Methane (CH₄), a significant greenhouse gas, ranks second only to carbon dioxide in its contribution to global warming. The application of Chinese herbs as a strategy to mitigate CH₄ emissions in ruminants has shown promise. However, there is limited information regarding the efficacy of Chinese herb straw in reducing CH₄ emissions in ruminants. This research aimed to investigate the beneficial effects of varying levels of Astragalus straw supplementation on methane emissions and to elucidate the underlying molecular mechanisms. The study examined the effects of different supplementation levels (0%, 5%, 10%, 15%, 20%) on in vitro rumen fermentation, CH₄ emissions, and ruminal microbial community in Lanzhou fat-tailed sheep using an in vitro fermentation method. The findings indicated that IVDMD, gas production, and CH₄ production significantly decreased with increasing levels of Astragalus straw supplementation (p < 0.05). Simultaneously, the lowest levels of AA, AA/PA, and NH₃-N, along with the highest concentrations of PA, BA, and MCP, were observed in the 20% supplementation group after 48 h of fermentation. In addition, supplementation with Astragalus straw resulted in an increased abundance of Bacteroidota, Spirochaetota, and Actinobacteriota, while decreasing the abundance of Firmicutes, Fibrobacterota, and Verrucomicrobiota. At the genus level, there was an observed increase in the abundance of Prevotella and Streptococcus, accompanied by a decrease in Rikenellaceae_RC9_gut_group. In conclusion, the supplementation of Astragalus straw has the potential to reduce CH₄ production by altering ruminal fermentation patterns, fermentation parameters, and microbial dynamics.

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