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To assess the potential endocrine disruptive effects through multiple nuclear receptors(NRs),especially non-steroidal NRs,in municipal wastewater,we examined the agonistic activities on four NRs(estrogen receptor α,thyroid hormone receptor α,retinoic acid receptor α and retinoid Ⅹ receptor α)of untreated and treated wastewater from municipal wastewater treatment plants(WWTPs)in Japan using a yeast two-hybrid assay.Investigation of the influent and effluent of seven WWTPs revealed that agonistic activities against steroidal and non-steroidal NRs were always detected in the influents and partially remained in the effluents.Further investigation of four WWTPs employing conventional activated sludge,pseudo-anoxic-oxic,anoxic-oxic and anaerobic-anoxic-oxic processes revealed that the ability to reduce the agonistic activity against each of the four NRs varies depending on the treatment process.These results indicated that municipal wastewater in Japan commonly contains endocrine disrupting chemicals that exert agonistic activities on steroidal and non-steroidal NRs,and that some of these chemicals are released into the natural aquatic environment.Although the results obtained in yeast assays suggested that measured levels of non-steroidal NR agonists in the effluent of WWTPs were not likely to cause any biological effect,further study is required to assess their possible risks in detail.