In this study, the sera collected from a variety of mammalian species (ass-mules, cat, cattle, dog, horse, human and sheep) in 10 representative provinces of Turkey, were surveyed for the presence of neutralizing antibodies to West Nile virus (WNV). Overall, 1 of 40 (2.5%) ass-mules, 4 of 100 (4%) cattle, 43 of 114 (37.7%) dogs, 35 of 259 (13.5%) horses, 18 of 88 (20.4%) humans and 1 of 100 (1%) sheep, tested positive for WNV-neutralizing antibodies. The results indicate that a wide range of mammals are exposed to a West Nile-related virus and this could contribute to the long-term survival of this virus in the absence of overt disease.