In order to clarify chiropteran resources in Hunan Province, a series of field surveys were conduct in July and September 2022. During our surveys, nine Myotis bats (7 males and 2 females) and five Murina bats (2 males and 3 females) were collected using harp traps in Taoyuan County of Changde City, Baojing County of Xiangxi Prefecture, Shuangpai County of Yongzhou City and Suiling County of Shaoyang City. To identify our sampling, morphological taxonomic analysis and phylogenetic inference were conducted. The Myotis specimens are medium-sized, with forearm length of 43.41﹣50.89 mm (Table 1) and reddish-brown pelage, and blackend edges of ears, nose, toes, claws, and the tail are black-brown (Fig. 1a, c). Triangular reddish-brown spots were presented between the palmar fingers of the wing membrane reaching the edge. The dorsal view of the skull was flat with well-developed zygomatic arch, and the cranial is high and prominent (Fig. 1e). These morphological characteristics and measurements accord with those of Myotis rufoniger. Meanwhile, the Murina specimens were medium-sized, with forearm length of 33.83﹣37.53 mm (Table 1), sepia dorsal hair and light yellow ventral hair (Fig. 2b, c). The greatest length of skull is 16.74﹣17.29 mm (Table 1), and the zygomatic is straight and broad (Fig. 2d, e). These features are consistent with those of Murina huttoni. Based on Cyt b and COI genes segments, our phylogenetic inferences also verified such identifications (Fig. 3). These findings represent the first records of M. rufoniger and Mu. huttoni in Hunan province. Presently, the specimens are deposited in Key Laboratory of Conservation and Application in Biodiversity of South China, Guangzhou University which the M. rufoniger specimens are numbered GZHU 22429, GZHU 22430, GZHU 22459, GZHU 22523, GZHU 22961, GZHU 22962, GZHU hun22011, GZHU 22549, GZHU 22703 andthe Mu. huttoni specimens are numbered GZHU hun22001, GZHU hun22008, GZHU 22431, GZHU 22437, GZHU hun22015.