The Laosicheng site in Yongshun County, Hunan Province, China, plays an important role in elucidating the animal population composition of the region prior to the middle Qing Dynasty (approximately 300 years ago). This site has yielded a substantial number of animal bones, 11 960 of which have been identified to the species level, serving as direct evidence of ancient animal populations (Hunan Provincial Institute of Cultural Relics and Archaeology, 2018). Notably, two mammalian skulls from this site, designated as Laosicheng1: 10YLG10 (101) and Laosicheng2: 10YLG10 (134), have garnered particular attention, with Laosicheng2 selected as the cover image for the above publication. Initially, these skulls were classified as Canids, with Laosicheng1 identified as a dog and Laosicheng2 identified as a wolf (Hunan Provincial Institute of Cultural Relics and Archaeology, 2018). However, the accuracy of these classifications has been called into question. In this context, a comprehensive analysis was undertaken, involving precise cranial morphological examination and mitochondrial genome (mitogenome) sequencing of the two Laosicheng samples using micro-computed tomography (micro-CT) and ancient DNA (aDNA) techniques to facilitate taxonomic reassessment. The results revealed that both skulls are attributable to leopards (Panthera pardus japonensis). Furthermore, phylogenetic analysis using data from this study and global leopard datasets, with a focus on East Asia, strongly suggested that East Asian leopards (P. p. orientalis, P. p. japonensis, and P. p. delacouri) belong to three distinct populations. This insight significantly advances our understanding of the maternal genetic history of East Asian leopards.