Snakes are model animals for studying vertebrate energy balance. The gut microbes help the host get energy and nutrients. Currently, researches focused on the captive animals, but little is known about the gut microbes in the wild snakes. We described intestinal microflora in large intestine, small intestine and cloaca in 3 wild Rhabdophis tigrinus by using a V4 region of 16S rRNA gene amplicon sequencing and analyzed the core composition and distribution characteristics of the intestinal microflora. The results showed no significant difference in the diversity of alpha index in different intestinal regions (Table 1). The three intestinal regions shared 534 Operational Taxonomic Units (OTUs), among which the large intestine contained 388 more unique OTUs (Fig. 3c). At the phylum level, Fusobacteria were found to be dominant in the small intestine (52.87% ± 14.49%), large intestine (41.12% ± 22.60%) and cloaca (65.70% ± 10.44%) (Fig. 1). At the genus level, we indicated that the genus of Cetobacterium had an advantage in all intestinal segments, including the small intestine (46.36% ± 13.86%), large intestine (21.95% ± 9.82%) and cloaca (58.18% ± 14.29%) (Fig. 2), and that Citrobacter showed a significant difference between small intestine and cloaca. In addition, many potentially pathogenic bacteria, for example, Citrobacter, Trichococcus and Erysipelothrix, were detected in the gut, which is helpful for. understanding the distribution of potential pathogens in the intestines of Rhabdophis tigrinus. In cluster analysis, the data repeatability of cloaca samples were well (Fig. 3d) and combined analysis with the Agkistrodon piscivorus data provided the theoretical basis for further comparison analysis of intestinal microflora among different snake species (Fig. 4).