Reproductive effects in fathead minnows (Pimphales promelas) following a 21 d exposure to 17α-ethinylestradiol.

17α-ethinylestradiol (EE2) is a synthetic estrogen that is an active ingredient in oral contraception and hormone replacement therapy. Surveys of wastewater treatment plant effluents and surface waters throughout the world have reported EE2 concentrations in the ng/L range, and these low levels can cause significant reproductive effects in fish. This study tested the effects of three environmentally relevant EE2 concentrations: 0.47, 1.54 and 3.92 ng/L using a 21 d short-term reproductive assay to investigate the effects of EE2 on fathead minnow (Pimephales promelas) reproduction. The two highest EE2 concentrations tested in this study caused significant liver gene expression and induction of vitellogenin plasma protein in male fathead minnows. Exposure to 3.92 ng EE2/L increased the production of plasma vitellogenin in the females. Plasma estradiol concentrations were significantly reduced in females exposed to 1.54 and 3.92 ng EE2/L. All three tested concentrations significantly reduced fathead minnow egg production after a 21 d exposure to EE2. The results of this study indicate that the previously reported no observed adverse effect concentration (NOAEC) for EE2 on fathead minnow egg production (1.0 ng/L) may be too high. Because all three treatments resulted in significantly reduced egg production, the lowest observed adverse effect concentration (LOAEC) for EE2 on fathead minnow egg production is 0.47 ng EE2/L. This research estimates a NOAEC for fathead minnow reproduction at 0.24 ng EE2/L following a 21 d exposure. Additionally, induction of vitellogenin is a sensitive indicator of estrogen exposure but does not appear to be predictive of fathead minnow egg production.

Determining the effects of a mixture of an endocrine disrupting compound, 17α-ethinylestradiol, and ammonia on fathead minnow (Pimephales promelas) reproduction.

Aquatic organisms are exposed to a multitude of contaminants and to fully understand the impact of multiple stressors on fish populations, we must first understand the mechanism of action for each toxicant and how the combined effects manifest at the level of the individual. 17α-ethinylestradiol (EE2) has been known to cause adverse reproductive effects including reduced fecundity and fertility, intersex and skewed sex ratios in fish by mimicking naturally produced estrogen at low concentrations. Ammonia can cause adverse reproductive and mortality effects in individual fish through effects or damage to the central nervous system. Both EE2 and ammonia are found in most municipal effluents in various concentrations. A flow-through diluter system was used to test the individual effects of these two contaminants at their respective no observable adverse effect concentration (NOAEC) as well as their combined effects on fathead minnow, (Pimephales promelas) reproduction in a mixture exposure. While neither contaminant nor their mixture altered reproduction in terms of fecundity, their mixture resulted in significant fathead minnow mortality during a 21 d exposure. This study demonstrated the need to consider mixture effects when assessing risk for toxicity testing with multiple stressors.

Determining the effects of ammonia on fathead minnow (Pimephales promelas) reproduction.

Ammonia can cause adverse reproductive and mortality effects in individual fish by interacting with the central nervous system. The last published study that assessed the effects of ammonia on fathead minnow reproduction was a lifecycle study conducted in 1986. Our study's main goal was to re-evaluate ammonia toxicity on fathead minnow Pimephales promelas reproduction using a 20-day fecundity flow-through diluter method. Flow-through diluter systems have been used by regulatory agencies, such as the U.S. Environmental Protection Agency, in the past as an effective way to estimate acceptable levels of contaminants. There was a significant difference in cumulative egg production among treatments (ANOVA; F=10.167, p≤0.01, df=3). All three concentrations of ammonia tested in this study significantly reduced fecundity after 20days of exposure (Dunnett's, p≤0.05 for each treatment). The lowest un-ionized ammonia concentration (0.06mg/L at a pH of 7.3 and temperature of 25.1°C) tested during this study resulted in a 29% decrease in cumulative fecundity. Because all tested ammonia concentrations caused an effect on P. promelas reproduction, the no effect concentration was estimated to be 0.025mg/L un-ionized ammonia (2.19mg/L total ammonia-nitrogen). This estimate was determined using the U.S. EPA Toxicity Relationship Analysis Program to calculate the 10% effect concentration of ammonia on P. promelas reproduction. This value is much lower than the previous reported no effect concentration on P. promelas reproduction (0.37mg/L un-ionized ammonia or 6.43mg/L total ammonia-nitrogen) as determined from the 1986 study, which was used to determine the ammonia water quality criteria by the U.S. Environmental Protection Agency. Our results should be considered in the next revision of water quality criteria.

An interlaboratory comparison of sediment elutriate preparation and toxicity test methods.

Elutriate bioassays are among numerous methods that exist for assessing the potential toxicity of sediments in aquatic systems. In this study, interlaboratory results were compared from 96-h Ceriodaphnia dubia and Pimephales promelas static-renewal acute toxicity tests conducted independently by two laboratories using elutriate samples prepared from the same sediment. The goal of the study was to determine if the results from the elutriate tests were comparable between two U.S. Environmental Protection Agency (USEPA) laboratories when different elutriate preparation procedures were employed by each lab. Complete agreement in site characterization was attained in 22 of the 25 samples for both bioassays amongst each lab. Of the 25 samples analyzed, 10 were found to be toxic to at least one of the species tested by either laboratory. The C. dubia elutriate tests conducted by the National Exposure Research Laboratory (NERL) indicated that 7 of the 25 sediment samples were toxic, while 8 sediment samples were characterized as such in testing conducted by USEPA Region 6 (Region 6). The P. promelas elutriate tests conducted by NERL determined 8 samples as toxic, while Region 6 tests displayed toxicity in 5 of the samples. McNemar's test of symmetry for C. dubia (S = 0.33, p = 0.5637) and P. promelas (S = 3.0, p = 0.0833) tests indicated no significant differences in designating a site toxic between NERL and Region 6 laboratories. Likewise, Cohen's kappa test revealed significant agreement between NERL and Region 6 C. dubia (K = 0.7148, p < 0.01) and P. promelas (K = 0.6939, p < 0.01) elutriate tests. The authors conclude that differences in interlaboratory elutriate preparation procedures have no bearing on the ability of either the C. dubia or P. promelas bioassay testing methods to detect toxicity while yielding similar results.

Sediment toxicity in mid-continent great rivers (USA).

As part of the Environmental Monitoring and Assessment Program for Great River Ecosystems (EMAP-GRE), sediment samples were collected from 447 randomly selected littoral sites along the main channels of the Ohio, Missouri, and Upper Mississippi Rivers between 2004 and 2006. Toxicity of these sediment samples was measured using a 7-day Hyalella azteca survival and growth test. Sixty-five sites (14.5%) exhibited lethal toxicity, and 130 sites (29.1%) exhibited decreased growth. In the EMAP-GRE probabilistic sampling design, each sampled site had a weight associated with it that determined the length (and proportion) of the river represented by that sample point in the population. Weighted whole-river estimates indicated that of the 4721 river km sampled, sediment from 15.9 ± 3.0% of the river (752 ± 50 km) were lethally toxic, 27.4 ± 3.5% (1289 ± 57 km) were toxic by way of growth inhibition, and 40.0 ± 3.7% (1887 ± 68 km) exhibited either lethal or growth toxicity. Selected toxic samples were analyzed for 21 pesticides, 20 polychlorinated biphenyl congeners, and 6 polybrominated diphenyl ether congeners. For all of the samples tested, the concentration levels of these analytes were mostly lower than known toxicity thresholds, and neither unionized ammonia concentration nor osmotic stress (as measured by conductivity) could account for the toxicity found in sediments. The spatial pattern of sediment toxicity cannot be readily explained by urbanization or agricultural land use at the subcatchment scale. We speculate that the distribution of toxic sediment is more likely due to a combination of localized sources, including polluted tributaries, and the redistribution of contaminated sediments from upriver. The sediment toxicity results from this study will be used, in combination with other sediment, biologic, and habitat metrics and indicators collected in the EMAP-GRE study, to help interpret and assess the condition of the Ohio, Upper Mississippi, and Missouri Rivers.

Comparison of bulk sediment and sediment elutriate toxicity testing methods.

Numerous methods exist for assessing the potential toxicity of sediments in aquatic systems. In this study, the results from 10-day bulk sediment toxicity test methods using Hyalella azteca and Chironomus tentans were compared to results from 96-h Pimephales promelas and Ceriodaphnia dubia renewed acute toxicity tests conducted using elutriate samples prepared from the same sediments. The goal of the study was to determine if the results from the elutriate tests were comparable to those obtained from the bulk sediment tests. Of the 25 samples analyzed, 16 were found to be toxic to at least one of the species tested, in either elutriate or bulk sediment tests. The C. tentans 10-day bulk sediment test was the most sensitive, with 12 sediment samples exhibiting toxicity to this species, whereas the H. azteca bulk sediment test and C. dubia 96-h elutriate test were the least sensitive, exhibiting toxicity in only 7 of the 25 sediments tested. The P. promelas elutriate test found 8 of the 25 sediments to be toxic. Based on the total number of sites found to show toxicity, results from testing indicate 96-h elutriate tests show a level of sensitivity comparable to 10-day bulk sediment tests in assessing toxicity quantitatively. However, the methods did not always find toxicity at the same sites, suggesting that the ability of elutriate tests to predict toxicity (quantitatively) is not statistically correlated with bulk sediment methods. This would indicate that a suite of toxicity test methods would provide the most complete measure of site condition; however, in circumstances where bulk sediment testing is not feasible, elutriate tests can provide a practical and credible alternative for toxicity assessment.

Development and validation of a Daphnia magna four-day survival and growth test method.

Zooplankton are an important part of the aquatic ecology of all lakes and streams. As a result, numerous methods have been developed to assess the quality of waterbodies using various zooplankton species. Included in these is the freshwater species Daphnia magna. Current test methods using D. magna involve acute lethality test methods ranging from 24 to 96 h in duration and chronic test methods with durations of 21 to 28 d. Whereas the current acute and chronic test methods are useful, a need exists for a shorter-duration test method that will provide a chronic or subchronic endpoint with this, species. In the present study, a 4-d, static-renewal survival and growth test was developed for use with D. magna. The test results were compared to performance criteria and results from 7-d survival and reproduction tests with Ceriodaphnia dubia to determine the level of comparability between the two methods. Results from the 4-d D. magna survival and growth test method indicated that this method will produce consistent results with various reference toxicant materials and provide data that are both reproducible and useful for detecting potential toxicity in aquatic environments.