Knowledge of genetic structure is essential for the long-term management and conservation of endangered species. We report the results from a genetic examination of the federally endangered Florida bonneted bat (Eumops oridanus) sampled from its range in southern Florida, USA. Bonneted bats are primarily found in four regions separated by approximately 100 to 250 kms, including three western natural areas (BW, PC, and CC) and one urban population on the east coast [Miami-Dade County (MD)]. We used 22 microsatellite loci and cytochrome b sequences to assess the extent of connectivity and levels of genetic diversity. Regional populations were highly differentiated (F ST = 0.178) and model-based and multivariate analyses showed that MD was the most distinct among pairwise comparisons. Regional populations are small (i.e., N e < 100) but demographically stable. Estimates of contemporary migration and historic gene ow suggest that regional populations do not frequently exchange migrants, but simulations suggest that the divergence among western regions is likely a result of recent genetic drift rather than long-term isolation. Signi cantly, mitochondrial DNA revealed that haplotypes from MD were similar or shared with those recognized as Eumops ferox from Cuba and Jamaica, and divergent (1.5%) from the remainder of bonneted bats in Florida. Our data support the management of each of the four populations as distinct population segments, and that BW, PC and CC combined are on an independent evolutionary trajectory from bats in MD. Critically, bonneted bats in Florida appear to harbor cryptic diversity that will require a reassessment of their taxonomy. (Moussy et al. 2013). In non-migratory species, genetic structure has been explained as an outcome of the extent of gene ow resulting from either contemporary processes such as natal dispersal, mating dispersal or recent colonization, or from historical isolation (i.e., phylogeographic processes Avise 2000).Distinguishing historical processes (i.e., predating anthropogenic impacts) from more recent processes (i.e., contemporary colonization and founder events) can be di cult because the latter may be strongly in uenced by the former (Keyghobadi 2007). However, genotypic arrays of microsatellites are expected to re ect the shortest and nest-scale contemporary processes whereas mitochondrial sequence data provides a better picture of deeper historical processes (Sunnucks 2000). In geographically restricted species having no obvious historical barriers to dispersal, contemporary processes are generally predicted to predominate the genetic signature of these marker types.The Florida bonneted bat (Eumops oridanus, family Molossidae) is a federally endangered bat that is endemic to southern Florida, USA. Despite having one of the most limited geographic distributions of any bat in the United States, and recent efforts to examine occupancy across its range (Bailey et al. 2017a), its distribution within Florida is still poorly understood. Initially known only from Miami-Dade Coun...