The endocrine disruptor 17ß-trenbolone alters the relationship between pre- and post-copulatory sexual traits in male mosquitofish (Gambusia holbrooki).

It is now well-established that reproduction in wildlife can be disrupted by anthropogenic environmental changes, such as chemical pollution. However, very little is known about how these pollutants might affect the interplay between pre- and post-copulatory mechanisms of sexual selection. Here, we investigated the impacts of 21-day exposure of male eastern mosquitofish (Gambusia holbrooki) to a field-realistic level (average measured concentration: 11 ng/L) of the endocrine-disrupting chemical 17ß-trenbolone (17ß-TB) on pre- and post-copulatory reproductive traits. We examined male reproductive behaviour by testing the time spent near a female behind a partition, as well as the number of copulation attempts made, and the time spent chasing a female in a free-swimming context. Sperm traits were also assayed for all males. We found that exposure of male fish to 17ß-TB altered the relationship between key pre- and post-copulatory reproductive traits. Furthermore, 17ß-TB-exposed males had, on average, a higher percentage of motile sperm, and performed fewer copulation attempts than unexposed males. However, there was no overall effect of 17ß-TB exposure on either the time males spent associating with or chasing females. Taken together, our findings demonstrate the potential for chemical pollutants to affect both pre- and post-copulatory sexual traits, and the interplay between these mechanisms of sexual selection in contaminated wildlife.

Context-specific behavioural changes induced by exposure to an androgenic endocrine disruptor.

Pharmaceutical contaminants are being detected with increased frequency in organisms and ecosystems worldwide. This represents a major environmental concern given that various pharmaceuticals act on drug targets that are evolutionarily conserved across diverse taxa, are often persistent in the environment, and can bioconcentrate in organisms and bioaccumulate in food chains. Despite this, relatively little is known about the potential for pharmaceutical contaminants to affect animal behaviour, especially across multiple fitness-related contexts. Here, we investigated impacts of 21-day exposure of wild-caught male eastern mosquitofish (Gambusia holbrooki) to a field-realistic level of the veterinary pharmaceutical 17ß-trenbolone-a growth-promoting steroid used extensively in beef production worldwide and a potent androgenic endocrine disruptor repeatedly detected in surface waters affected by livestock effluent run-off. First, we examined male boldness, activity, and exploratory behaviour in a novel environment (maze arena) and found no significant effect of 17ß-trenbolone exposure. Second, the same males were tested in a reproductive assay for their tendency to associate with a stimulus (unexposed) female behind a partition. Exposed males exhibited reduced association behaviour, taking longer to first associate with, and spending less time within close proximity to, a female. Third, all males were assayed for sperm function (computer-assisted sperm analysis, sperm viability) or quantity (total sperm count) and, although no significant main effects of 17ß-trenbolone were seen on sperm traits, exposure altered the relationship between male morphology and sperm function. Lastly, morphological traits were assessed and exposed males were found to have, on average, increased mass relative to length. In combination, these results demonstrate that exposure to a field-realistic level of 17ß-trenbolone can produce subtle but important trait alterations in male fish-including context-specific behavioural changes, disruption of key sperm function trade-offs, and altered morphology-with potential impacts on exposed wildlife.

Impact of the widespread pharmaceutical pollutant fluoxetine on behaviour and sperm traits in a freshwater fish.

Pharmaceutical pollutants are detected in aquatic habitats and wildlife tissues globally. One widespread contaminant of major concern is the antidepressant fluoxetine, which can affect behavioural and physiological processes in non-target species. Despite this, effects of fluoxetine on wildlife behaviour have seldom been investigated across multiple fitness-related contexts, especially at environmentally realistic concentrations. Accordingly, we examined impacts of 35-day fluoxetine exposure at two environmentally relevant concentrations (31 and 374 ng/L) across a suite of fitness-related contexts in wild-caught male mosquitofish (Gambusia holbrooki). First, we investigated anxiety-related behaviours (boldness, exploration and activity) in a novel environment (maze arena) and found no significant impacts of exposure. Second, we tested effects of fluoxetine in a reproductive context, including mating behaviour and sperm quality. We found that, relative to controls, fluoxetine exposure resulted in males spending a greater amount of time pursuing females. Further, low-exposed males were more likely to attempt copulation than unexposed males. Lastly, we investigated across-context behavioural correlations, and how fluoxetine exposure might affect such relationships. A significant positive correlation was detected in control fish between activity levels in the maze and time spent pursuing females in the reproductive assay. This relationship was disrupted by fluoxetine at both exposure levels. This is the first evidence that field-detected concentrations of a pharmaceutical pollutant can disturb across-context behavioural correlations in wildlife. Our findings provide clear evidence that fluoxetine can produce context-specific behavioural effects in fish and underscore how pharmaceutical exposure at field-detected concentrations can induce important shifts in wildlife behaviour.

Field-realistic exposure to the androgenic endocrine disruptor 17ß-trenbolone alters ecologically important behaviours in female fish across multiple contexts.

The capacity of pharmaceutical pollution to alter behaviour in wildlife is of increasing environmental concern. A major pathway of these pollutants into the environment is the treatment of livestock with hormonal growth promotants (HGPs), which are highly potent veterinary pharmaceuticals that enter aquatic ecosystems via effluent runoff. Hormonal growth promotants are designed to exert biological effects at low doses, can act on physiological pathways that are evolutionarily conserved across taxa, and have been detected in ecosystems worldwide. However, despite being shown to alter key fitness-related processes (e.g., development, reproduction) in various non-target species, relatively little is known about the potential for HGPs to alter ecologically important behaviours, especially across multiple contexts. Here, we investigated the effects of exposure to a field-realistic level of the androgenic HGP metabolite 17ß-trenbolone-an endocrine-disrupting chemical that has repeatedly been detected in freshwater systems-on a suite of ecologically important behaviours in wild-caught female eastern mosquitofish (Gambusia holbrooki). First, we found that 17ß-trenbolone-exposed fish were more active and exploratory in a novel environment (i.e., maze arena), while boldness (i.e., refuge use) was not significantly affected. Second, when tested for sociability, exposed fish spent less time in close proximity to a shoal of stimulus (i.e., unexposed) conspecific females and were, again, found to be more active. Third, when assayed for foraging behaviour, exposed fish were faster to reach a foraging zone containing prey items (chironomid larvae), quicker to commence feeding, spent more time foraging, and consumed a greater number of prey items, although the effect of exposure on certain foraging behaviours was dependent on fish size. Taken together, these findings highlight the potential for exposure to sub-lethal levels of veterinary pharmaceuticals to alter sensitive behavioural processes in wildlife across multiple contexts, with potential ecological and evolutionary implications for exposed populations.

The antidepressant fluoxetine alters mechanisms of pre- and post-copulatory sexual selection in the eastern mosquitofish (Gambusia holbrooki).

Contamination of aquatic habitats with pharmaceuticals is a major environmental concern. Recent studies have detected pharmaceutical pollutants in a wide array of ecosystems and organisms, with many of these contaminants being highly resistant to biodegradation and capable of eliciting sub-lethal effects in non-target species. One such pollutant is fluoxetine, a widely prescribed antidepressant, which is frequently detected in surface waters globally and can alter physiology and behaviour in aquatic organisms. Despite this, relatively little is known about the potential for fluoxetine to disrupt mechanisms of sexual selection. Here, we investigate the impacts of 30-day exposure to two environmentally realistic levels of fluoxetine (low and high) on mechanisms of pre- and post-copulatory sexual selection in the eastern mosquitofish (Gambusia holbrooki). We tested 1) male mating behaviour in the absence or presence of a competitor male, and 2) sperm quality and quantity. We found that high-fluoxetine exposure increased male copulatory behaviour in the absence of a competitor, while no effect was detected under male-male competition. Further, fluoxetine exposure at both concentrations increased total sperm count relative to males from the control group, while no significant change in sperm quality was observed. Lastly, low-fluoxetine males showed a significant reduction in condition index (mass relative to length). Our study is the first to show altered mechanisms of both pre- and post-copulatory sexual selection in an aquatic species resulting from environmentally realistic fluoxetine exposure, highlighting the capacity of pharmaceutical pollution to interfere with sensitive reproductive processes in wildlife.