Elucidating gut microbial composition profiles of Tibetan pig based on 16S rRNA gene and metagenomic sequencing

<bold>Objective</bold> This study aims to investigate the relationship between the intestinal flora of Tibetan pigs and their roughage tolerance and resilience performance by analyzing the composition of intestinal flora and its functional annotation. <bold>Method</bold> Microbial DNA was extracted from the feces of 66 Tibetan pigs，and 16S rRNA gene sequencing was used to analyze the composition of the intestinal flora of Tibetan pigs at the phylum level and genus level.A flora interactions network of shared core bacteria was constructed.Combined with the metagenomic sequencing of intestinal microbes in 27 Tibetan pigs，the annotation analysis of the KEGG pathway and the carbohydrate metabolizing enzymes （CAZymes） database were performed to elucidate functional pathways related to the fiber metabolism and disease resistance in the Tibetan pigs. <bold>Result</bold> （1） The results showed that in the study of intestinal flora composition of Tibetan pigs，16S rRNA sequencing analysis revealed that Bacteroidetes （39.34%-60.72%） and Firmicutes （24.65%-38.5%） were dominant at the level of intestinal microbial phylum，while at the genus level，<italic>Prevotella</italic>，<italic>Treponema</italic> and <italic>Succinivibrio</italic> had the highest abundance.（2） Cytoscape software was used to construct the interactions network between the 26 core genera shared in the Tibetan pig intestinal samples，and there were cooperative and competitive relationships among the groups in the network，which proved that the structure of the intestinal flora of Tibetan pigs was relatively stable.（3） Tibetan pig intestine was mainly enriched in pathways related to nutrient anabolism and genetic information processing in KEGG functional annotation based on the results of macrogenomic birdshot sequencing，including amino acid biosynthesis，carbon metabolism，and group sensing pathways.Furthermore，annotation using the CAZymes database showed the highest relative abundance of the glycosyltransferases （<italic>GTs</italic>） gene family in the intestine of Tibetan pigs，mainly <italic>GT2</italic>，<italic>GT4</italic> and <italic>GT41</italic> genes，followed by the glycoside hydrolase gene family （<italic>GHs</italic>） of <italic>GH13</italic>，<italic>GH2</italic> and <italic>GH3</italic> genes. <bold>Conclusion</bold> Tibetan pigs have rich and unique microbial composition and functional pathways.Verrucomicrobia and Akkermansia，which are related to immunity and disease resistance，are high abundance genera specific to the intestinal tract of Tibetan pigs，which may be related to the adaptation of Tibetan pigs to the environment of high altitude and low oxygen and low temperature and their disease resistance performance；those associated with crude fiber metabolism，such as <italic>Treponema</italic>，<italic>Sphaerochaeta</italic> and <italic>Fibrobacter</italic> are enriched in the intestinal tract of Tibetan pigs，which can help Tibetan pigs degrade crude fiber in the ration and adapt to the semi-grazing feeding mode.These results help us to understand the role of gut microbiota in good adaptability to roughage forage and harsh environmental stress of Tibetan pigs，and provide new ideas for the development and utilization of the excellent characteristics of Tibetan pigs in the future.