The analysis of molecular data within a historical biogeographical framework, coupled with ecological characteristics can provide insight into the processes driving diversification. Here we assess the genetic and ecological diversity within a widespread horseshoe bat Rhinolophus clivosus sensu lato with specific emphasis on the southern African representatives which, although not currently recognized, were previously described as a separate species R. geoffroyi comprising four subspecies. Sequence divergence estimates of the mtDNA control region show that the southern African representatives of R. clivosus s.l. are as distinct from samples further north in Africa than they are from R. ferrumequinum, the sister-species to R. clivosus. Within South Africa, five genetically supported geographic groups exist and these groups are corroborated by echolocation and wing morphology data. The groups loosely correspond to the distributions of the previously defined subspecies and Maxent modelling shows a strong correlation between the detected groups and ecoregions. Based on molecular clock calibrations, it is evident that climatic cycling and related vegetation changes during the Quaternary may have facilitated diversification both genetically and ecologically.