To investigate whether male carp (Cyprinus carpio) are really feminized by estrogen-like substances in effluent from sewage treatment plants (STPs) in Japan, exposure tests were performed using carp in water tanks that were receiving effluent from a STP. During eight weeks of exposure, we had measured their serum vitellogenin (VTG), a precursor of yolk protein specific for females that is synthesized in their liver by estrogen-like substances and that is, therefore, a biomarker of feminization. For this research, a STP with representative effluent estrogenic activity level was selected. In the first run of the tests that launched in the early spring of 2000, male carp were exposed. Results showed that the VTG concentration in male carp exposed to the effluent increased with time, while it was reduced after exposure to the effluents was stopped and the exposure was changed to dechlorinated tap water. On the other hand, in the control experiment, VTG was not detected in male carp exposed to dechlorinated tap water. Therefore, male carp might be reversibly feminized by the STP effluent. To assure whether the feminization of male carp was caused by the STP effluent at any time, two more runs of the test were performed in the summer of 2000 and February to April of 2001. However, significant level of VTG in male carp could not be observed for eight weeks. The estrogenic activities in the effluents were almost the same level throughout the tests; therefore the seasonal timing of exposure might be important for understanding this inconsistent phenomenon.