A population of western mosquitofish (Gambusia affinis) living below a wastewater treatment plant in the vicinity of Birmingham (AL, USA) was studied for evidence of exposure to estrogens. Mosquitofish are sexually dimorphic live-bearing fish. Males have an elongated and modified anal fin, called a gonopodium, used in mating. It has been hypothesized that exposure to estrogens and/or anti-androgens in treated wastewater might inhibit the androgen-dependent development of the gonopodium. The population in this study showed no evidence of having been exposed to endocrine-disrupting chemicals. The difference in adjusted (for overall fish size via analysis of covariance) mean gonopodium length between effluent-exposed and control populations was not significant. No detectable levels of vitellogenin were observed in the blood of any of the male mosquitofish, either from the effluent-exposed or the control population. Testes and livers were weighed and examined histologically. The fish exposed to treated wastewater effluent had significantly larger adjusted mean weights for both testis and liver, but no histological changes indicating exposure to estrogens were observed.