Place-based screening of mixtures of dominant emerging contaminants measured in Lake Michigan using zebrafish embryo gene expression assay

Determining impacts of emerging contaminants is difficult due to the different concentrations of mixtures of these chemicals over a landscape. Assessment approaches need to account for absorption, distribution, metabolism and excretion of the chemicals in an organism, and potential crosstalk between molecular pathways. The goal of this study was to assess the utility of employing a modified zebrafish embryo toxicity (ZFET) assay that assesses morphological alterations and measurements of estrogen-associated mRNA transcripts, to exposure of a mixtures of chemicals at concentrations measured in several locations in Lake Michigan. The 5 pharmaceuticals in this study were carbamazepine, diltiazem, fluoxetine, gemfibrozil and metformin. Exposures consisted of 4 concentrations of each individual chemical, mixture concentrations measured at seven locations in Lake Michigan, or 17β-estradiol. The relative expression of Estrogen Receptor-alpha, brain aromatase (CYP19A2), and gonadotropin releasing hormone 3 mRNA were measured at the end the 6-d exposure to determine estrogenicity of the individual chemical or mixture. In this study, there was significant induction of CYP19A2 in individual exposures of diltiazem, fluoxetine, gemfibrozil and metformin at concentrations measured in Lake Michigan. Exposure to 5 of the 7 chemical mixtures altered the expression of one of the three biomarkers. Transcripts varied across mixtures, indicating that biological screening of whole water samples for potential estrogenicity may need to include alternative molecular pathways other than just steroid receptor binding. This research demonstrates that pairing chemical measurements with a modified ZFET assay, twhich incorporates molecular biomarkers and morphological endpoints, could provide location and mixture specific toxic profiling.