Electro-fermentation enhances H2 and ethanol co-production by regulating electron transfer and substrate transmembrane transport

Electro-fermentation systems (EFSs) are an emerging technology capable of regulating microbial fermentation pathways by tuning oxidation-reduction potentials and electron flow. However, there is limited understanding of the bioenergy conversion and metabolic regulation of fermentative bacteria in EFSs. In this study, we investigated how electrode potentials in EFSs affect the metabolic products and global transcriptome expression of Ethanoligenens harbinense. The E. harbinense-inoculated anodic EFS (AEFS) with a poised potential of 0 V or the cathodic EFS (CEFS) with a poised potential of 0 V (vs. Ag/AgCl reference electrode) obtained the maximum H2 production of 1888-1986 mL/L-medium, which increased by 23-26% compared with open-circuit fermentation (OC-EFSs). The highest H2 yield of 1.190 +/- 0.009-1.197 +/- 0.001 mol-H2/mol-glucose was obtained by the AEFS0 and the OC-EFS. Ethanol production of AEFS-0.4 increased by 30.7 +/- 13.3% compared with OC-EFSs. In addition, glucose uptake and cell growth in the EFS were enhanced with an increase in cellular energy supply. Transcriptome analysis revealed that overexpression of the [FeFe]-hydrogenase, ferredoxin, and rubredoxin genes in the AEFS with a poised potential of 0 V promoted the H2 production rate. Genes involved in electron transfer and reduced nicotinamide adenine dinucleotide (NADH) regeneration were upregulated in the AEFS, leading to more ethanol production. In addition, substrate transmembrane transport was suppressed by underexpression of adenosine triphosphate (ATP)-binding cassette (ABC) transporter system-related genes at lower or higher potentials. These results confirm that an EFS effectively regulates the metabolite spectrum of H2-producing bacteria by coordinating electron transfer, NADH regeneration, and substrate transmembrane transport to provide a flexible approach for improving bioenergy production by fermentative bacteria.