Directed microbial community assembly enhances biohydrogen production from cheese whey during continuous dark fermentation

Dark fermentation of cheese whey represents a promising strategy for sustainable biohydrogen production. In this study, a microbial community assembly strategy was evaluated to promote synergistic interactions between native lactic acid bacteria (LAB) rich communities and hydrogen-producing bacteria (HPB) enriched from anaerobic sludge. Continuous reactors were operated under different HPB/native LAB inoculum ratios (RHN) and hydraulic residence times (HRT = 6 and 10 h). Hydrogen production rates reached up to 3.84 L H 2 L −1 d −1 , with the highest values observed under RHN = 0.3 at an HRT of 10 h. Microbial community taxonomy was restructured, with up to 98% of the community composed of emergent amplicon sequence variants (ASVs) related to fermentative lineages such as Clostridium , Megasphaera , and Anaerotignum . Functional prediction indicated the prevalence of hydrogen-associated pathways. These results demonstrate that directed microbial community assembly can promote functional redundancy, enabling efficient hydrogen production while reducing reliance on specialized inocula.