES
C1 -NAPHTHALENES
C1 -NAPHTHALENES
C1 -NAPHTHALENES
C1 -NAPHTHALENES
C1 -NAPHTHALENES
C1 -NAPHTHALENES
C1 -NAPHTHALENES
C1 -NAPHTHALENES
C1 -NAPHTHALENES
C1 -NAPHTHALENES
CHRYSENE
CHRYSENE
CHRYSENE
CHRYSENE
CHRYSENE
CHRYSENE
CHRYSENE
CHRYSENE
CHRYSENE
CHRYSENE
CHRYSENE
CHRYSENE
CHRYSENE
FLUORANTHENE
FLUORANTHENE
FLUORANTHENE
FLUORANTHENE
FLUORANTHENE
FLUORANTHENE

4
4
4
4
4
4
4
4
4
4
4
4
4













12
12
12
12
12
12
12
12
12
12
12
12
12
11
11
11
11
11
11

4
5
6
7
9
10
11
12
13
14
16
19
22
4
5
6
7
9
10
11
12
13
14
16
19
22
4
5
6
7
9
10
11
12
13
14
16
19
22
4
5
6
7
9
10
2.30
1.06709
1.52488
1.95013
2.31239
1.78193
1 .35955
1.58544
1.72667
2.52996
1.38104
1 .64953
1.75647
1.27716
2.77457
3.58325
4.28271
7.39486
5.75268
3.63483
3.81527
3.72231
5.59283
4.52395
4.39785
3.80030
4.18440
3.02049
1.67312
5.27471
3.97181
2.01778
1.41153
1.41262
1.54802
2.65747
2.09014
3.98941
1 .93023
2.33951
3.69733
2.72360
5.09321
4.74009
2.53183
1.78517

11.9630
2.8980
2.6632
2.4737
3.5088
1 .6200
2.1031
3.6531
1 .4242
2.5676
2.7237
2.4023
1.8646
19.6111
7.8980
10.2421
6.2105
14.4386
3.9700
7.2371
13.1224
9.0455
11.7477
12.7500
7.2644
5.1563
4.9074
1.5306
3.0947
3.0175
1.2807
1.2500
2.2474
1.6122
1.1364
1.1351
1.7500
0.9425
1.2188
6.2778
1 .3265
3.2842
2.5789
1 .8947
1.2300

11.2109
1 .9004
1 .3656
1.0698
1 .9691
1.1916
1 .3265
2.1157
0.5629
1 .8592
1.6512
1.3677
1 .4599
7.0682
2.2041
2.3915
0.8398
2.5099
1.0922
1 .8969
3.5254
1.6173
2.5968
2.8991
1.9115
1 .2323
1 .6247
0.9148
0.5867
0.7597
0.6347
0.8856
1.5910
1.0415
0.4276
0.5431
0.4387
0.4883
0.5209
1 .6979
0.4871
0.6448
0.5441
0.7484
0.6890

0.08920
0.52619
0.73226
0.93480
0.50785
0.83923
0.75386
0.47266
1.77636
0.53788
0.60563
0.73116
0.68496
0.14148
0.45369
0.41815
1.19070
0.39842
0.91557
0.52718
0.28366
0.61830
0.38509
0.34493
0.52314
0.81152
0.61550
1.09311
1.70441
1.31624
1.57553
1.12922
0.62855
0.96016
2.33857
1.84131
2.27966
2.04792
1.91960
0.58896
2.05317
1.55082
1 .83799
1 .33624
1.45136

-------
Appendix 2.5
                DBS
                      CHEMICAL
                                           CHEM #    POOL
                                                           LLCONC
                                                                     FLCONC
                                                                               F:L
                                                                                       L:F
136
137
138
139
140
141
142
143
144
145
146
147
148
149
150
151
152
153
154
155
156
157
158
159
160
161
162
163
164
165
166
167
168
169
170
171
172
173
174
175
176
177
178
179
180
FLUORANTHENE
FLUORANTHENE
FLUORANTHENE
FLUORANTHENE
FLUORANTHENE
FLUORANTHENE
FLUORENE
FLUORENE
FLUORENE
FLUORENE
FLUORENE
FLUORENE
FLUORENE
FLUORENE
FLUORENE
FLUORENE
NAPHTHALENE
NAPHTHALENE
NAPHTHALENE
NAPHTHALENE
NAPHTHALENE
NAPHTHALENE
NAPHTHALENE
NAPHTHALENE
NAPHTHALENE
NAPHTHALENE
NAPHTHALENE
NAPHTHALENE
NAPHTHALENE
PERYLENE
PERYLENE
PERYLENE
PERYLENE
PERYLENE
PERYLENE
PERYLENE
PERYLENE
PERYLENE
PERYLENE
PERYLENE
PERYLENE
PERYLENE
PHENANTHRENE
PHENANTHRENE
PHENANTHRENE
11
11
11
11
11
11
6
6
6
6
6
6
6
6
6
6
1
1
1
1
1
1
1
1
1
1
1
1
1
15
15
15
15
15
15
15
15
15
15
15
15
15
8
8
8
12
13
14
16
19
22
6
7
9
10
12
13
14
16
19
22
4
5
6
7
9
10
11
12
13
14
16
19
22
4
5
6
7
9
10
11
12
13
14
16
19
22
4
5
6
2.167
2.470
2.418
4.333
3.188
2.559
2.410
1.600
1.057
0.971
1.071
0.805
1.128
1.346
1.101
1.335
3.728
4.299
3.688
5.014
4.952
5.103
3.610
3.986
5.511
4.064
5.478
5.075
4.253
24.790
48.628
13.956
190.789
70.149
113.346
129.307
54.336
92.593
49.203
26.337
41.635
25.099
6.382
9.429
10.321
3.9184
1.8182
1.6577
3.8684
2.5747
2.4896
1.8526
1.3158
2.2456
0.9100
3.7959
0.8333
1.2973
1 .4868
1.2529
1.5521
26.0370
8.3673
5.6421
5.9123
6.0877
4.6900
7.2371
11.4694
3.9242
5.2432
7.3816
3.4023
4.1146
20.7963
21 .6735
5.9053
62.2456
35.6667
52.9900
48.6082
26.2653
37.9545
15.3694
14.3816
18.0115
18.6667
11.2222
3.9184
3.6000
1.80861
0.73624
0.68552
0.89285
0.80760
0.97273
0.76882
0.82251
2.12404
0.93722
3.54323
1 .03481
1.15012
1.10498
1.13824
1.16242
6.98461
1 .94651
1 .52973
1.17919
1.22923
0.91909
2.00455
2.87759
0.71211
1.29029
1 .34760
0.67034
0.96754
0.83891
0.44570
0.42314
0.32625
0.50844
0.46751
0.37591
0.48339
0.40991
0.31237
0.54607
0.43260
0.74371
1 .75850
0.41558
0.34880
0.55291
1 .35826
1 .45875
1.12001
1.23823
1.02804
1 .30069
1.21578
0.47080
1.06699
0.28223
0.96636
0.86947
0.90499
0.87855
0.86027
0.14317
0.51374
0.65371
0.84804
0.81351
1 .08804
0.49887
0.34751
1.40429
0.77502
0.74206
1.49178
1.03355
-1.19202
2.24367
2.36330
3.06510
1 .96680
2.13901
2.66019
2.06873
2.43956
3.20138
1.83127
2.31160
1 .34461
0.56867
2.40627
2.86693
                                               2.31
-------
Appendix 2.5
               DBS
                      CHEMICAL
                                          CHEH #   POOL
                                                           LLCONC
                                                                     FLCONC
                                                                               F:L
                                                                                      L:F
181
182
183
184
185
186
187
188
189
190
191
192
193
194
195
196
197
198
199
200
201
202
203
204
205
206
207
208
209
210
211
212
213
214
215
216
PHENANTHRENE
PHENANTHRENE
PHENANTHRENE
PHENANTHRENE
PHENANTHRENE
PHENANTHRENE
PHENANTHRENE
PHENANTHRENE
PHENANTHRENE
PHENANTHRENE
PYRENE
PYRENE
PYRENE
PYRENE
PYRENE
PYRENE
PYRENE
PYRENE
PYRENE
PYRENE
PYRENE
PYRENE
PYRENE
TOTAL PCBs
TOTAL PCBs
TOTAL PCBs
TOTAL PCBs
TOTAL PCBs
TOTAL PCBs
TOTAL PCBs
TOTAL PCBs
TOTAL PCBs
TOTAL PCBs
TOTAL PCBs
TOTAL PCBs
TOTAL PCBs
8
8
8
8
8
8
8
8
8
8
10
10
10
10
10
10
10
10
10
10
10
10
10













7
9
10
11
12
13
14
16
19
22
4
5
6
7
9
10
11
12
13
14
16
19
22
4
5
6
7
9
10
11
12
13
14
16
19
22
9.197
7.274
1.916
1.735
2.053
1.927
1.938
6.344
5.295
3.908
7.094
2.711
5.861
5.646
2.744
2.581
2.051
2.587
3.165
2.371
4.837
3.518
3.185
133.321
26.355
28.644
14.275
15.253
23.962
39.490
25.261
10.182
14.400
58.854
50.689
25.622
2.351
4.667
1.630
2.464
19.224
1.773
3.279
4.526
4.379
2.771
10.907
2.673
5.316
3.105
2.772
1.400
2.443
4.510
2.045
1.946
4.224
2.632
2.698
149.278
66.061
13.768
99.316
17.000
14.350
19.979
21.204
8.197
27.045
25.026
6.310
5.813
0.25562
0.64156
0.85076
1.41975
9.36506
0.92014
1.69212
0.71351
0.82707
0.70903
1.53757
0.98599
0.90694
0.54999
1.01032
0.54242
1.19119
1.74316
0.64631
0.82078
0.87318
0.74829
0.84720
1.11969
2.50663
0.48067
6.95756
1.11453
0.59887
0.50593
0.83940
0.80506
1 .87813
0.42523
0.12449
0.22686
3.9T206
1.55870
1.17542
0.70435
0.10678
1.08679
0.59098
1.40152
1 .20909
1.41037
0.65038
1.01421
1.10261
1 .81823
0.98978
1 .84359
0.83949
0.57367
1 .54726
1.21836
1.14524
1 .33639
1.18036
0.89310
0.39894
2.08043
0.14373
0.89724
1 .66982
1 .97656
1.19132
1.24214
0.53245
2.35168
8.03273
4.40802
                                                2.32
-------
Chapter 3: Assessing Sediment Toxicity from Upper Mississippi River Navigational Pools Using a
Benthic Invertebrate Community Evaluation and the Sediment Quality Triad Approach

Canfield, T.J., Brunson, E.L., Dwyer, F.J., Ingersoll, C.G., and Kemble, N.E.

Introduction

The Mississippi River is the central catchment for a majority of the water runoff between the
west side of the Appalachian mountain range to the east side of the Rocky mountain range.  This
makes the Mississippi River system the largest in the United States, the third largest drainage
worldwide, and the seventh largest average discharge worldwide (Van der Leeden, Troise and
Todd 1990).  The river receives inputs from municipal, agricultural, and industrial sources.
Previous studies have examined the concentrations of organic and inorganic contaminants in the
sediments from select pools in the Upper Mississippi River (Wiebe 1927; Bailey and Rada 1984;
Weiner et al.  1984; Rada et al. 1990). Recently, some studies have reported a decline in the levels
of contaminants in the sediments of the Upper Mississippi River (Rada et al. 1990).
    The Upper Mississippi River (UMR), that part of the river north of the confluence with the
Ohio River at Cairo, IL, is divided into a series of large runs and pools by 26 locks and dams
constructed for navigational purposes (Rada et al. 1990). This lock and dam system,  which runs
from Minneapolis, MN to St. Louis, MO, provides areas for deposition of large quantities of fine
grained sediments during normal and low flows (Nielson et al. 1984).  Contaminants are often
associated with fine-grained sediments and settle along with these  sediments (Forstner and
Wittmann 1979, Hassett et al.  1980).  Sediments often serve as a  sink for an array of organic and
inorganic contaminants when the water to sediment gradient is high, and  these sediments can act
as a source of contamination when the water to sediment gradient is low (Shimp et al. 1971;
Oschwald 1972; Medine and McCutcheon 1989).
    Benthic macroinvertebrates inhabiting the sediments are presumably exposed continuously to
any contaminants contained in the sediments. Benthic macroinvertebrate abundance, community
structure, and ecological function have long been used to characterize water quality in freshwater
ecosystems (Davis and Lathrop 1992). Numerous studies have documented potential changes in
benthic invertebrate community structure associated with the impacts of contaminants (Cook and
Johnson 1974; Rosenberg and Wiens 1976; Hilsenhoff 1982, 1987; Waterhouse and Farrell 1985;
Clements et al. 1992). Most studies in lotic environments have examined the responses of benthic
macroinvertebrate communities in riffle areas due to ease of collection and observed higher taxa
richness.  However, only a limited number of assessments have been conducted in depositional
soft-sediments (Canfield et al.  1996).
    The spatial and temporal distribution of resident organisms may reflect the degree to which
chemicals in the sediments are bioavailable and toxic. Field surveys of invertebrates can provide
an important component of biological assessments of toxicity associated with contaminated
sediments for several reasons:  (1) macroinvertebrates are abundant, relatively sedentary, easy to
collect, and ubiquitous across a broad array of sediment types; (2) many indigenous benthic
organisms complete all or most of their life cycles in the aquatic environment and may serve as
continuous monitors of sediment quality; and (3) results of an assessment of indigenous
                                           3.1
-------
populations may be useful for quantifying resource damage (Cook 1976, Pratt and Coler 1976,
Davis and Lathrop 1989).
    The United States Geological Survey (USGS) has been monitoring the UMR since 1987 to
document the fate, transport, and distribution of contaminated sediments (Moody and Meade
1995). Concern with regard to the fate of contaminated sediments in the UMR arose after the
flood of 1993, because of the potential for re-exposure of deeply buried, potentially highly
contaminated sediments. Further, the flood inundated numerous riparian areas known to contain
both diffuse and concentrated (i.e. fuel tanks, warehouses) sources of contaminants.  This study
was designed to evaluate the current status of sediments in the UMR and is one chapter in a series
designed to assess the extent of sediment contamination in the navigational pools of the river.
The overall study consisted of the following components: (1) monitoring concentrations of
contaminants in the Mississippi River sediments (Moody et al. 1996);  (2) toxicity testing with
whole-sediments collected from the river (Chapter 1); (3) bioaccumulation tests with whole-
sediments collected from the river (Chapter 2);  and (4)  analysis of benthic invertebrate
community structure. The objective of this portion of the study was three-fold:  (1)  describe
distributions and abundances of benthic invertebrates in soft-sediments from selected locations in
pools of the UMR; (2) evaluate impacts of contaminants associated with these sediments using
measures of benthic invertebrate community structure;  and  (3) evaluate the concordance of
benthic invertebrate assessments to sediment toxicity and sediment chemistry using the sediment
quality triad approach.

Materials and Methods

Sampling Locations

Stations were selected for assessment of sediment toxicity, sediment chemistry and benthic
macroinvertebrate communities based on historical chemistry data (Moody et al. 1996) and the
availability of soft sediments (Chapter 1). Upper Mississippi River pools were sampled from June
11 to July 5, 1994. Stations were located in 23 of 26 pools in the UMR from pool 1 near
Hastings, MN to pool 26 near St. Louis, MO (Figure 1.1, Chapter 1). A complete description of
the sampling locations in each pool is described in Kemble et al. (Chapter 1) and bioaccumulation
data is contained in Brunson et al. (Chapter 2).

Sediment Collection, Handling, and Storage

Locations of stations for field sampling were determined with a Global Positioning System. A
stainless steel standard Ponar grab (23 x 23 cm, 529 cm2 area) was used to collect bulk sediments
from about the upper 6 to 10 cm of the sediment for chemistry analyses, laboratory toxicity
assessments and benthic invertebrates assessments at one station per pool (Chapter 1). Each
sample was a composite of 35 to 80 L of sediment/station (identified as C samples in Chapter 1).
Sediments were placed in a 120-L high density polyethylene drum and homogenized with a hand-
held power drill and a stainless steel auger. A 2.5 liter subsample of sediment for evaluations of
benthos was obtained taken from the composite C sample before subsamples were obtained for
                                           3.2
-------
 chemistry and laboratory analyses (Chapter 1).  To isolate the benthos, these 2.5 liter subsamples
 were sieved through an ASTM No.30 (533 um) and an ASTM No. 60 (250 urn) bucket
 connected in series using screened river water for rinsing. Material containing benthos retained by
 the sieves was combined and transferred into 1L high density polyethylene jars, preserved with
 10% buffered formalin, and transported to the laboratory. Subsamples for use in toxicity and
 bioaccumulation testing (10L), for chemical characterization (250 ml for metals, 250 ml for
 organics), and for physical characterization (250 ml) were taken and stored in high-density
 polyethylene containers or amber glass I-CHEM bottles (chemical characterizations only). All
 samples were stored at 4 °C in the dark (Chapter 1).

 Taxonomic Identification

 The preserved samples of benthos were placed in a sieve (250 um) and rinsed thoroughly with tap
 water in the laboratory to remove formalin and excess silt or mud before sorting. The samples
 were drained of excess water, returned to the original jars, filled with 95% ethanol and allowed to
 soak for at least 24 h to facilitate extraction of volatile compounds. Aliquots of the sample were
 sequentially removed from the jar to sort benthic invertebrates until the entire sample had been
 sorted.
     A binocular dissecting microscope (4x to 12x power) was used to sort and pick the entire
 sample. Invertebrates were initially sorted and enumerated into the following orders or families:
 Oligochaeta, Chironomidae, Bivalvia, Gastropoda, Ephemeroptera, Odonata, Plecoptera,
 Hemiptera, Megaloptera, Trichoptera, Coleoptera, Diptera, Hirudinea, and Amphipoda. Taxa
 were identified to the lowest practical level using appropriate taxonomic keys (Wiederholm 1983;
 Merritt  and Cummins 1984; Pennak 1989; Thorp and Covich 1991).  The following benthic
 macroinvertebrate metrics were calculated:  macroinvertebrate abundance (number/m2), species
 composition, and taxa richness (Appendix 3.1, 3.2, 3.3).   All taxa were either identified or
 verified by personnel at the Aquatic Resources Center in Franklin, TN.
     Chironomid larvae (midge) were examined for deformities in mouthpart structures. These
 deformities,  which included various types of asymmetry,  missing teeth, extra teeth, fusion among
 various  teeth, and labial separation, have been described by several investigators (Saether 1970;
Hamilton and Saether 1971; Hare and Carter 1976; Warwick et al.  1987; Warwick 1989).
 Individual midge were mounted on slides and their mouthparts were examined for deformities in
the mentum  and ligula (Tanypodinae only).  Occurrence of deformities was expressed as a
 proportion of the total number of midges at each station.

Physical and Chemical Characterizations of Sediment

 Sediment physical characteristics included the following: (1) sediment particle size, (2) total
 organic carbon, (3) inorganic carbon and (4) percent water.   Sediment chemical parameters
 included the following: (1) chlorinated pesticides, (2) polychlorinated biphenyls (PCB), (3) select
 aliphatic and polynuclear aromatic hydrocarbons (PAH), (4) simultaneously extracted metals
 (SEM), (5) acid volatile sulfide (AVS), and (6) total metals.  See Chapter 1 for additional
 information on chemical and physical characteristics of the sediments.
                                           3.3
-------
Statistical Analyses

Statistical analysis was performed with the Statistical Analysis System (SAS, Statistical Analysis
System 1994). Comparisons between benthic invertebrate abundance and physical and chemical
data were made with a Spearman Rank correlation and multivariate regression.  If not reported,
statements of statistical significance indicate p< 0.05.

Sediment Quality Triad Assessments

The sediment quality triad (Triad) approach was used as an effects based approach to integrate
data from chemical and physical analyses (e.g., PAHs, metals, grain size), sediment laboratory
toxicity exposures (e.g., Hyalella azteca survival and growth) and benthic community structure
(e.g. biotic index, taxa richness, midge mouth-part deformities) in order to evaluate the level of
concordance between these three measures and  the degree of contaminant-induced degradation in
aquatic communities in soft-sediment depositional areas (Chapman et al. 1992).  Toxicity,
benthos,  and chemistry data were scored using procedures developed by Kreis (1988) and data
were plotted using procedures described by Canfield et al. (1994, 1996). Values for each
individual variable for all samples were scaled proportionally between 1 and 100 (e.g., 1 is
indicative of the lowest concentration or least impacted, and 100 is the greatest concentration or
most impacted). Scaling data retains proportional differences between measurements and results
in an identical range for all variables.  Typically, more than one variable is determined for a
particular Triad component (e.g. Hyalella azteca survival and growth). In these instances Kreis
(1988) recommends: (1) scaling each individual variable among samples, (2) summing the scaled
values for each variable, and then  (3) re-scaling the sums for all samples. This results in scaled
scores (e.g., toxicity, benthos, or chemistry) between 1 and 100 for each Triad component which
can be compared graphically or in tabular form.
    The high and low values used to establish scores for each of the sets of information for
benthos, chemistry, and laboratory toxicity were previously reported in Canfield et al. (1996). In
order to  evaluate the extent of contamination of the UMR in the context of other areas of concern
in North America, we used data from three Great Lakes Areas of Concern (Canfield  et al. 1996)
and data from a study of the upper Clark Fork River, including Milltown Reservoir, in Montana
(Canfield et al. 1994).  Inclusion of these data sets provided a larger number of stations with a
broad range in levels of contamination that could be used in the analyses of the relative responses
of benthic communities in select sampling locations in the UMR and evaluate the relative
contamination of sediments in the UMR sediments when compared to other areas. Six benthic
invertebrate indices were used to evaluate the extent of sediment contamination in the UMR
system: (1) total taxa richness, (2) chironomid genera richness,  (3) chironomid mouthpart
deformities, (4) chironomid biotic index, (5) chironomid/oligochaete ratio, and (6) oligochaete
biotic index. The Hilsenhoff index of Biotic Integrity was used to calculate the biotic indices for
both the midges and oligochaetes.  Species sensitivity within a genera was obtained primarily from
those assigned by Hilsenhoff (1982, 1987) and secondarily by Lenat (1993).
     To evaluate the chemistry portion of the Triad, we used Effect-Range Median (ERM)
 concentrations calculated by Ingersoll et al. (1996). An ERM is defined as the concentration of a
                                            3.4
-------
chemical in sediment above which effects are frequently or always observed (Long et al. 1995).
We used seven ERM values which correctly classify laboratory toxicity >70% of the time in
Hyalella azteca 28-d tests (Ingersoll et al. 1996). These seven ERMs would more closely identify
cause and effect toxicity rather than correlative toxicity (Canfield et al. 1996).  These ERMs
included: cadmium, nickel, lead, zinc, chrysene,  benzo(a)pyrene, and benzo(g,h,i)perylene.
    To evaluate the laboratory toxicity portion of the Triad we used amphipod (Hyalella azteca)
28-d growth and survival to score laboratory toxicity (Chapter 1).  Amphipods are sensitive to
contaminated sediments and frequently exhibit reduced survival and growth following exposures
to contaminated sediments (Burton et al. 1996, Ingersoll et al. 1996). Each sample was
designated as toxic when either survival or growth of Hyalella were significantly reduced relative
to the control or reference sediment  (Chapter 1; Kemble et al.  1994; Ingersoll et al.  1996;
USEPA 1993 ).  Associations between benthic indices or laboratory toxicity tests and sediment
chemistry were evaluated by plotting the scores  of either the benthic indices or laboratory toxicity
data against the sum of the ERM quotient (SERM-Q: ERM-Q=concentratidn of a chemical in
sediment sample / ERM for that chemical) for all seven chemicals in a sample.  This approach is
similar to a toxic unit approach.
    Well defined guidelines have not been developed for distinguishing impacts of contaminant
effects on benthos found in soft sediments in either lakes, streams, or rivers.  Canfield et al.
(1996) incorporated data plots (partitioned by using quadrants defined by no effect concentration
data for plotting) and frequency analysis to identify the distribution of the data points to identify
relations between sediment  chemistry, laboratory toxicity and benthic invertebrate distributions.
This quadrant frequency analysis (essentially a frequency analysis to identify correct classification
and Type I and Type II error) was conducted in order to evaluate which benthic indices were
most sensitive to elevated contaminant concentrations.  In this analysis, scores for benthic indices
are plotted against scores for chemical contamination in sediments.  Quadrants were then defined
which identified one of four possible conditions:  (1) low chemical concentration and benthos not
adversely impacted, (2) elevated chemical concentration and benthos adversely impacted, (3)
low chemical concentration and benthos adversely impacted (Type I error, false positive), and  (4)
elevated chemical concentration and benthos not adversely impacted (Type II error, false
negative). Various combinations of benthic indices were evaluated by adding the individual scores
and re-scoring.  These analyses were conducted for all possible combinations of the six scored
benthic indices listed above.
    Sediment toxicity studies were conducted on sediments from all of the UMR pools (except
pools 3 and  17, Chapter 1). The results of the tests on the UMR sediments were combined with
data from 19 Great Lakes sediment samples (Ingersoll et al. 1996) and 13 Clark Fork
River/Milltown Reservoir samples (Kemble et al. 1994) in order to evaluate the toxicity of the
UMR sediments in the context of other samples previously evaluated. Based on previous plots of
toxicity scores (Canfield et al. 1996), the vertical quadrant line above which no non-toxic samples
were observed and above which chemical contamination was considered toxic was sum of the
ERM quotient of 39 (Figure 3.1).  The horizontal quadrant line depicting laboratory toxicity was
a score of 30, which corresponded to the greatest laboratory score above which no non-toxic
samples were observed.   This selection procedure for establishing quadrant lines may be less
environmentally protective since some of the samples that had a sum of the ERM quotient score
                                           3.5
-------
less than 39 were toxic to Hyalella azteca in the laboratory studies (Chapter 1; Kemble et al.
1994; Ingersoll et al. 1996).
    Scores for each of the benthic indices and all combinations of scores were plotted against the
sum of the ERM quotient. The position of quadrant lines for benthic indices were determined in 3
steps: (1) plotting the data,  (2) drawing the vertical quadrant line at 39 for the Sum of the ERM
quotient, (3) by evaluating the distribution of the data and selecting a benthic score (horizontal
quadrant line) which maximized the number of points in quadrants which would be considered
"correctly classified" and minimized the number of samples with "Type I, false positive" and
"Type n, false negative" error results.

Results and Discussion

Benthic Invertebrate Assessments

Abundance:  Benthic invertebrates from the UMR exhibited a wide range of abundance values.
Benthic invertebrate abundance (number/m2) in samples ranged from 250/m2 in sample 1C  to a
maximum of 22,389/m2 in sample 19C (Table 3.1). Total abundance values were less than
8,000/m2 in 21 of 24 samples with the remaining 3 samples having abundance values two-fold
greater than any of the other samples.  Oligochaetes were numerically dominant in 12 of 24
samples. Midge comprised the majority of the community in 8 of 24 samples with the bivalves
(2), mayflies (1) and nematodes (1) comprising the majority of the community in 4 of 24 samples
(Table 3.1; Appendix 3.1, 3.2, 3.3).
    Oligochaete abundance ranged from 63/m2 in sample 5C to 12,111/m2 in sample 19C (Table
3.1).  Across the pools there were order of magnitude differences in abundance values. In
general, oligochaete abundance is lowest in samples from the upper pools (1 to 7) and higher in
the lower pools (Table 3.1).  We expected these differences to be explained by organic carbon
and grain size although no significant correlations were observed in the correlation analysis (Table
3.2) evident with this data set.
    Chironomid abundance ranged from zero in samples from station 7C to 8,889/m2 in samples
from station 15C (Table 3.1).  Distribution of midge was fairly even across this range. These
values for chironomid abundances were generally higher than those reported from contaminated
sediments in Milltown Reservoir/Clark Fork River (Canfield et al. 1994) or the Great Lakes
(Canfield et al. 1996).

Community Composition:    Samples from the UMR had a fairly diverse benthic invertebrate
community (Table 3.1). Overall taxa richness was greater in samples from the lower two-thirds of
the river than in the upper 8 pools.  Oligochaete abundance accounted for 5 to 90% of the
community in all samples. Combined oligochaete and midge abundance accounted for 8 to 100%
of the total benthic invertebrate community in all samples, with the remainder of the benthic
community abundance coming primarily from the Bivalvia and Ephemeroptera.
    The oligochaete community was comprised of 2 families, 5 genera and 9 species (Appendix
3.1).  Samples from 20C and 11C had the highest number of species, while samples from 1C, 4C
and 5C  each had only one species. Except for IOC, the oligochaete community was made up
                                          3.6
-------
entirely of the family Tubificidae.  Limnodrilus spp., generally considered tolerant of organic and
metal contamination (Kennedy 1965, Brinkhurst et al. 1972, Burt et al. 1991), was the most
common genera occurring in samples from the UMR.
    The midge community was comprised of 4 subfamilies (Chironomini, Tanipodinae,
Tanytarsini, Orthocladinae) and 18 genera (Appendix 3.2). The sample from station IOC had the
highest number of genera present (8), while sample from station 7C had no genera present.
Chironomus spp. was the most abundant genera present in 17 of 24 samples, with Procladius spp.
the most abundant in 3 of the remaining samples.
    The Bivalvia (clams) and aquatic insects (excluding midge) comprised a large part (>20%) of
the community collected in 11 of 24 samples (Appendix 3.3).  Bivalvia abundance ranged from
zero in 7 samples to 16,722/m2 in sample 9C (Table 3.1).  The Bivalvia were present in 17 of 24
samples. Bivalvia abundance was greater than or equal to 1,000/m2 in 5 of 24 samples. Bivalvia
abundance of 16,722/m2 in sample 9C is 1 to 2 orders of magnitude greater than all other samples
collected and comprises 77% of the overall community abundance (Table 3.1). The Bivalvia
community was made up almost entirely ofMusculium transversum.
    The Ephemeroptera (mayflies) were present in 16 of 24 samples.  Ephemeroptera abundance
ranged from absent in 8 samples to 3,278/m2 in sample 19C (Table 3.1). Ephemeroptera
abundance was greater than or equal to 500/m2 in 6 of 24 samples, but were entirely absent in 8 of
24 samples.  The Ephemeroptera community was comprised of 2 families, 2 genera and 3 species.
The majority of the insect community (chironomidae excluded) was comprised offfexagenia sp.
    The estimated abundance values of benthic invertebrates collected in this study are
comparable with the values of invertebrates collected in previous studies of the UMR (Eckblad et
al. 1977; Butts and  Sparks 1982; Neuswanger, Taylor and Reynolds 1982; Eckblad 1986; Jahn
and Anderson 1986; Hornbach et al. 1989). Although there is some variation among  studies,
abundances were within the same range regardless of the study. In 1991, the Environmental
Management Program, Long Term Resource Monitoring Program (LTRMP) issued an
observation bulletin (LTRMP Observational Bulletin NO. 1, Eckblad 1991) which reported on the
observed decline of macroinvertebrate communities (primarily the fingernail clams) in the UMR.
Data from our study does not support the trends reported in the LTRMP report.  Benthos
abundances were above the low level warnings issued in the LTRMP report.  Differences in
abundances may be due to natural spatial or temporal variation in the invertebrate communities or
conditions in the river and sediments have changed between the time when the LTRMP report
was issued and when we conducted our study.

Deformities in chironomids

The frequency of mouth part deformities in the midge community ranged from a low of zero in
samples from 11 stations to a maximum of 13% in sample 20C (Figure 3.2). Deformities were
present in 13 out of 24 samples in the UMR, although only 4  of 24 samples had deformities which
could be considered above the identified background levels of 3 to 4% (Dickman, Brindle and
Benson 1992).
    Different genera of midge exhibit different levels of susceptibility or tolerance to
contaminants (Hamilton and Saether 1971; Hare and Carter 1976; Warwick 1985, 1988;
                                         3.7
-------
Wiederholm 1984).  Some genera are quite intolerant and are eliminated from locations with
relatively low levels of contaminants, while other genera such as Procladius spp., Chironomus
spp. and Cryptochironomus spp. are more tolerant and may persist in contaminated locations
(Warwick 1985; Bode 1988).  An association between increased contamination and the presence
of midge deformities has been observed by several investigators (Hamilton and Saether 1971;
Warwick 1985; Tennessen and Gottfried 1983; Cushman 1984; Wiederholm 1984; Diggins and
Stewart 1993).  Deformities reported in these studies include thickening of the exoskeleton,
enlargement and darkening of the head capsule, asymmetry in mouth parts, missing or fused
lateral teeth, and antenna! deformities. None of the specimens examined in the present study
exhibited antennal deformities.  Deformities observed in this study occurred only in Procladius
spp. and Chironomus spp.
    The occurrence of midge deformities is reportedly less than 1% in non-impacted or pre-
industrialization communities (Wiederholm 1984; Warwick et al. 1987). Background levels  have
been estimated at 3% to 4% (Dickman, Brindle and Benson 1992), and investigators have
suggested that frequency of deformities in the range of 5 to 25% or greater are generally
associated with moderate to severe contamination (Wiederholm 1984; Warwick et al. 1987).
Based on these criteria, deformities in midge  from the UMR indicate that sediments from only 4
samples would be classified as "moderately contaminated" (Figure 3.2).  Deformities of midges in
samples from the UMR were considerably lower than those from contaminated sediments in
studies from either the Milltown Reservoir/Clark Fork River (Canfield et al. 1994) or the Great
Lakes (Canfield et al.1996). These data indicate that overall the sediments in the UMR are
uncontaminated relative to other locations with documented occurrences of deformities in midges.

Correlation Data

Spearman rank correlations were used to compare associations of physical and chemical measures
to benthic responses because of non-normal distribution of data (Snedecor and Cochran 1982).
Few significant correlations were detected between benthic parameters and either contaminants or
abiotic factors evaluated (Table 3.2).  Significant negative correlations were observed between
total Ephemeroptera abundance (r=-0.43) and total abundance (r=-0.54) with percent sand.
Conversely, significant positive correlations were observed between  clay and total numbers
(r=0.59), bivalve abundance (r=0.49), chironomid abundance (r=0.48, Ephemeroptera abundance
(r=0.47), number of chironomid genera (r=0.46), and number of chironomid taxa (r=0.46).
Positive correlations with clay and negative correlations with sand imply that hydrological factors
such as current velocity may have been determinants of benthic distributions. For example, clay
dominated areas may support greater benthic nymphs due to increased stability of physical habitat
and increased deposition of organic matter compared to sandy areas. However, abiotic causality
is difficult to infer without additional, manipulative studies.
     Significant correlations with measures of chemical contamination were sporadic, observed
comparing the measures of total abundance (TOTAL) with zinc (Zn), number of oligochaete taxa
(OTAXA) with cadmium (Cd), the oligochaete biotic index (OLBI) with chrysene (CHRYS) and
cadmium, and total taxa richness (TXRICH) with chrysene, benzo(a)pyrene (BAP), cadmium and
nickel (Ni, Table 3.2).  Although these correlations were significant, they still explained no  more
                                           3.8
-------
than 35% of the total variability.  Further, the number of positive and negative correlations varied
within a particular chemical. This makes interpretation difficult, but may not be unexpected given
that the measured chemicals in almost all the sediments were extremely low compared to
sediments in other locations in the United States and the relative weakness of non-parametric
correlations as statistical tools.

Sediment Quality Triad

Spearman rank correlations described above were used to make initial comparisons between
measures of the benthic invertebrate community to measures of sediment chemistry or overlying
water quality at the sampling stations. While rank correlation analysis can be used to demonstrate
association among variables, this ranking of data eliminates proportional relationships among
variables by re-ranking data to simple rank-order (e.g. 1,2,3,).  Thus, this ranking can not be used
to adequately evaluate dose response relationships. For these reasons, we evaluated benthic
community and laboratory toxicity data using a quadrant classification approach described below
(see also Canfield et al. 1996).
    Results of toxicity and chemistry evaluations of UMR sediments presented in Chapter 1
indicate these sediment samples were relatively uncontaminated compared to other locations in
the United States (Kemble et al. 1994; Ingersoll et al  1996). We used the sediment quality triad
approach in order to evaluate  how benthic communities sampled from the UMR compared to
other locations in the U.S. we previously have evaluated (Canfield et al.  1994, 1996).
    Scores for various benthic indices relating benthic alterations to contaminant levels were
previously identified using data sets from the Great Lakes (Canfield et al. 1996) and the Clark
Fork River in Montana (Canfield et al. 1994).  The scores were used to evaluate the scores for
samples from the current study.   In the present study, benthos samples were not classified as
impacted or non-impacted a priori,  but rather samples were considered to be classified as
"incorrect" only if the  scores were in the false positive or false negative error quadrants as
established in Canfield et al. (1996) (Figure 3.1).
    Four benthic indices (midge biotic index, midge richness, percent midge deformities, and
taxa richness) were previously found to provide some degree of discrimination among samples
from the  Great Lakes with differing degrees of contamination (Canfield et al. 1996). In the
present study, midge deformities (19%) had the smallest combined false positive and false
negative error rate relative to the sum of the ERM quotient score (Table 3.3). Midge oligochaete
ratio, midge biotic index, midge taxa richness, and total taxa richness had a combined false
positive and false negative error rate of 34% to 35%.   A benthos score required to obtain this
degree of discrimination was always > 75%.
    In addition to assessments using single indices, the combined scores of benthic indices were
evaluated which provided the  best classification (smallest combined false positive and false
negative error). Quadrant classification using the  sum of the ERM quotient score of two to three
combined benthic indices reduced the false positive and false negative error rate to 19 to 24% ,
which is less than all individually scored benthic indices except midge deformities (Table 3.3).
The various combinations of four to all six benthic indices were not included in or discussed since
the accuracy of classification did not increase with combinations of more than three benthic
                                           3.9
-------
indices (Table 3.3).  The combinations were restricted so that the benthos score required to
minimize false positive and false negative error was a score no greater than 80 to 81.  We were
unable to identify a combined score of less than 80 which minimized both false positive and false
negative error. The combined metric of midge oligochaete ratio, midge taxa richness and total
taxa richness provided the combination which had the lowest false positive and false negative
error of 19% (Table 3.3).
    Table 3.4 summarizes the classification of sediment samples based on exceedances of scores
for toxicity, chemistry, or benthos by quadrant analyses as  described in Canfield et al. (1996).
Twenty-one of 24 samples (88%) showed good agreement (i.e. all "minuses") among all three
measures of the Triad, which indicated that no contaminant induced degradation was observed
(Table 3.4). None of.the samples were scored with all pluses (i.e. evidence of contaminant
induced degradation). In one of the 24 (4%) samples laboratory toxicity and sediment chemistry
measures were in agreement, however the benthic component was not in accordance. Similarly in
one of the samples sediment chemistry and benthos response are in agreement, yet toxicity did not
occur. High concordance among laboratory toxicity, chemistry, and benthos is evidence that these
sediment samples from the UMR were relatively low in contamination or toxic effects compared
to other locations we have previously evaluated (Canfield et al. 1994, 1996).

Summary

Benthic invertebrate abundance values in sediment samples from the UMR were comparable to
values reported from relatively uncontaminated sediments. The percent composition of the
benthic invertebrate community also indicates a relatively healthy community compared to more
contaminated locations. Oligochaetes and chironomids constituted over 90% of the benthic
invertebrate communities collected in 10 of 24 samples from the UMR, which is expected given
the pre-dominance of soft sediments. However, most of the UMR pools had a relatively high
diversity of representatives from orders other than the oligochaetes and chironomids, which is
different from observations from other highly contaminated areas (Canfield et al.  1994, 1996).
Further benthic community indices were only weakly correlated with sediment contaminants.
    The occurrence of midge  deformities ranged from 0 to 13% in the UMR pool samples, which
were relatively low compared  to those chironomids from more highly contaminated sediments.
Sediment Quality Triad analyses classified a high percentage of the samples (88%) to be not
impacted.  These data indicate that these sediment samples were relatively uncontaminated.
    Additional studies are needed to evaluate specific contaminant, biotic, and abiotic factors
controlling benthic communities in soft sediments associated with backwater areas of both lotic
and lentic environments. Studies designed to evaluate benthic distributions in relation to factors
influencing variation on a local microhabitat scale are necessary in order to reduce the variation in
the relations between sediment chemistry, habitat,  and measures of benthic invertebrate
communities. These studies should greatly expand our ability to evaluate environmental quality of
ecosystems such as the UMR.
                                          3.10
-------
Acknowledgements The authors would like to thank Doug Hardesty, Shane Reussler, Jeff
Steevens and David Whites for all their field and laboratory help on this project. We would like
to thank James Fairchild, David Mount and Parley Winger for constructive criticism and review of
this manuscript. We thank Pamela Haverland and Ellen Eherhart for their statistical help in
developing the benthic metrics.
                                         3.11
-------
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                                         3.18
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