Coordinated Interfacial Electron Transfer and Metabolic Coupling Drive Acetate Electrosynthesis in a Synthetic Microbial Consortium

Microbial consortia in microbial electrosynthesis (MES) can couple cathodic electron utilization with CO2 fixation through interspecies electron transfer and metabolite cross-feeding. However, how defined functional members coordinate interfacial electron input with downstream acetogenic metabolism remains insufficiently understood. Here, we constructed a defined three-member consortium comprising the Shewanella oneidensis MR-1 and the acetogens Clostridium aceticum and Acetobacterium woodii to investigate how functional complementarity supports acetate electrosynthesis. The three-member consortium reached a maximum acetate concentration of 2.33 ± 0.17 g L–1, 2.62-fold that of the A. woodii and C. aceticum binary consortium, accompanied by a higher current density, stronger redox activity, and more pronounced riboflavin-related signals. Phase-resolved fhs abundances, metabolomic profiles, and flavin perturbation experiments further supported spatially differentiated roles within the consortium: S. oneidensis enhanced cathode-associated electron input, while phase-resolved fhs abundances and H2/formate dynamics indicated that acetogenic functions were maintained across both biofilm and planktonic phases and coupled to soluble-intermediate turnover. Despite varying initial inoculation ratios, the consortium consistently converged on similar acetate production and community functions, indicating that steady-state performance of the consortium was not strongly dependent on the precise initial community composition. These results suggest that the enhanced performance of the three-member consortium was associated with coordinated electron transfer and metabolic handoff among functionally complementary members. This study demonstrates that enhanced CO2 electrosynthesis arises from the reorganization of electron-metabolic interactions rather than the simple addition of functional members, providing a mechanistic basis for the rational design of electrically driven synthetic microbial communities.