Arousing the bioelectroactivity and syntrophic metabolic functionality of microorganisms in a recirculated two-phase anaerobic digestion bioreactor for enhanced biohythane recovery from high-solids biowaste

Recirculated two-phase anaerobic digestion (R-TPAD) offers a promising approach for converting organic wastes into value-added chemicals such as biohythane. However, the inherent defects of slow hydrolysis-acidification, low H₂ yield, toxicity of metabolic intermediates from methanogenesis phase limit the practical application of such biotechnology. This study here developed a bioelectro-hybrid R-TPAD system by incorporating an in-situ bioelectrochemical intervention therapy into the acidification phase to evoke bioelectroactivity and syntrophic metabolic functionality of microorganisms for higher biohythane recovery. H₂ biotransformation process, impact factors and metabolic mechanisms were analyzed comprehensively. The results revealed that the highest biohythane yield (69.3 ± 7.6 %, 32.4 ± 3.5 H₂ mL/g COD; and 71.9 ± 3 %, 223.5 ± 20.1 CH₄ mL/g COD) was achieved at an applied voltage of 0.8 V and a digestate recirculation ratio of 0.3, resulting in the highest energy recovery of 22.1 kWh/m³. The bioelectrochemical intervention effectively boosted electron transfer activity and conductivity of sludge suspension while mediating the development and spatial distribution of the electrobiofilm on electrode surface by provoking the secretions of extracellular biopolymers. The syntrophic interactions of co-cultured Thermoanaerobacterium and Clostridium were enhanced by in-situ bioelectrochemical intervention, sustaining a robust proliferating and co-existing circumstance for H₂-producing microorganisms. The metagenomic analysis further demonstrated that bioelectrochemical intervention upregulated key functional gene families associated with H₂ production (such as por, acsB, buk, and ack) by 1.9–2.9-fold, promoting the potential genetic information exchange and syntrophic metabolic functionality of anaerobic microorganisms. These findings outline the key role of in-situ bioelectrochemical intervention in arousing the bioelectroactivity and multiple syntrophic metabolic functionalities of microorganisms required for biohythane recovery and provide an alternative strategy for advancing the practical applications of R-TPAD technology in real-world biowaste management.