Understanding the habitat selection and population genetic structure of an endangered species can play important roles in its protection. The Wuchuan odorous frog (Odorrana wuchuanensis) is endemic to the karst regions of southwest China. This frog is currently listed as “Critically Endangered” by the IUCN, but little is known about its habitat selection and population genetics. In this study, we conducted analyses of habitat selection with occurrence/absence sites and environmental data, and assessed the genetic structure between north and south populations in Guizhou provinces in China using three mitochondrial markers. The results revealed that the probability of this frog occupying cave habitats increased with higher average humidity in July and higher lowest temperature in January, but was negatively related to precipitation in January. Analyses of F statistics combined with analyses of median-joining haplotype networks and the phylogenetic tree showed low genetic differentiation between the two populations of O. wuchuanensis. Considering the small population size and geographic isolation because of the complex karst terrains, we suggest careful management practices are needed to protect this species.