Hipposideros pratti is species of bat distributed in caves from Southeast Asia to the Qinling-Funiu Mountain area in China. To understand the dynamic changes in its distribution and evaluate the current health status of the species, we reviewed 48 caves in the northernmost area of the H. pratti distribution and conducted the first analysis of genetic diversity for the species in the region. H. pratti was only detected in three caves, the four previously distributed caves and the remaining 41 previously undistributed caves were not observed. An analysis based on microsatellite markers revealed a few key points. 1) The average number of observed alleles (Na) of H. pratti in the region was 3.94, and the average observed heterozygosity (Ho) was 0.5293. All three populations were in Hardy–Weinberg equilibrium. 2) Intra-population was the predominant genetic variation of the H. pratti population. 3) H. pratti in the region experienced a bottleneck effect. We found that the three caves where H. pratti are currently distributed face varying degrees of human interference and the populations are threatened. Management strategies, including appropriate countermeasures to reduce human interference in caves, are urgently needed.