Adult fathead minnows were exposed to dilutions of a historically estrogenic wastewater treatment plant effluent in a 21-d reproduction study. This dataset is comprised of a variety of endpoints representing key events along adverse outcome pathways linking estrogen receptor activation and other molecular initiating events to reproductive impairment. This study demonstrates the value of using an integrative approach that encompasses analytical chemistry, in vitro bioassays, and in vivo apical and pathway-based approaches with endpoints spanning from molecular- (e.g., gene expression) to organismal- (e.g., reproduction) levels of biological organization to help infer causal relationships between chemistry and potential effects on reproduction. This dataset is associated with the following publication: Cavallin , J., K. Jensen , M. Kahl , D. Villeneuve , K. Lee, A. Schroeder , J. Mayasich, E. Eid, K. Nelson, R. Milsk, B. Blackwell, J. Berninger , C. LaLone, C. Blanksma, T. Jicha , C. Elonen , R. Johnson , and G. Ankley. Pathway-based approaches for assessment of real-time exposure to an estrogenic wastewater treatment plant effluent on fathead minnow reproduction. ENVIRONMENTAL TOXICOLOGY AND CHEMISTRY. Society of Environmental Toxicology and Chemistry, Pensacola, FL, USA, 35(3): 702-716, (2016).

The purpose of this study was twofold. First, we sought to identify candidate markers of exposure to antiandrogens by analyzing endogenous metabolite profiles in the urine of male fathead minnows (mFHM, Pimephales promelas). Based on earlier work, we hypothesized that unidentified lipids in the urine of mFHM were selectively responsive to exposure to androgen receptor antagonists,which is otherwise difficult to confirm using established fish toxicity assays. A second goal was to evaluate the feasibility of non-lethally and repeatedly sampling urine from individual mFHMs over the time course of response to a chemical exposure. Accordingly, we exposed mFHM to the model anti-androgens vinclozolin or flutamide. Urine was collected from each fish at 48 hour intervals over the course of a 14 day exposure. Parallel experiments were conducted with mFHM exposed to bisphenol A or control water. The frequent handling/sampling regime did not cause apparent adverse effects on the fish. Endogenous metabolite profiling was conducted with gas chromatography–mass spectrometry (GC–MS), which exhibited lower variation for the urinary metabolome than was found in earlier work with nuclear magnetic resonance (NMR) spectroscopy. Specifically, for inter- and intra-individual variations, the median spectrum-wide relative standard deviation (RSD) was 32.6% and 33.3%, respectively, for GC–MS analysis of urine from unexposed mFHM. These results compared favorably with similar measurements of urine from other model species, including the Sprague Dawley rat. In addition, GC–MS allowed us to identify several lipids (e.g., certain saturated fatty acids) in mFHM urine as candidate markers of exposure to androgen receptor antagonists. The dataset that is uploaded here is the complete processed data from GC-MS instrument. This dataset is associated with the following publication: Collette , T., D. Skelton, J. Davis , J. Cavallin , K. Jensen , M. Kahl , G. Ankley , G. Ankley , D. Martinovic-Weigelt, and D. Ekman. Metabolite profiles of repeatedly sampled urine from male fathead minnows (Pimephales promelas) contain unique lipid signatures following exposure to anti-androgens. COMPARATIVE BIOCHEMISTRY AND PHYSIOLOGY - PART D: GENOMICS AND PROTEOMICS. Elsevier BV, AMSTERDAM, NETHERLANDS, 19: 190-198, (2016).

Aromatase inhibition is one of the chemical modes of action of concern to EPA's Endocrine Disruptor Screening Program (EDSP). In vitro bioassays that can detect aromatase inhibition are part of both the EDSP tier 1 screening program and are included subset of ToxCast assays employed for EDSP21 screening. An adverse outcome pathway (AOP) linking aromatase inhibition to reproductive dysfunction in fish has been described and endorsed by the OECD, establishing a scientifically sound connection between aromatase inhibition and adverse apical outcomes relevant to risk assessment and regulatory decision-making. Further, computational models that allow for quantitative prediction of dose-response time-course behaviors and the potential severity of the adverse outcome based on in vitro screening data have been developed. The present study provides further weight of evidence to support this AOP and its use in regulatory decision-making. In particular, it identifies rapid responses to aromatase inhibition that can be expected to occur within the first 24 h of exposure, examines the dynamic stability of gene expression responses over that period to help identify appropriate time periods in which characteristic gene expression responses may serve as effective biomarkers of exposure to aromatase inhibitors, and provides insights into different gene regulatory mechanisms that may be operating over the first few hours of exposure versus more systemic endocrine-related regulation that appear to take over after 6-12 h of exposure. These data continue to refine our understanding of this important mode of endocrine disruption and how to more efficiently and effectively both model and test for it to support regulatory decision-making. This dataset is associated with the following publication: Schroeder, A., G. Ankley, T. Habib, N. Garcia-Reyero, B. Escalon, K. Jensen, M. Kahl, E. Durhan, E. Makynen, J. Cavallin, D. Martinovic-Weigelt, E. Perkins, and D. Villeneuve. Rapid effects of the aromatase inhibitor fadrozole on steroid production and gene expression in the ovary of female fathead minnows (Pimephales promelas). GENERAL AND COMPARATIVE ENDOCRINOLOGY. Academic Press Incorporated, Orlando, FL, USA, 252: 79-87, (2017).

High-resolution mass spectrometry is advantageous for monitoring physiological impacts and contaminant biotransformation products in fish exposed to complex wastewater effluent. We evaluated this technique using skin mucus from male and female fathead minnows (Pimephales promelas) exposed to control water or treated wastewater effluent at 5%, 20%, and 100% levels for 21 d, using an onsite, flow-through system providing real-time exposure. Both sex-specific and non-sex-specific responses were observed in the mucus metabolome, the latter suggesting the induction of general compensatory pathways for xenobiotic exposures. Altogether, 85 statistically significant treatment-dependent metabolite changes were observed and 30 of those annotated with probable structures. The mummichog software package was used to elucidate impacted biochemical pathways and enhance metabolite annotation. Partial least squares regression models revealed relationships between the mucus metabolomes and upregulated hepatic mRNA transcripts reported previously for these same fish. These regression models suggest that mucus metabolomic changes reflected, in part, processes by which the fish biotransformed xenobiotics in the effluent. Further, we detected a phase II transformation product of bisphenol A in the skin mucus of male fish. Collectively, these findings demonstrate the utility of mucus as a minimally invasive matrix for simultaneously assessing exposures and effects of real-world mixtures of contaminants. This dataset is associated with the following publication: Mosley, J., D. Ekman, J.E. Cavallin, D. Villeneuve, G. Ankley, and T. Collette. High‐resolution mass spectrometry of skin mucus for monitoring physiological impacts and contaminant biotransformation products in fathead minnows exposed to wastewater effluent. ENVIRONMENTAL TOXICOLOGY AND CHEMISTRY. Society of Environmental Toxicology and Chemistry, Pensacola, FL, USA, 37(3): 788-796, (2018).

This data set describes experiments that were conducted to investigate whether exposure of male fathead minnows to environmentally-relevant estrone concentrations would result in elevated plasma 17β-estradiol concentrations in the fish. Secondly, we sought to establish whether observed elevations in plasma 17β-estradiol occurred as a result of the conversion of external estrone by the fish using an approach involving exposure of the fish to 13C-labeled estrone. Endpoints reported in the dataset include plasma 17β-estradiol and estrone concentrations, plasma vitellogenin concentrations, hepatic vitellogenin mRNA, 13C-labeled plasma 17β-estradiol and estrone concentrations, and testicular and/or hepatic expression of aromatase and several hydroxysteroid dehydrogenases involved in estrone metabolism. This dataset is associated with the following publication: Ankley, G., D. Feifarek, B. Blackwell, J. Cavallin, K. Jensen, M. Kahl, S. Poole, E. Randolph, T. Saari, and D. Villeneuve. Reevaluating the significance of estrone as an environmental estrogen (article). ENVIRONMENTAL SCIENCE & TECHNOLOGY. American Chemical Society, Washington, DC, USA, 51: 4705-4713, (2017).

This dataset provides exposure and effects data for two experiments regarding the dietborne toxicity of inorganic arsenic on fathead minnows survival and growth. There are separate spreadsheets for the two experiments, at the replicate exposure chamber level, of fish exposure, survival, and weights and calculated growth metrics. Each spreadsheet provides footnote descriptions of the data and calculations provided. Descriptions of experimental protocols are also provided as supporting documents. This dataset is associated with the following publication: Erickson, R., D. Mount, T. Highland, R. Hockett, D. Hoff, C. Jenson, and T. Lahren. The relative importance of waterborne and dietborne As exposure on survival and growth of juvenile fathead minnows. AQUATIC TOXICOLOGY. Elsevier Science Ltd, New York, NY, USA, 211: 18-28, (2019).

Fathead minnows (Pimephales promelas) were exposed to 100 ug/L indomethacin, 200 ug/L ibuprofen, or 20 ug/L celecoxib for 96 h. Effects on cycloxygenase enzyme activity in ovary, prostaglandin F2alpha concentrations in plasma, 17beta-estradiol concentrations in plasma, and vitellogenin concentrations in plasma were measured. Gene expression in ovary samples was evaluated using a 15,000 probe oligonucleotide microarray. Transcriptomics data (raw data and normalized) are available through the National Center for Biotechnology Information, Gene Expression Omnibus (GEO), accession number GSE72976. Metabolite profiles in liver tissue were measured by proton nuclear magnetic resonance. In addition to these data, the data set also contains identification of differentially expressed genes, pathway enrichment and gene set enrichment analyes, ToxCast data for indomethacin and celecoxib, chemical-gene interaction data derived from the Comparative Toxicogenomics database, and results from Level 1, Level 2, and Level 3 SeqAPASS analyses that examine conservation of target proteins across species (https://seqapass.epa.gov/seqapass/). This dataset is associated with the following publication: Martinovic-Weigelt, D., A. Mehinto, G. Ankley , J. Berninger, T. Collette , J. Davis , N. Denslow, E. Durhan, E. Eid, D. Ekman , K. Jensen , M. Kahl , C. LaLone , Q. Teng , and D. Villeneuve. Derivation and evaluation of putative adverse outcome pathways for the effects of cyclooxygenase inhibitors on reproductive processes in female fish. TOXICOLOGICAL SCIENCES. Society of Toxicology, 156(2): 344-361, (2017).