Estrogens play a major role in the sexual differentiation, gonad development, and oocyte growth of most oviparous organisms. They also stimulate vitellogenesis, the formation of high-density glycolipophosphoprotein that serves as an energy source for the developing embryo. Surface waters from the St. Lawrence River, obtained in the vicinity of an urban area (Montreal, Quebec, Canada), were studied with respect to their estrogenic potential to the freshwater mussel Elliptio complanata. Estrogenicity was measured in water extracts by means of a competitive assay of estradiol binding to cytosolic proteins and by the vitellin-inducing ability of mussel hemolymph following direct extract injection. Surface-water samples drawn downstream of a municipal outfall plume and in a river draining a large farming and agricultural area had high levels of total and fecal coliform bacteria. High levels of estrogen competitors were also found and were able to induce vitellins in injected mussels. Moreover, the estrogen-competing potential of the extracts was found to be significantly correlated with total and fecal coliform bacteria (R = 0.9, p < 0.01) and with the levels of vitellins in the hemolymph (R = 0.62, p = 0.03). The results indicate that water samples drawn from within the municipal effluent plume and from a river draining an agricultural area are estrogenic to freshwater mussels. Thus, the environmental inputs of estrogens are likely to be associated with human sewage and pesticide products.