BACKGROUND: Rumen methane emissions (RMEs) significantly contribute to global greenhouse gas emissions, underscoring the essentials to identify effective inhibitors for RME mitigation. Despite various inhibitors shown potential in reducing RME by modulating rumen microbes, their impacts include considerable variations and inconsistency. OBJECTIVES: We aimed to quantitatively assess the impacts of various methane inhibitors on RME, rumen microbial abundance, and fermentation in ruminants. Additionally, the relationships between microbial abundance and RME were examined through meta-regressions. METHODS: Meta-analysis and meta-regression were conducted to assess the impacts of methane inhibitions, including 3-nitrooxypropanol, ionophores, nitrate, triglycerides, phytochemicals, and co-inhibitors, on RME and rumen microbiota in beef, dairy cattle, and sheep. RESULTS: Analyses of 922 datasets from 274 experiments revealed that inhibitors, except ionophores (P = 0.43), significantly reduced RME, with co-inhibitors displaying the highest efficacy (standardized mean difference -2.1, P <
0.01). Inhibitors' effects were more pronounced in sheep relative to beef and dairy cattle. Inhibitors decreased the abundance of ciliates and methanogens, with positive correlations observed between Dasytrichidae (P = 0.05), Entodinomorphs (P ≤ 0.001), Methanobacteriale (P = 0.001), and fungi (P <
0.01) with RME. Among inhibitors, triglycerides exhibited simultaneous reduction in methanogen, ciliate, and fungal abundances. 3-Nitrooxypropanol and triglycerides increased H CONCLUSIONS: Microbes, including Dasytrichidae, Entodinomorphs, Methanobacteriale, and fungi, significantly attribute to RME, and co-inhibitors have the highest efficacy in limiting RME and reducing microbial abundances. This study underscores the roles of both host and microbiota in modulating the inhibitor efficacy in RME, informing the refinement of rumen additives to mitigate RME from meat and milk production.