Natural infection of sentinel red foxes (Vulpes fulva) and free-ranging red foxes, gray foxes (Urocyon cinereoargenteus) and raccoons (Procyon lotor) with La Crosse (LAC) virus was demonstrated. One isolate of LAC virus was obtained from a sentinel red fox in an enzootic area. The viremia titer of the LAC virus-infected red fox was above the threshold of infection for Aedes triseriatus mosquitoes. Antibody responses were measured by the microneutralization test employing four California group viruses: LAC, snowshoe hare, trivittatus, and Jamestown Canyon. Four of six sentinel red foxes developed LAC virus neutralizing antibody. Antibody titers peaked between days 7 and 34 and were still detectable by day 345 post-infection. Homologous LAC virus antibody titers were consistently 4-fold or greater than heterologous titers. Serological evidence for natural LAC virus infection was demonstrated in 33 of 57 (58%) free-ranging red foxes, 18 of 32 (57%) gray foxes, and 4 of 16 (25%) raccoons. Antibody titers were comparable in free-ranging foxes infected naturally and those infected experimentally by mosquito bite. The prevalence of infection was significantly different (P < 0.001) for foxes trapped in three distinct areas within the enzootic region. Rates of infection in foxes by area coincided with LAC virus antibody prevalence observed in free-ranging chipmunks and with reported cases of human LAC encephalitis in Wisconsin.