Rapid Test Kit for Quantifying Hormonal Activity in Animal Feeding Operation Wastewater

Passage in 1996 of the Food Quality Protection Act (FQPA) and Amendments to the Safe Drinking Water Act (SDW) reflected these concerns and required EPA to develop a screening program, using appropriate validation test systems and other scientifically relevant information, to determine whether certain substances may have an endocrine effect in wildlife and humans. The proposed work will result in the development of an assay which tests for hormonal substances which are capable of altering development and reproductive processes in aquatic animals exposed to waste run-off from animal feed operations (AFOs). This proposal incorporates recommendations of previous reviewers to: 1) expound on the predictability of the model and determine the propensity of false positive and false negative responses, 2) evaluate the potential for interactive effects in complex wastewaters from the AFOs, 3) demonstrate the quantitative nature of the model in relation to potential regulations of wastewater from AFOs, and 4) demonstrate the link between proposed Phase I and II activities. The primary goal of the proposed research is to develop and commercialize a rapid and cost-effective test kit to measure contaminants released in to surface waters and ground water from animal feeding operations. Specifically, we will develop a 24-h test model for evaluating the toxicological activity of the waste run-off from animal feeding operations by monitoring the hormonally induced process of oocyte maturation in X. laevis oocytes in vitro. Evaluation will entail testing neat chemicals with known activities to evaluate the potential for false positive and negative responses, complex wastewater mixtures wastewater samples collected from AFOs, and complex mixtures from AFOs spiked with EDCs. The complex wastewater mixture from the AFOs will be evaluated with and without the standard additions of known EDCS to evaluate the potential for additive, non-interactive responses and interactive responses including synergism and antagonism using a binary mixture model. Inhibition of GVBD will be evaluated relative to the classical inhibition ethinyl estradiol to establish a quantitivative measure of estrogen equivalents. When GVBD stimulation or inhibition is detected, complementary progesterone and androgen receptor binding assay will be performed to determine if the effect is the result of receptor binding. The successful completion of the proposed technology will provide the scientific and regulatory community with an ecologically-relevant, rapid means of evaluating the toxicological effects of pharmaceuticals and hormones contained released to the environment from animal feed operations.