Cultivation and sequencing of microbiota members unveil the functional potential of yak gut microbiota.

The animal gut microbiota exhibits extensive taxonomic diversity, yet cultivated isolates and complete genomes from animal hosts remain scarce, hindering functional and ecological insights. We present a cultivated Yak () Fecal bacteria genome Reference (YFR), comprising 548 high-quality genomes based on aerobic and anaerobic cultivation. Notably, 216 strains represented novel taxa, classified into 29 species-level clusters spanning 4 phyla and 14 genera. The YFR increased the proportion of cultured ruminant gut bacterial species by 19.39%, significantly expanding the reference database for this ecosystem. Among these, 11 species harbor abundant CAZymes Gene Clusters (CGCs), indicating a high capacity for digesting complex polysaccharides. Biosynthetic Gene Clusters (BGCs) are predicted and demonstrated to possess distinct novelty in YFR genomes, demonstrating a potential for future applications. We demonstrated that the symbiotic relationship between host bacterial strains and bacteriophages can be effectively studied using cultured strains by enabling precise mapping of viral genes to host metabolic adaptations. Culturing animal gut bacterial species not only expands the resources of culturable strains but also provides a basis for subsequent functional mining.IMPORTANCEAs a representative species in high-altitude extreme environments, yaks rely on their gut microbiota to support critical physiological functions and adapt to harsh conditions. This study established a comprehensive pipeline by integrating innovative single-bacterium culture conditions with optimized strategies for the yak gut microbiota. The resulting genomic repository not only expands the culturable microbial resources for extremophile mammals but also reveals unique metabolic traits, including polysaccharide-digesting CAZyme clusters, novel BGCs, and phage-host interactions. This approach provides essential microbial resources for advancing our understanding of host-microbial adaptations to extreme environments and offers tangible tools for industrial enzyme discovery and synthetic biology applications.