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Introduction: Bamboo rats are rodents that eat bamboo, and their robust capacity for bamboo digestion is directly correlated with their gut flora. Chinese bamboo rat ( Rhizomys sinensis ) is a common bamboo rat in Chinese central and southern regions. As a single-stomach mammal, bamboo rats are a famous specificity bamboo-eating animal and their intestinal microbial composition may also play a key role in the digestion of cellulose and lignin. So, the gut microbiota of bamboo rat may play an important role in the adaptation of bamboo rats for digesting lignocellulose-based diet., Methods: To study the microbiome differences of bamboo rats from different sexes, the microbial genomic DNA was extracted from each fecal sample and the V4 region of 16S rRNA genes was amplified and sequencing on an IlluminaHiSeq6000 platform. The operational taxonomic units (OTUs) were classified, the OTUs in different sexes was identified and compared at phylum and genus levels. For isolation and screening of cellulose degradation bacteria from bamboo rats, fresh feces from randomly selected bamboo rats were collected and used for the isolation and screening of cellulose degradation bacteria using Luria Bertani (LB) Agar medium containing Carboxymethyl cellulose. The cellulase activity, biochemical characterization and phylogenetic analysis of the purified bacteria strains were characterized., Results and Discussion: A total of 3,833 OTUs were classified. The total microbial diversity detected in the female and male rats was 3,049 OTUs and 3,452 OTUs, respectively. The Shannon index revealed significant differences between the two groups ( p < 0.05), though they were all captive and had the same feeding conditions. At the phylum level, Firmicutes, Bacteroidota , and Proteobacteria were prominent in the microbial community. At the genus level, the microbial community was dominated by Lachnospiraceae, Lactobacillus, Bacteroides , and Prevotella , but there was a significant difference between the two groups of bamboo rats; ~90 bacteria genus in the female group was significantly higher than the male group. Among them, Bacteroides, Colidextribacter , and Oscillibacter were significantly higher genera, and the genera of Lachnoclostridium, Oscillibacter , and Papillibacter had the highest FC value among the male and female bamboo rats. The KEGG function annotation and different pathways analysis revealed that membrane transport, carbohydrate metabolism, and amino acid metabolism were the most enriched metabolic pathways in the two groups, and multiple sugar transport system permease protein (K02025 and K02026), RNA polymerase sigma-70 factor (K03088), and ATP-binding cassette (K06147) were the three different KEGG pathways ( p < 0.05). Two cellulose degradation bacteria strains- Bacillus subtilis and Enterococcus faecalis -were isolated and characterized from the feces of bamboo rats., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Gan, Wu, Dong, Li, Wu, Jin and Lu.)