The purpose of this study was to refine our knowledge of the natural host relationships of Whitewater Arroyo (WWA) virus. Two hundred eight rodents, representing nine species, were captured in July 1999 on the Chaparral Wildlife Management Area in southern Texas and tested for evidence of arenavirus infection. Antibody to an arenavirus was found in seven (21.9%) of 32 southern plains woodrats (Neotoma micropus) and none of 168 other rodents. Infectious WWA virus was isolated from four antibody-positive southern plains woodrats, one of 25 antibody-negative southern plains woodrats, and none of 176 other rodents. Collectively, the results indicate that the southern plains woodrat is a principal host of WWA virus in southern Texas. Analyses of viral gene sequence data revealed substantial genetic diversity among WWA virus strains isolated from the woodrats, suggesting that multiple variants of the virus can coexist in a single woodrat species in a small geographic area.