New Insights into Microbial Nitrogen Utilization in the Rumen Enabled by Genome-Resolved Multi-Omics

Optimizing nitrogen (N) utilization in ruminant production systems holds both economic and environmental significance. However, traditional paradigms of N metabolism, derived primarily from well-studied model rumen bacteria, may not fully capture the diverse and complex N metabolic dynamics within the rumen ecosystem. To address this gap, we utilized comparative genomics and genome-resolved multi-omics analyses to investigate N assimilation and regulation in rumen microbes. We discovered that canonical ammonia assimilation and regulation mechanisms, such as the glutamine synthetase (GS)/glutamate synthase (GOGAT) (GS-GOGAT) pathway and its regulatory proteins, are absent in many rumen bacteria, which appear to utilize alternative pathways for ammonia assimilation. These findings challenge the applicability of E. coli-based N regulation models to rumen bacteria. We further linked polysaccharides utilization and N assimilation across hundreds of rumen microbial species. Furthermore, we identified specific microbial species involved in ureolysis, denitrification, and phages carrying auxiliary metabolic genes that perform N assimilation. Using an animal trial with 11 pairs of lamb twins in a crossover design, we demonstrated that dietary crude protein (CP) levels had minimal impact on rumen microbiome composition and expression of N assimilation genes. Instead, dietary shifts triggered alternative responses, including increased expression of amino acid biosynthesis pathways. These findings indicate a nuanced, species-specific microbial response to dietary interventions, highlighting the limitations of traditional N metabolism models and the need for more granular studies of rumen microbial ecosystems. This study provides foundational insights for developing tailored dietary strategies to enhance N utilization efficiency in ruminants.