TOXICITY REDUCTION EVALUATION ACTIVITY REPORT THIRD QUARTER 2014

Transcription

1

2 TOXICITY REDUCTION EVALUATION ACTIVITY REPORT THIRD QUARTER 2014 UMETCO MINERALS CORPORATION FORMER WILSON MINE SITE OUTFALL 001 NPDES PERMIT NO. AR AFIN: Prepared for Umetco Minerals Corporation 2754 Compass Drive, Suite 280 Grand Junction, CO Prepared by FTN Associates, Ltd. 3 Innwood Circle, Suite 220 Little Rock, AR FTN No. R October 31, 2014

3 EXECUTIVE SUMMARY A toxicity reduction evaluation (TRE) was implemented to investigate potential causes of inconsistent chronic sublethal toxicity to the cladoceran Ceriodaphnia dubia at Umetco Minerals Corporation s Outfall 001. A TRE action plan for this work was approved by the Arkansas Department of Environmental Quality (ADEQ) on December 17, In correspondence from ADEQ approving the plan, ADEQ specified that quarterly activity reports will be due on the last day of the month for April, July, and October of 2014, and for January, April, and July The final report will be due November 2, TRE activities completed during the first quarter were reported in April 2014, and activities completed during the second quarter were reported in July Activities conducted during the third quarter of 2014 are described in the current report. Grab samples were collected August 11, 2014, from three locations: SWT10-10, which represents the Outfall 001 discharge; a sump at the south detention pond (SDP) representing seepage from the South Lecroy area that is pumped to the treatment facility; and effluent from the treatment facility prior to discharge to the settling pond. The grab samples were collected per the approved TRE action plan. The samples were sent to three aquatic toxicity testing laboratories for toxicity screening to evaluate factors affecting bioavailability (e.g., hardness of the water used as diluent/control) and potential sources of toxicity. Additional samples representing Outfall 001 were fortified with laboratory-specific food preparations used in chronic C. dubia tests. These fortified samples were used to assess the potential influence of food as another factor affecting bioavailability, which might account for interlaboratory variability in dose-response relationships observed in previous biomonitoring results. Results from screening tests on the sample collected from SWT10-10 indicated a modest level of sublethal toxicity that was consistent among the laboratories; however, the sublethal toxicity was not of sufficient magnitude (i.e., at least a 50% reduction in C. dubia reproduction as compared to the control) to justify further toxicity identification evaluation (TIE) follow-up testing. i

4 Testing of a neutralized sample collected from the South Lecroy area (SDP sample) indicates that the neutralized sample has the potential to cause sublethal toxicity in the Outfall 001 discharge if the neutralized input from the South Lecroy area comprises approximately 5% or more of the total volume of water treated and discharged from Outfall 001. Examination of available flow data indicates that the flows from the SDP in the South Lecroy area can comprise more than 11% of the Outfall 001 discharge over 50% of the time. This finding suggests that the contribution of water from the South Lecroy area to the Outfall 001 discharge can be sufficient to cause toxicity in Outfall 001. This possibility will be evaluated further in future testing. In some tests conducted during the third quarter, statistical significance of C. dubia reproduction depended on whether the comparison was based on control performance using the soft or moderately hard laboratory water. These results suggest that the use of soft water (as opposed to moderately hard water) as a control in routine biomonitoring tests may influence the statistical outcome of the comparison of the control to the critical dilution (100% effluent). However, at this point in the project, there are insufficient data available to fully evaluate this possibility. Results from tests to assess the potential influence of food as another factor affecting bioavailability indicated that differences in the concentration of dissolved organic carbon and total suspended solids due to the addition of food can vary by up to a factor of two between laboratories. Such differences in food composition, while compliant with method requirements, could introduce interlaboratory variability due to different effects on bioavailability. While not definitive, these results begin to suggest potential reasons (one or more) for the inconsistent and non-concentration-dependent results observed for the Outfall 001 discharge in previous biomonitoring. It is too early in the TRE to offer definitive conclusions regarding specific factors affecting bioavailability, interlaboratory variability, or potential sources/causes of toxicity. However, preliminary analyses indicate that the water from the South Lecroy area may have sufficient influence on the Outfall 001 discharge to cause episodes of sublethal toxicity. In accordance with the approved TRE action plan, routine screening of samples collected from ii

5 SWT10-10 will continue in order to evaluate interlaboratory variability and identify toxic samples for TIE follow-up. Similarly, sample analysis and testing will continue in order to further evaluate potential sources of toxicity and factors affecting bioavailability, including the potential confounding effects of artifacts associated with test methods (e.g., soft diluents, differences in food composition). iii

6 TABLE OF CONTENTS EXECUTIVE SUMMARY... i 1.0 INTRODUCTION Previous Findings Third Quarter 2014 Evaluations Routine Screening of the Discharge Evaluation of Factors Affecting Bioavailability Evaluation of Potential Toxicant Sources METHODS Description of Sampling Locations and Sample Collection Routine Screening of the Discharge Evaluation of Factors Affecting Bioavailability Evaluation of Potential Toxicant Sources RESULTS AND DISCUSSION Routine Screening of the Discharge Evaluation of Factors Affecting Bioavailability Evaluation of Potential Toxicant Sources EFFLP Sample SDP Sample CONCLUSIONS REFERENCES...15 iv

7 LIST OF TABLES Table 1 Table 2 Table 3 Table 4 Table 5 Sampling locations, volumes, and distribution for samples collected on August 11, Summary of percent survival in effluent exposures of chronic screening tests on SWT10-10 sample using C. dubia... 8 Summary of reproduction in effluent exposures of chronic screening tests on SWT10-10 sample using C. dubia... 8 Summary of percent survival in laboratory controls of chronic screening tests on SWT10-10 sample using C. dubia... 9 Summary of reproduction in laboratory controls of chronic screening tests on SWT10-10 sample using C. dubia... 9 Table 6 Results of the analysis of food-fortified SWT10-10 samples Table 7 Results of chronic screening tests on the EFFLP sample Table 8 Test results using the neutralized SDP sample Table 9 Summary of flows from the SDP in the South Lecroy area to the treatment facility expressed as a percent of Outfall 001 flows based on weekly averages LIST OF FIGURES Figure 1 Schematic diagram showing locational relationships and sample collection locations... 4 v

8 1.0 INTRODUCTION A toxicity reduction evaluation (TRE) was implemented to investigate potential causes of inconsistent chronic sublethal toxicity to the cladoceran Ceriodaphnia dubia at Umetco Minerals Corporation s Outfall 001. A TRE action plan for this work was approved by the Arkansas Department of Environmental Quality (ADEQ) on December 17, In correspondence from ADEQ approving the plan, ADEQ specified that quarterly activity reports will be due on the last day of the month for April, July, and October of 2014, and for January, April, and July The final report will be due November 2, TRE activities completed during the first quarter were reported in April 2014, and activities completed during the second quarter were reported in July Activities conducted during the third quarter of 2014 are described in the current report. 1.1 Previous Findings Results and findings through the second quarter of 2014 were as follows: 1. Results from toxicity tests conducted on spiked effluent samples indicated that it is unlikely that sublethal toxicity previously reported in testing of Outfall 001 was due to total dissolved solids (TDS) and/or sulfate (SO 4 ). 2. Results from toxicity testing of split samples collected from Outfall 001 and completed at two different laboratories suggested that laboratory variability is a factor that needs to be considered in evaluating previous reports of sublethal toxicity to C. dubia. 3. Evaluation of past biomonitoring test results illustrated an inverse gradient of increasing bioavailability and toxicity with increasing Outfall 001 sample dilution. This gradient could explain the flattened dose-response relationship that has been typical of routine biomonitoring tests showing sublethal toxicity. 4. Results from screening tests (single-exposure tests using undiluted effluent and a laboratory control) completed with SWT10-10 (representing Outfall 001) indicated a modest level of sublethal toxicity (15.5% to 26.4% reduction in reproduction in undiluted effluent relative to the control) that was consistent among the laboratories but was not of sufficient magnitude (i.e., at least a 50% reduction in C. dubia reproduction as compared to the control) to justify further toxicity identification evaluation (TIE) follow-up testing. 1

9 5. Results from tests to assess the potential influence of food as another factor affecting bioavailability indicated that differences in the concentration of dissolved organic carbon and total suspended solids due to the addition of food can vary by up to a factor of two between laboratories. 6. Insufficient data were available at the end of the second quarter of 2014 to draw conclusions regarding specific factors affecting bioavailability, interlaboratory variability, or potential sources/causes of toxicity. However, the inverse hardness/bioavailability gradient described above could result in a flattened dose-response relationship, which may help explain previously reported inconsistent and confounded results (i.e., non-concentration-dependent results observed for the Outfall 001 discharge). 1.2 Third Quarter 2014 Evaluations Grab samples were collected August 11, 2014, from three locations: SWT10-10, which represents the Outfall 001 discharge; a sump at the south detention pond (SDP) representing seepage from the South Lecroy area that is pumped to the treatment facility; and effluent from the treatment facility prior to discharge to the settling pond (EFFLP). This evaluation focused on three topics: (1) routine screening of the discharge, (2) evaluation of factors affecting bioavailability, and (3) evaluation of potential toxicant sources Routine Screening of the Discharge Per the approved study plan, samples were collected from the discharge and screened for toxicity using three different laboratories 1. The purpose of this testing is to evaluate interlaboratory variability and to identify samples with sufficient toxicity to warrant TIE follow-up Evaluation of Factors Affecting Bioavailability Previous testing and analysis identified significant differences in the level of toxicity in Outfall 001 samples in tests conducted at different laboratories. One factor in biomonitoring tests using C. dubia that has the potential to influence test results is the daily addition of food to the 1 ENVIRON International Corp., 201 Summit View, Suite 300, Brentwood, TN 37027; Atkins North America, Inc., Environmental Toxicology Laboratory, 888 West Sam Houston Parkway, Houston, TX 77042; American Interplex Corporation Laboratories (AIC), 8600 Kanis Road, Little Rock, AR

10 test chambers, which adds binding capacity in the form of organic carbon (dissolved and total) and suspended solids. A working hypothesis was developed proposing that differences in toxicity tests among laboratories could result from interlaboratory differences in the binding capacity of the food used for C. dubia. These differences could also potentially explain flat dose-response curves if the binding capacity of food used by a particular laboratory was insufficient to overcome the bioavailability gradient that is present due to the use of laboratory water as diluent. As a first step toward evaluating this possibility, testing for this quarter included analysis of total organic carbon (TOC), dissolved organic carbon (DOC), and total suspended solids (TSS) in the food suspensions added to test exposures by the three laboratories participating in this study Evaluation of Potential Toxicant Sources Stormwater runoff and seepage from the upper Wilson Creek watershed along with seepage pumped from the SDP in the South Lecroy area are treated using neutralization with hydrated lime, air oxidation, precipitation, and settling. Settling occurs in East Wilson Pond, which receives the treated discharge from the treatment plant. Outfall 001 discharges from East Wilson Pond to lower Wilson Creek (Figure 1). Water from the South Lecroy area is a minor component of the total inflow to the treatment plant. The source evaluation for this quarter focused on the water that is pumped to the treatment facility from the South Lecroy area and on the discharge from the treatment facility to East Wilson Pond. Samples collected from the SDP location (Figure 1) represent water that is pumped from the South Lecroy area to the treatment facility. Samples collected from the EFFLP location (Figure 1) represent the combination of water from the South Lecroy area and upper Wilson Creek that has been treated and discharged to East Wilson Pond. 3

11 Figure 1. Schematic diagram showing locational relationships and sample collection locations. 4

12 2.0 METHODS 2.1 Description of Sampling Locations and Sample Collection Samples were collected as grab samples from all locations (see Figure 1) on August 11, The samples shipped to ENVIRON and Atkins were shipped on ice with chain-of-custody (COC) documentation via overnight carrier on August 11, FTN delivered the remaining samples to AIC. Sampling locations, volumes, distribution, and testing of samples are summarized in Table 1. Table 1. Sampling locations, volumes, and distribution for samples collected on August 11, Sampling Location 10 m downstream of Outfall 001 NPDES compliance point Sump at south detention pond that collects seepage from the South Lecroy area before being pumped to the treatment facility Effluent from treatment facility immediately after ph adjustment and aeration but before settling Sample Designation Volume Collected (L) SWT Laboratory AIC ENVIRON Atkins SDP 9 ENVIRON EFFLP 9 ENVIRON *In accordance with guidelines set forth in US Environmental Protection Agency (EPA) Sample Manipulations and Tests to be Performed Single-dilution (100%) chronic screening toxicity tests* using C. dubia. Single-dilution (100%) chronic screening toxicity tests* using C. dubia. TIE follow-up as appropriate. Single-dilution (100%) chronic screening toxicity tests* using C. dubia. Adjust sample ph to 7.0 su with sodium hydroxide (NaOH). Filter through glass fiber filter. Perform range-finding test as needed using C. dubia to identify a range of toxic and non-toxic concentrations to be used in a chronic multi-dilution toxicity test. Perform multi-dilution chronic toxicity tests as needed (C. dubia) based on results of range-finding test to determine a non-toxic concentration/proportion of the SDP sample. Allow precipitated solids to settle for 2 hours. Collect supernatant and perform single-dilution (100%) chronic screening toxicity tests using C. dubia. 5

13 Sampling location SWT10-10 (Figure 1) refers to a location downstream of the National Pollutant Discharge Elimination System (NPDES) permit compliance point for Outfall 001 that is representative of the Outfall 001 discharge. The EFFLP sampling location in the treatment facility is located immediately after the addition of hydrated lime and air oxidation but before the treated stream has had time to settle. The SDP sample was collected from a sump near the SDP that collects seepage from the South Lecroy area for pumping to the treatment facility. 2.2 Routine Screening of the Discharge Sample collected from SWT10-10 was split into aliquots for shipment to each of the three laboratories involved in the study. COC forms and laboratory coordination included instructions regarding when to begin testing for each laboratory such that each laboratory began tests on the same day (to allow for comparable holding times among laboratories). Toxicity tests were single-dilution (100%) chronic screening toxicity tests using C. dubia (Method , EPA 2002) and included laboratory controls using both moderately hard and soft water. 2.3 Evaluation of Factors Affecting Bioavailability To obtain preliminary information to evaluate potential bioavailability factors in the C. dubia food preparations and in the Outfall 001 discharge, laboratories were instructed as follows. Upon sample arrival, each laboratory added an amount of C. dubia food to 1 L of SWT10-10 sample equivalent to the amount that would be present in a chronic test exposure. Atkins and ENVIRON shipped their food-fortified samples to FTN, who delivered the samples to AIC for testing. COC forms and laboratory coordination included instructions regarding preparation and shipment of the food-fortified sample such that the holding times for samples from all three laboratories would be comparable. Samples were analyzed for TOC, DOC (EPA Method 415), and TSS (EPA Method 160.2). For purposes of this evaluation, the DOC sample was filtered through a 0.1-µm pore-size membrane filter instead of the 0.45-µm pore size that is typically used for DOC analysis. This modification was chosen to eliminate organic carbon due to bacteria from the DOC fraction. Sample filtration and analyses were performed by AIC. 6

14 2.4 Evaluation of Potential Toxicant Sources The EFFLP sample was shipped to ENVIRON with instructions to allow the solids in the sample to settle (Table 1) before decanting the supernatant, which would be used for the toxicity test. The toxicity test was a single-dilution (100%) chronic screening toxicity test using C. dubia (Method , EPA 2002) with a moderately hard laboratory water control. The SDP sample was shipped to ENVIRON. Because water from the South Lecroy area is known to show low ph and high TDS, the laboratory was instructed to adjust the ph to neutrality with NaOH (Table 1) and filter through a glass fiber filter before using the sample for toxicity testing. A single aliquot of sample was prepared in a quantity sufficient to perform the entire test. The objective of the toxicity testing was to estimate the concentration/proportion of neutralized SDP sample that would cause toxicity in chronic tests using C. dubia. 7

15 3.0 RESULTS AND DISCUSSION For ease of comparison, results from this quarter are summarized and discussed along with results of previous testing, as appropriate. 3.1 Routine Screening of the Discharge Results of chronic screening tests on SWT10-10 samples collected on June 26 and August 11 (for the second and third quarters of 2014, respectively) are summarized in Tables 2 through 5. Survival and reproduction in the effluent exposures are presented in Tables 2 and 3, respectively. Survival and reproduction in the laboratory controls are presented in Tables 4 and 5, respectively. Table 2. Summary of percent survival (n = 10) in effluent exposures of chronic screening tests on SWT10-10 sample using C. dubia. Sampling Date AIC Atkins ENVIRON June 26, August 11, * * Statistically less (P < 0.05) than both the moderately hard and soft water control. Table 3. Summary of reproduction (average three-brood neonate production per female) in effluent exposures of chronic screening tests on SWT10-10 sample using C. dubia. Sampling Date AIC Atkins ENVIRON June 26, (a) 31.4 (a) 18.4 (b) August 11, (c) 24.1 (c) 26.9 (c) Notes: a. Statistically less (P < 0.05) than the moderately hard water control only (no soft water control included). b. Statistically less (P < 0.05) than the soft water control only. c. Statistically less (P < 0.05) than both the soft and moderately hard water controls. 8

16 Table 4. Summary of percent survival (n = 10) in laboratory controls of chronic screening tests on SWT10-10 sample using C. dubia. Sampling Date Water Type AIC Atkins ENVIRON June 26, 2014 Moderately Hard Soft ND ND 100 August 11, 2014 Moderately Hard Soft ND = No data; test not performed. Table 5. Summary of reproduction (average three-brood neonate production per female) in laboratory controls of chronic screening tests on SWT10-10 sample using C. dubia. Sampling Date Water Type AIC Atkins ENVIRON June 26, 2014 Moderately Hard Soft ND ND 25.5 August 11, 2014 Moderately Hard Soft ND = No data; test not performed. Sublethal effects observed with the sample collected on June 26, 2014 (as part of the second quarter 2014 TRE activities) were not of sufficient magnitude (i.e., at least a 50% reduction in C. dubia reproduction as compared to the control) or consistency among laboratories to warrant additional follow-up testing using TIE procedures. Neither AIC nor Atkins observed the marginally significant lethal effect that was observed by ENVIRON using the sample collected on August 11, In the judgment of the project personnel, the lethal effect was not a reliable result based on the timing of the organism deaths (at or shortly after the release of the third brood). Although results obtained by Atkins and AIC also indicated sublethal effects, they were not of sufficient magnitude to warrant additional follow-up testing using TIE procedures. Accordingly, no TIE follow-up testing has been conducted to date due to the absence of sufficient levels of toxicity. The results from the sample collected on August 11, 2014, showed the same statistical results for C. dubia reproduction (in terms of null hypothesis rejection) using either the 9

17 moderately hard or soft water control for all three laboratories. In contrast, the ENVIRON results from the sample collected on June 26, 2014, showed statistical significance only with the soft water control (no soft water controls were included in the tests performed at AIC and Atkins for the June 26 sample). This result suggests that the use of soft water as a diluent/control in routine biomonitoring may influence the statistical outcome of the tests. However, at this point in the project, there are insufficient data available to fully evaluate this possibility. 3.2 Evaluation of Factors Affecting Bioavailability Results of the analysis of food-fortified SWT10-10 samples are provided in Table 6. Food-fortified samples contained higher TOC, DOC, and TSS than the sample without food. Food-fortified samples prepared by AIC and ENVIRON show similar TOC, DOC, and TSS levels, which are approximately two times the levels measured in the sample provided by Atkins. The relative differences among laboratories were similar between sampling dates, which indicates that the holding time dissimilarities among the samples collected on June 26, 2014, may not have strongly influenced those results. These analyses will be continued until such time as the investigating team concludes that sufficient information has been collected to evaluate the differences among laboratories and the potential influence on whole effluent toxicity (WET) test results. Table 6. Results of the analysis of food-fortified SWT10-10 samples. Sample Collection Date Food-Fortified Sample Source TSS (mg/l) TOC (mg/l) DOC (mg/l) AIC June 26, 2014 * Atkins ENVIRON Effluent without food < AIC August 11, 2014 Atkins ENVIRON Effluent without food < *Holding times not comparable among laboratories. 10

18 3.3 Evaluation of Potential Toxicant Sources EFFLP Sample Results of the toxicity tests for the EFFLP sample are provided in Table 7. Tests on these samples were conducted only at ENVIRON. The test on the sample collected on August 11 showed the same unreliable effects on survival that were seen with the SWT10-10 sample (Table 2). The data available at this point in the project do not allow evaluation of the discharge from the EFFLP location as a source of toxicity. Table 7. Results of chronic screening tests on the EFFLP sample. Sample Collection Date Percent Survival (n = 10) Average 3-Brood Neonate Production Per Female June 26, August 11, * 25.6* Note: Corresponding controls are the same as those given in Tables 5 and 6. * Statistically less (P < 0.05) than both the soft and moderately hard controls (Table 6) SDP Sample Results of the toxicity tests for the neutralized SDP sample are provided in Table 8. Neutralization of the SDP sample with NaOH to circumneutral ph produces an iron flocculent, which was filtered out of the sample prior to testing (Table 1). Based on the two SDP samples collected to date, the concentration/proportion of neutralized sample from the SDP in the discharge that could cause toxicity in chronic tests using C. dubia is approximately 5% to 10%. The potential implications of this result can be evaluated by comparing the flows from the SDP to the treatment facility with the flows from Outfall 001. Table 9 provides a summary of the SDP flow to the treatment facility expressed as a percent of Outfall 001 flows based on weekly averages. The summary shows that the flows from the SDP in the South Lecroy area can comprise more than 11% of the discharge from the outfall approximately 50% of the time. This finding suggests that the contribution to the total discharge attributable to the South Lecroy area can be sufficient to cause toxicity. 11

19 Table 8. Test results using the neutralized SDP sample. Sample Collection Date Test Concentration (percent neutralized SDP sample) Percent Survival (n = 10) Average 3-Brood Neonate Production Per Female Laboratory Control June 26, * Laboratory Control August 11, * * *Statistically less (P < 0.05) than control. Table 9. Summary of flows from the SDP in the South Lecroy area to the treatment facility expressed as a percent of Outfall 001 flows based on weekly averages. Summary Statistic Percent of Outfall 001 from South Lecroy Area* Percentile Minimum 0 Average 12 Maximum 52 * Period of record from July 2013 through August Flow data for Outfall 001 and sampling location representing flow pumped from the SDP to the treatment plant provided by CH2M Hill. 12

20 4.0 CONCLUSIONS Results from screening tests on the sample collected from SWT10-10 indicated a modest level of sublethal toxicity that was consistent among the laboratories; however, the sublethal toxicity was not of sufficient magnitude (i.e., at least a 50% reduction in C. dubia reproduction as compared to the control) to justify further TIE follow-up testing. Testing of a neutralized sample collected from the South Lecroy area (SDP sample) indicates that the neutralized sample has the potential to cause sublethal toxicity in the Outfall 001 discharge if the neutralized input from the South Lecroy area comprises approximately 5% or more of the total volume of water treated and discharged from Outfall 001. Examination of available flow data indicates that the flows from the SDP in the South Lecroy area can comprise more than 11% of the Outfall 001 discharge approximately 50% of the time. This finding suggests that the contribution of water from the South Lecroy area to the Outfall 001 discharge can be sufficient to cause toxicity in Outfall 001. This possibility will be evaluated further in future testing. In some tests conducted during the third quarter, statistical significance of C. dubia reproduction depended on whether the comparison was based on control performance using the soft or moderately hard laboratory water. These results suggest that the use of soft water (as opposed to moderately hard water) as a control in routine biomonitoring tests may influence the statistical outcome of the comparison of the control to the critical dilution (100% effluent). However, at this point in the project, there are insufficient data available to fully evaluate this possibility. Results from tests to assess the potential influence of food as another factor affecting bioavailability indicated that differences in the concentration of DOC and TSS due to the addition of food can vary by up to a factor of two between laboratories. Such differences in food composition, while compliant with method requirements, could introduce interlaboratory variability due to different effects on bioavailability. While not definitive, these results begin to suggest potential reasons (one or more) for the inconsistent and non-concentration-dependent results observed for the Outfall 001 discharge in previous biomonitoring. 13

21 It is too early in the TRE to offer definitive conclusions regarding specific factors affecting bioavailability, interlaboratory variability, or potential sources/causes of toxicity. However, preliminary analyses indicate that the water from the South Lecroy area may have sufficient influence on the Outfall 001 discharge to cause episodes of sublethal toxicity. In accordance with the approved TRE action plan, routine screening of samples collected from SWT10-10 will continue in order to evaluate interlaboratory variability and identify toxic samples for TIE follow-up. Similarly, sample analysis and testing will continue in order to further evaluate potential sources of toxicity and factors affecting bioavailability, including the potential confounding effects of artifacts associated with test methods (e.g., soft diluents, differences in food composition). 14

22 5.0 REFERENCES EPA Short-term methods for estimating the chronic toxicity of effluents and receiving waters to freshwater organisms, fourth edition [EPA-821-R ]. Washington, DC: US Environmental Protection Agency, Office of Water. 15