Homoacetogenic Clostridium magnum WY-4 as a novel bio-stabilizing agent in anaerobic digestion: enhanced methane yield and metabolic modulation

Anaerobic digestion is widely applied for organic waste treatment, but its efficiency is often constrained by volatile fatty acid accumulation, elevated hydrogen partial pressure and unstable microbial interactions. This study proposed using a novel homoacetogenic bacterium Clostridium magnum WY-4 (WY-4) as a bio-stabilizing agent to improve methane production of anaerobic digestion. Biochemical methane potential tests revealed a clear dose-dependent response to WY-4. The optimal dosage of WY-4 (5 %) increased methane yield by 17 % compared to the control group without WY-4, reaching 8.79 ± 0.29 mol CH₄/kg VS, while the maximum methane production rate increased by 29 % to 1.87 ± 0.10 mol CH₄/kg VS/d. Correspondingly, energy recovery reached 7046 ± 165 kJ/kg VS, which was 17 % higher than that of the control. The addition of WY-4 reduced hydrogen accumulation, increased acetate kinase activity, and lowered coenzyme F₄₂₀ levels, indicating a redirection of electron and carbon fluxes toward homoacetogenesis, which subsequently stimulated acetoclastic methanogenesis. The relative abundance of Methanosarcina increased by 5 % at the optimal WY-4 dosage. The restructuring of microbial community was accompanied by a selective shift in quorum sensing, characterized by the dominance of C₆-HSL, and a change in extracellular polymeric substances composition toward a higher protein-to-polysaccharide ratio in the loosely bound fraction. The results demonstrate that bioaugmentation with a pure homoacetogenic culture is an effective strategy for stabilizing anaerobic digestion, enhancing methane production and diverting CO₂ toward acetate formation, providing a previously underexplored route for process regulation.