The knowledge that exposure to estrogenic wastewater treatment work (WwTW) effluents induces a range of reproductive abnormalities in fish has highlighted the need to understand the wider health effects of effluents. Access to laboratory-based testing systems for WwTW effluents could greatly facilitate this endeavour. In this investigation, a laboratory-based test system was developed and applied for WwTW effluents using fathead minnows (Pimephales promelas). Sexually maturing fathead minnows were exposed, under flow-through conditions in the laboratory, for up to 21 days to graded concentrations of effluent from three different UK (temperate) WwTWs. The stability of the estrogenic component within the test system was assessed via measurements for estradiol and estrone concentrations in the effluent, and through determining estrogenic responses in an in vitro recombinant yeast estrogen screen (rYES) and in fish (plasma vitellogenin induction). The estrogen component of the effluents was stable within the holding system used (chilled <10 degrees C) for up to 7 days and measured concentrations of estradiol and estrone were shown to differ by less than 20% between the first and final day of use for each batch of effluent. Total estrogenic activity as measured in the rYES was found to be more variable (up to 66% variance between measurements for the two time points) but there was no consistent trend for a reduction in estrogenic activity. Vitellogenin was induced in males in a concentration-dependent manner and the magnitude of the response observed was proportional to the average measured concentrations of estradiol and estrone in the exposure effluent. The system described, thus, provides a robust test method for evaluating the estrogenic effects of temperate WwTW effluents that could be further applied to assess wider health effects, including population-relevant endpoints such as reproduction, using model OECD warm-water fish species such as the fathead minnow.
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