mitigates ruminant methane emissions via microbial modulation and inhibition of methyl-coenzyme M reductase.

INTRODUCTION

() has shown great potential to mitigate methane (CH) emissions in recent years. This study aims to evaluate the impact of on methane emissions and to fill the knowledge gap regarding its mechanisms of action in affecting CH metabolism and rumen fermentation.

METHODS

The experimental design consisted of a control group (CON) and test groups supplemented with 2% (Low), 5% (Mid), and 10% (High) of dried and freeze-dried treatment , respectively, for 48 h of rumen fermentation. The optimal combination strategy for mitigating CH emissions was confirmed by analyzing nutrient degradation, CH production and rumen fermentation parameters, and the mechanism of action was analyzed by metagenomic and metabolomic approaches.

RESULTS AND DISCUSSION

The results showed that freeze-dried treatment had better potential to mitigate CH emissions than dried treatment, and supplementation of freeze-dried treatments at Low, Mid, and High groups significantly reduced CH production by 32.44%, 98.53%, and 99.33%, respectively. However, the High group exhibited a huge negative impact on rumen fermentation. Therefore, subsequent analyses focused on the Low and Mid groups to explore the underlying mechanisms. Metagenomics analyses showed that supplementation of freeze-dried treatment with the Mid-level supplementation significantly increased the relative abundance of propionate-producing bacteria such as , , and , while inhibited acetate-producing bacteria such as , altered the pattern of volatile fatty acid (VFA) synthesis in the rumen, and reduced H availability for methanogenesis and promoted propionate production, indirectly alleviating CH production. Moreover, by suppressing the relative abundance of , CH production in the rumen was directly suppressed. Furthermore, KEGG pathway analysis showed that significantly inhibited the abundance of K00399, methyl-coenzyme M reductase alpha subunit, which directly inhibited CH synthesis. Metabolomics analysis of supplementation significantly enriched ketoglutarate, malate, isocitrate, and melatonin, which may have reduced the release of rumen fermented H, thereby mitigating CH emissions. In summary, freeze-dried treatment at the 5% supplementation level achieved the optimal balance between CH mitigation and rumen fermentation efficiency.