The taxonomic status of subspecies of Hipposideros pomona (Mammalia: Chiroptera: Hipposideridae) in China is controversial and uncertain. To explore the differences and subspecies differentiation in H. pomona from different regions of China, we conducted a series of bat surveys from 2011 to 2014. We performed multivariate morphometric analyses using 16 external and 21 skull measurements based on 100 samples and compared DNA sequence divergence. The results revealed distinct differences in both the external and skull measurements of H. pomona. The Cytb and COΙ; sequence divergences of H. pomona from 3 regions ranged from 6.0% to 8.5% and 5.2% to 8.0%, respectively, which reached the levels required for subspecies classification. In contrast, the RAG1 gene was not considered to be a good molecular marker for identifying subspecies because the sequence divergence was 0–0.7% within a region and 0–0.8% among regions. Both the morphological and molecular results of this study supported the conclusion that H. pomona from the Min-Guang coastal (SM), South Yunnan (SY), and Hainan Island (SH) subregions in South China belonged to different subspecies. The SM population represents H. p. sinesis, and the SH population may be a unique subspecies. Whether the SY population belongs to H. p. gentilis requires further study.