Emerging wastewater contaminant metformin causes intersex and reduced fecundity in fish

AbstractThe occurrence of intersex fish, where male reproductive tissues show evidence of feminization, have been found in freshwater systems around the world, indicating the potential for significant endocrine disruption across species in the ecosystem. Estrogens from birth control medications in wastewater treatment plant effluent have been cited as the likely cause, but research has shown that endocrine disruption is not solely predictable based on hormone receptor interactions. Many other non-hormone pharmaceuticals are found in effluent at concentrations orders of magnitude higher than estrogens, yet there is little data indicating the impacts of these other medications. The widely prescribed anti-diabetic metformin is among the most abundant of pharmaceuticals found in effluent and is structurally dissimilar from hormones. However, we show here that exposing fathead minnows (Pimephales promelas) to a concentration of metformin found in wastewater effluent causes the development of intersex gonads in males, reduced size of treated male fish, and reduction in fecundity for treated pairs. Our results demonstrate that metformin acts as an endocrine disruptor at environmentally relevant concentrations

Transcriptome signatures of wastewater effluent exposure in larval zebrafish vary with seasonal mixture composition in an effluent-dominated stream

Wastewater treatment plant (WWTP) effluent-dominated streams provide critical habitat for aquatic and terrestrial organisms but also continually expose them to complex mixtures of pharmaceuticals that can potentially impair growth, behavior, and reproduction. Currently, few biomarkers are available that relate to pharmaceutical-specific mechanisms of action. In the experiment reported in this paper, zebrafish (Danio rerio) embryos at two developmental stages were exposed to water samples from three sampling sites (0.1 km upstream of the outfall, at the effluent outfall, and 0.1 km below the outfall) during base-flow conditions from two months (January and May) of a temperate-region effluent-dominated stream containing a complex mixture of pharmaceuticals and other contaminants of emerging concern. RNA-sequencing identified potential biological impacts and biomarkers of WWTP effluent exposure that extend past traditional markers of endocrine disruption. Transcriptomics revealed changes to a wide range of biological functions and pathways including cardiac, neurological, visual, metabolic, and signaling pathways. These transcriptomic changes varied by developmental stage and displayed sensitivity to variable chemical composition and concentration of effluent, thus indicating a need for stage-specific biomarkers. Some transcripts are known to be associated with genes related to pharmaceuticals that were present in the collected samples. Although traditional biomarkers of endocrine disruption were not enriched in either month, a high estrogenicity signal was detected upstream in May and implicates the presence of unidentified chemical inputs not captured by the targeted chemical analysis. This work reveals associations between bioeffects of exposure, stage of development, and the composition of chemical mixtures in effluent-dominated surface water. The work underscores the importance of measuring effects beyond the endocrine system when assessing the impact of bioactive chemicals in WWTP effluent and identifies a need for non-targeted chemical analysis when bioeffects are not explained by the targeted analysis

Gene set enrichment analysis of microarray data from Pimephales promelas (Rafinesque), a non-mammalian model organism

<p>Abstract</p> <p>Background</p> <p>Methods for gene-class testing, such as Gene Set Enrichment Analysis (GSEA), incorporate biological knowledge into the analysis and interpretation of microarray data by comparing gene expression patterns to pathways, systems and emergent phenotypes. However, to use GSEA to its full capability with non-mammalian model organisms, a microarray platform must be annotated with human gene symbols. Doing so enables the ability to relate a model organism's gene expression, in response to a given treatment, to potential human health consequences of that treatment. We enhanced the annotation of a microarray platform from a non-mammalian model organism, and then used the GSEA approach in a reanalysis of a study examining the biological significance of acute and chronic methylmercury exposure on liver tissue of fathead minnow (<it>Pimephales promelas</it>). Using GSEA, we tested the hypothesis that fathead livers, in response to methylmercury exposure, would exhibit gene expression patterns similar to diseased human livers.</p> <p>Results</p> <p>We describe an enhanced annotation of the fathead minnow microarray platform with human gene symbols. This resource is now compatible with the GSEA approach for gene-class testing. We confirmed that GSEA, using this enhanced microarray platform, is able to recover results consistent with a previous analysis of fathead minnow exposure to methylmercury using standard analytical approaches. Using GSEA to compare fathead gene expression profiles to human phenotypes, we also found that fathead methylmercury-treated livers exhibited expression profiles that are homologous to human systems & pathways and results in damage that is similar to those of human liver damage associated with hepatocellular carcinoma and hepatitis B.</p> <p>Conclusions</p> <p>This study describes a powerful resource for enabling the use of non-mammalian model organisms in the study of human health significance. Results of microarray gene expression studies involving fathead minnow, typically used for aquatic ecological toxicology studies, can now be used to generate hypotheses regarding consequences of contaminants and other stressors on humans. The same approach can be used with other model organisms with microarray platforms annotated in a similar manner.</p

Psychoactive Pharmaceuticals Induce Fish Gene Expression Profiles Associated with Human Idiopathic Autism

Idiopathic autism, caused by genetic susceptibility interacting with unknown environmental triggers, has increased dramatically in the past 25 years. Identifying environmental triggers has been difficult due to poorly understood pathophysiology and subjective definitions of autism. The use of antidepressants by pregnant women has been associated with autism. These and other unmetabolized psychoactive pharmaceuticals (UPPs) have also been found in drinking water from surface sources, providing another possible exposure route and raising questions about human health consequences. Here, we examined gene expression patterns of fathead minnows treated with a mixture of three psychoactive pharmaceuticals (fluoxetine, venlafaxine & carbamazepine) in dosages intended to be similar to the highest observed conservative estimates of environmental concentrations. We conducted microarray experiments examining brain tissue of fish exposed to individual pharmaceuticals and a mixture of all three. We used gene-class analysis to test for enrichment of gene sets involved with ten human neurological disorders. Only sets associated with idiopathic autism were unambiguously enriched. We found that UPPs induce autism-like gene expression patterns in fish. Our findings suggest a new potential trigger for idiopathic autism in genetically susceptible individuals involving an overlooked source of environmental contamination

Sets associated with human neurological disorders.

<p>Sets associated with human neurological disorders.</p

Single drug treatments & ASD sets.

<p>Single drug treatments & ASD sets.</p

Single drug treatments & ND sets.

<p>Single drug treatments & ND sets.</p

Observed values of psychoactive pharmaceuticals in various systems.

<p>Observed values of psychoactive pharmaceuticals in various systems.</p

“ND”: Sets of genes associated with various human neurological disorders.

<p>“ND”: Sets of genes associated with various human neurological disorders.</p