The Bonin flying fox (Pteropus pselaphon) is endemic to the Ogasawara Islands, a collection of small oceanic islands in the Pacific Ocean. It inhabits only five islands: Chichi-jima, Haha-jima, Kita-iwo, Iwo, and Minami-iwo (arranged from north to south). Hahajima and Kita-iwo, the most widely spaced islands, are separated by a distance of about 160 km. The islands have different histories in the modern era with respect to human activity. At present, P. pselaphon population sizes exceed 100 on Chichi-jima and Minamiiwo, but the species is rare on the other three islands. Loss of genetic diversity is of concern because of the small population sizes. We obtained samples from three of the five islands — Chichi-jima, Kita-iwo, and Minami-iwo — and investigated species genetic diversity and genetic structure based on mitochondrial DNA (mtDNA) control region sequences and microsatellite markers. Different mtDNA haplotypes were found in each island population. Based on the mtDNA sequence data, P. pselaphon displayed a cryptic genealogy, as haplotypes on each island did not cluster together. The microsatellite marker data, however, revealed a clear genetic structure among the island populations, suggesting the absence of recent inter-island gene flow. Based on these results, we propose that the individual island populations are not evolutionarily significant units, but should be conserved collectively as a single management unit.