Sustained anaerobic valorization of refractory wastewater into bioenergy via abiotic-biotic hybrid systems: Unraveling synergistic hydrolysis and optimized interspecies electron transfer.

Anaerobic digestion (AD) of refractory industrial wastewater remains challenging due to structural recalcitrance and thermodynamic limitations. While abiotic-biotic hybrid systems incorporating electroactive materials (EMs) offer promise, their long-term efficacy and mechanisms at a system level, particularly with real industrial waste streams, are underexplored. This study demonstrates the sustained, system-level enhancement of anaerobic valorization using magnetite (Mag)- or pyrogenic carbon (PC)-amended reactors during 140-day continuous operation treating real membrane-making wastewater laden with N,N-dimethylformamide (DMF). EMs-augmented systems achieved superior COD removal of 75.5 % or 67.0 % and methane production of 255 or 228 mL/(g VSS·day), respectively, at an organic loading rate of 10.0 g COD/(L·day), representing an 88.5 % or 68.6 % increase in methanogenesis over the control. The enhancement stemmed from EMs synergistically boosting enzymatic hydrolysis to deconstruct refractory amide bonds and conjugated structures, and redirecting interspecies electron transfer (IET) pathways with greater efficiency and conductivity. Mag excelled under shock loads due to conductive Fe(III/II) lattice, whereas PC performed better under moderate loads leveraging pseudocapacitive functionalities. Microbial evolution revealed EM-specific colonization of hydrolytic/acidogenic consortia and IET-active methanogens. Metagenomic reconstruction indicated EMs enhanced cleavage of recalcitrant bonds, redirected DMF biodegradation toward (di)methylamine dehydrogenases-catalyzed hydrolysis pathway, and stimulated CO2-reducing/acetoclastic methanogenesis. Furthermore, EMs appeared to reconstruct intra-extracellular IET channels via membrane-spanning redox-active complexes, reinforcing energy conservation and stress resilience while channeling electrons from fermentation to methanogenesis. These findings establish a sustainable strategy for valorizing recalcitrant wastewater into bioenergy, offering insights into microbial electron flux redirection in abiotic-biotic systems.