Semiquantitative Detection of Hydrogen-Associated or Hydrogen-Free Electron Transfer within Methanogenic Biofilm of Microbial Electrosynthesis

Hydrogen-entangled electron transfer has been verified as an important extracellular pathway of sharing reducing equivalents to regulate biofilm activities within a diversely anaerobic environment, especially in microbial electrosynthesis systems. However, with a lack of useful methods for in situ hydrogen detection in cathodic biofilms, the role of hydrogen involvement in electron transfer is still debatable. Here, a cathodic biofilm was constructed in CH4-produced microbial electrosynthesis reactors, in which the hydrogen evolution dynamic was analyzed to confirm the presence of hydrogen-associated electron transfer near the cathode within a micrometer scale. Fluorescent in situ hybridization images indicated that a colocalized community of archaea and bacteria developed within a 58.10-gm-thick biofilm at the cathode, suggesting that the hydrogen gradient detected by the microsensor was consumed by the collaboration of bacteria and archaea. Coupling of a microsensor and cyclic voltammetry test further provided semiquantitative results of the hydrogen-associated contribution to methane generation (around 21.20% +/- 1.57% at a potential of -0.5 V to -0.69 V). This finding provides deep insight into the mechanism of electron transfer in biofilm on conductive materials. IMPORTANCE Electron transfer from an electrode to biofilm is of great interest to the fields of microbial electrochemical technology, bioremediation, and methanogenesis. It has a promising potential application to boost more value-added products or pollutant degradation. Importantly, the ability of microbes to obtain electrons from electrodes and utilize them brings new insight into direct interspecies electron transfer during methanogenesis. Previous studies verified the direct pathway of electron transfer from the electrode to a pure-culture bacterium, but it was rarely reported how the methanogenic biofilm of mixed cultures shares electrons by a hydrogen-associated or hydrogen-free pathway. In the current study, a combination method of microsensor and cyclic voltammetry successfully semiquantified the role of hydrogen in electron transfer from an electrode to methanogenic biofilm.