Jonathan M. Ali

Bioavailability and Fate of Sediment-Associated Progesterone in Aquatic Systems

The environmental fate and bioavailability of progesterone, a steroid hormone known to cause endocrine-disrupting effects in aquatic organisms, is of growing concern due to its occurrence in the environment in water and sediment influenced by wastewater treatment plant and paper mill effluents, as well as livestock production. The objective of this study was to evaluate the fate of progesterone in two natural sediments and the corresponding alteration of gene expression in three steroid-responsive genes; vitellogenin, androgen receptor and estrogen receptor alpha. When exposed to progesterone-spiked sand, fathead minnows (Pimephales promelas) exhibited significant reductions in the expression of vitellogenin and androgen receptor expression. In contrast, fish exposed to progesterone associated with the silty loam sediment did not show a biological response at 7 days and only realized a significant reduction in vitellogenin. In both sediments, progesterone degradation resulted in the production of androgens including androsteinedione, testosterone, and androstadienedione, as well as the antiestrogen, testolactone. Differences in compound fate resulted in organism exposure to different suites of metabolites either in water or associated with the sediment. Results from this study suggest that environmental progestagens will lead to defeminization at environmentally relevant concentrations, and that exposure is influenced by sediment properties.

Impact of Sediment on Agrichemical Fate and Bioavailability to Adult Female Fathead Minnows: A Field Study

Precipitation induced runoff is an important pathway for agrichemicals to enter surface water systems and expose aquatic organisms to endocrine-disrupting compounds such as pesticides and steroid hormones. The objectives of this study were to investigate the distribution of agrichemicals between dissolved and sediment-bound phases during spring pulses of agrichemicals and to evaluate the role of suspended sediment in agrichemical bioavailability to aquatic organisms. To accomplish these objectives, suspended sediment and water samples were collected every 3 days from a field site along the Elkhorn River, located at the downstream end of a heavily agricultural watershed, and were screened for 21 pesticides and 21 steroids. Adult female fathead minnows (Pimephales promelas) were exposed in field mesocosms to river water containing varying sediment loads. Changes in organism hepatic gene expression of two estrogen-responsive genes, vitellogenin (VTG) and estrogen receptor alpha (ERα), as well as the androgen receptor (AR) were analyzed during periods of both low and high river discharge. Trends in agrichemical concentrations of both the dissolved and sediment phases as a function of time show that, while sediment may act as both a source and a sink for agrichemicals following precipitation events, the overall driver for molecular defeminization in this system is direct exposure to the sediment-associated compounds. This study suggests that endocrine disrupting effects observed in organisms in turbid water could be attributed to direct exposure of contaminated sediment.

On‐site, serial exposure of female fathead minnows to the Elkhorn River, Nebraska, USA, spring agrichemical pulse

In the Midwestern United States, waterways such as the Elkhorn River experience an annual spring pulse of runoff that carries sediments, nutrients, and organic compounds downstream. The objective of the present study was to elucidate relationships between contaminant load in Elkhorn River water and biological impacts on female fathead minnows throughout the entire spring agrichemical pulse. Fish were maintained in on-site outdoor microcosms at the Elkhorn River Research Station. The start of the spring pulse was determined using commercially available atrazine strips that detected atrazine when concentrations exceeded 3 ppb. Once the pulse began, 5 serial 7-d exposures were conducted. Concentrations of atrazine, its metabolites, and 5 other herbicides were quantified using gas chromatography-mass spectrometry. Agrichemicals peaked during the first and second weeks of the pulse, with a smaller peak occurring during week 4, but the peaks were not directly associated with runoff events (as estimated from river discharge). Elevated agrichemical concentrations were associated with biological impacts, but not solely responsible. In the present study, differences in the abiotic environment were found to play a significant role in the defeminization of exposed female fathead minnows.

Response and recovery of fathead minnows (Pimephales promelas) following early life exposure to water and sediment found within agricultural runoff from the Elkhorn River, Nebraska, USA

Agricultural runoff is a non-point source of chemical contaminants that are seasonally detected in surface water and sediments. Agrichemicals found within seasonal runoff can elicit endocrine disrupting effects in organisms as adults, juveniles and larvae. The objectives of this study were (1) to determine if exposure to water, sediment or the water-sediment combination collected from an agricultural runoff event was responsible for changes in endocrine-responsive gene expression and development in fathead minnow larvae, and (2) whether such early life exposure leads to adverse effects as adults. Larvae were exposed during the first month post-hatch to water and sediment collected from the Elkhorn River and then allowed to depurate in filtered water until reaching sexual maturity, exemplifying a best-case recovery scenario. Gas chromatography mass spectrometry (GC/MS) analysis of the water and sediment samples detected 12 pesticides including atrazine, acetochlor, metolachlor and dimethenamid. In minnow larvae, exposure to river water upregulated androgen receptor gene expression whereas exposure to the sediment downregulated estrogen receptor α expression. Adult males previously exposed to both water and sediment were feminized through the induction of an ovipositor structure whereas no impacts were observed in other reproductive or sex characteristic endpoints for either sex based on exposure history. Results from this study indicate that both water and sediments found in agricultural runoff elicit responses from minnow larvae, and larvae can recover following early life exposure under a best-case scenario.

Biological Impacts in Fathead Minnow Larvae Following a 7-Day Exposure to Agricultural Runoff: A Microcosm Study

The objective of this study was to investigate the impact of agricultural runoff on growth and development in fathead minnow larvae. Identifying these impacts in the field is difficult due to the complexity of pulsatile events and the challenge of maintaining larval fish under environmental conditions. This paper presents evidence of sublethal impacts on minnow larvae, maintained in microcosms, following a 7-day exposure to agricultural runoff. Fathead minnow larvae (5–12 dph) were exposed to agricultural runoff in the Elkhorn River, NE, USA. At 28 dph, 16 days after the field exposure, river exposed larvae showed reductions in body mass and length compared to controls. Female larvae exposed to river water also showed a reduction in the expression of cyp19a compared to controls. Further research will be necessary to separate the impact of agrichemicals from that of other stressors, such as suspended sediment and daily oscillation in water temperature.

Compensatory response of fathead minnow larvae following a pulsed in-situ exposure to a seasonal agricultural runoff event

Agriculturally-dominated waterways such as those found throughout the Midwestern United States often experience seasonal pulses of agrichemical contaminants which pose a potential hazard to aquatic organisms at varying life stages. The objective of this study was to characterize the developmental plasticity of fathead minnow larvae in a natural environment subject to a seasonal episodic perturbation in the form of a complex mixture of agricultural stressors. Fathead minnow larvae were maintained at the Elkhorn River Research Station for a 28-d in situ exposure to an agrichemical pulse event. Minnow larvae were sampled after 14 and 28days to characterize developmental plasticity through growth measures and relative gene expression. Concentrations of agrichemical contaminants measured in water using polar organic chemical integrative samplers and composite sediment samples throughout the 28-d exposure were quantified using gas chromatography-mass spectrometry. Elevated concentrations of acetochlor, atrazine, and metolachlor were indicative of inputs from agricultural sources and were associated with reductions in body mass, condition factor, and androgenic gene expression in river exposed fathead minnow larvae. However, following a 14-d in situ depuration during the post-pulse period, river exposed larvae overcompensated in previously suppressed biological endpoints. These results indicate that fathead minnow larvae are capable of compensatory responses following episodic exposure to agrichemical stressors.

De novo Assembly and Analysis of the Chilean Pencil Catfish Trichomycterus areolatus Transcriptome

Trichomycterus areolatus is an endemic species of pencil catfish that inhabits the riffles and rapids of many freshwater ecosystems of Chile. Despite its unique adaptation to Chiles high gradient watersheds and therefore potential application in the investigation of ecosystem integrity and environmental contamination, relatively little is known regarding the molecular biology of this environmental sentinel. Here, we detail the assembly of the Trichomycterus areolatus transcriptome, a molecular resource for the study of this organism and its molecular response to the environment. RNA-Seq reads were obtained by next-generation sequencing with an Illumina® platform and processed using PRINSEQ. The transcriptome assembly was performed using TRINITY assembler. Transcriptome validation was performed by functional characterization with KOG, KEGG, and GO analyses. Additionally, differential expression analysis highlights sex-specific expression patterns, and a list of endocrine and oxidative stress related transcripts are included.

Comparing the effects of atrazine and an environmentally relevant mixture on estrogen-responsive gene expression in the northern leopard frog and the fathead minnow

In Nebraska, fish are exposed to herbicides in agricultural runoff. The study objectives were to determine 1) if fathead minnows and northern leopard frogs exposed to atrazine experience alterations in gene expression, and 2) whether these changes are elicited by a simulated herbicide mixture. Following a 7-d exposure to atrazine, female minnows were defeminized, whereas male frogs were feminized. The mixture did not elicit statistically significant effects in either species. Environ Toxicol Chem 2018;37:1182-1188.

Estrogenic effects following larval exposure to the putative anti-estrogen, fulvestrant, in the fathead minnow (Pimephales promelas)

The objective of the present study was to investigate the consequences of early-life exposure to fulvestrant on estrogenic gene expression in fathead minnow larvae. To address this objective, fathead minnow larvae were exposed to fulvestrant (ICI 182,780) during the window of sexual differentiation between 0 to 30 days post-hatch (dph). The four treatment groups in this study included: filtered water controls (never exposed), solvent controls (ethanol 0.01%), and nominally low (0.10μg/L) and high (10.0μg/L) doses of fulvestrant. Following 30 d exposure to their respective treatment, larvae were transferred to filtered water aquaria and assessed for alterations in endocrine-responsive gene expression (i.e., RT-qPCR), body size and survival. The remaining fish depurated in filtered water until reaching sexual maturity (180dph) for assessment of persistent effects on sex characteristics, reproductive performance and sex ratio. Following the 30-d early life exposure, larvae showed upregulations of the endocrine-responsive genes ar, erβ and vtg in response to both low and high doses of fulvestrant, but showed no differences in survival or body mass. Upon reaching sexual maturity under depuration conditions, male minnows previously exposed to fulvestrant as larvae showed reductions in gonad mass along with the feminization of secondary sex characteristics with no observed effects in females. Exposure to fulvestrant had no effects on gonadal histology, reproductive performance or final sex ratio as adults. Results from this study demonstrate that aqueous exposure to fulvestrant is estrogenic in fathead minnow larvae and is capable of feminizing male fish as adults following early life exposure.