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Macronycteris commersoni (Hipposideridae), a bat species endemic to Madagascar, is widespread across the island and utilizes a range of habitat types including open woodland, degraded habitats, and forested areas from sea level to 1,325 m. Despite being widely distributed, there is evidence that M commersoni exhibits morphological and bioacoustic variation across its geographical range. We investigated the fine-scale phylogeographic structure of populations in the western half of the island using extensive spatial sampling and sequence data from two mitochondrial DNA regions. Our results indicated several lineages within M. commersoni. Individuals collected from northern Madagascar formed a single monophyletic clade (clade C). A second clade (clade B) included individuals collected from the south-western portion of the island. This second dade displayed more phylogeographical partitioning with differences in mtDNA haplotypes frequency detected between populations collected in different bioclimatic regions. Lineage dispersal, genetic divergence, and timing of expansion events of M. commersoni were probably associated with Pleistocene climate fluctuations. Our data suggest that the northern and the central western regions of Madagascar may have acted as refugia for this species during periods of cooler and drier climate conditions associated with the Pleistocene.