This study describes the diversity of ectoparasitic bat flies (Diptera: Streblidae) in the state of Yucatan, Mexico. Fieldwork was carried out from June 2010 to January 2012 in seven municipalities of Yucatan, where 13 sampling sites were selected to capture bats using mist nets. Over 156 sampling nights a total of 910 bats were captured; these belonged to 19 species in four families: Mormoopidae, Phyllostomidae, Natalidae, and Vespertilionidae. Phyllostomidae was the richest family (13 bat species), followed by Mormoopidae (3 spp.), Vespertilionidae (2 spp.), and Natalidae (1 spp.). After careful inspection of the bats, a total of 2,134 Streblid bat flies were collected, belonging to 17 species in six genera (Nycterophilia coxata Ferris, N. natali Wenzel, Trichobius diphyllae Wenzel, T. dugesii Townsend, T. galei Wenzel, T. hirsutulus Bequaert, T. intermedius Peterson and Hurka, T. parasiticus Gervais, T. uniformis Curran, T. yunkeri Wenzel, Megistopoda aranea Coquillett, M. proxima Séguy, Aspidoptera delatorrei Wenzel, Strebla alvarezi Wenzel, S. diphyllae Wenzel, S. wiedemanni Kolenati, and Metelasmus pseudopterus Coquillett). The richest and most diverse genus was Trichobius. Five species—N. natali, T. diphyllae, M. proxima, A. delatorrei, and M. pseudopterus, are new records for Yucatan, and T. galei is a new record for the country, increasing the total number of Streblidae species for Mexico to 49.