Evolutionary History of the Rhinolophus pusillus Species Complex Reveals Phylogeographic Diversification Across the East Asian Mainland–Island System

Abstract

Simple Summary The Rhinolophus pusillus species complex is widely distributed across East Asia and includes several geographically isolated or taxonomically debated forms. Because morphology, echolocation calls and genetic markers have previously given partly conflicting results, its evolutionary history has remained unclear. In this study, we combined mitochondrial DNA, nuclear loci, microsatellites, morphology, acoustic data, demographic analyses, ancestral-area reconstruction and ecological niche modelling using range-wide samples from mainland China, Hainan and Taiwan. Our results show that R. pusillus contains several geographically structured mitochondrial lineages, while the Taiwan endemic R. monoceros represents a closely related but distinctive island lineage. South China appears to have been an important refugial and ancestral region, whereas Hainan, Taiwan and Southwest China experienced stronger historical isolation. Nuclear markers showed weaker differentiation, suggesting incomplete lineage sorting and historical connectivity. These findings indicate that the complex is still undergoing diversification and that several regional populations deserve conservation attention.