Anodic microbial communities in acetate-fed microbial fuel cells (MFCs) were analyzed using stable-isotope probing of 16S rRNA genes followed by denaturing gradient gel electrophoresis. The results revealed that Geobacter sulfurreducens and Hydrogenophaga sp. predominated in the anodic biofilm. Although the predominance of Geobacter sp. as acetoclastic exoelectrogens in acetate-fed MFC systems has been often reported, the ecophysiological role of Hydrogenophaga sp. is unknown. Therefore, we isolated and characterized a bacterium closely related to Hydrogenophaga sp. (designated strain AR20). The newly isolated strain AR20 could use molecular hydrogen (H₂), but not acetate, with carbon electrode as the electron acceptor, indicating that the strain AR20 was a hydrogenotrophic exoelectrogen. This evidence raises a hypothesis that acetate was oxidized by G. sulfurreducens in syntrophic cooperation with the strain AR20 as a hydrogen-consuming partner in the acetate-fed MFC. To prove this hypothesis, G. sulfurreducens strain PCA was cocultivated with the strain AR20 in the acetate-fed MFC without any dissolved electron acceptors. In the coculture MFC of G. sulfurreducens and strain AR20, current generation and acetate degradation were the highest, and the growth of strain AR20 was observed. No current generation, acetate degradation and cell growth occurred in the strain AR20 pure culture MFC. These results show for the first time that G. sulfurreducens can oxidize acetate in syntrophic cooperation with the isolated Hydrogenophaga sp. strain AR20, with electrode as the electron acceptor.

Autotrophic theories for the origin of metabolism posit that the first cells satisfied their carbon needs from CO ₂ and were chemolithoautotrophs that obtained their energy and electrons from H ₂ . The acetyl-CoA pathway of CO ₂ fixation is central to that view because of its antiquity: Among known CO ₂ fixing pathways it is the only one that is i) exergonic, ii) occurs in both bacteria and archaea, and iii) can be functionally replaced in full by single transition metal catalysts in vitro. In order to operate in cells at a pH close to 7, however, the acetyl-CoA pathway requires complex multi-enzyme systems capable of flavin-based electron bifurcation that reduce low potential ferredoxin—the physiological donor of electrons in the acetyl-CoA pathway—with electrons from H ₂ . How can the acetyl-CoA pathway be primordial if it requires flavin-based electron bifurcation? Here, we show that native iron (Fe ⁰ ), but not Ni ⁰ , Co ⁰ , Mo ⁰ , NiFe, Ni ₂ Fe, Ni ₃ Fe, or Fe ₃ O ₄ , promotes the H ₂ -dependent reduction of aqueous Clostridium pasteurianum ferredoxin at pH 8.5 or higher within a few hours at 40 °C, providing the physiological function of flavin-based electron bifurcation, but without the help of enzymes or organic redox cofactors. H ₂ -dependent ferredoxin reduction by iron ties primordial ferredoxin reduction and early metabolic evolution to a chemical process in the Earth’s crust promoted by solid-state iron, a metal that is still deposited in serpentinizing hydrothermal vents today.

A Gram-stain-negative, non-spore-forming, rod-shaped bacterium, designated strain 1GB^T, was isolated from anodic biofilms of a glucose-fed microbial fuel cell. Strain 1GB^T was facultatively anaerobic and chemo-organotrophic, having both a respiratory and a fermentative type of metabolism, and utilized a wide variety of sugars as carbon and energy sources. Cells grown aerobically contained Q-8 as the major quinone, but excreted Q-9 and a small amount of Q-10 when cultured with an electrode serving as the sole electron acceptor. The G+C content of the genomic DNA of 1GB^T was 54.5 mol%. Multilocus sequence typing (MLST) analysis showed that strain 1GB^T represented a distinct lineage within the genus Raoultella (98.5–99.4 % 16S rRNA gene sequence similarity and 94.0–96.5 % sequence similarity based on the three concatenated housekeeping genes gyrA, rpoB and parC. Strain 1GB^T exhibited DNA–DNA hybridization relatedness of 7–43 % with type strains of all established species of the genus Raoultella . On the basis of these phenotypic, phylogenetic and genotypic data, the name Raoultella electrica sp. nov. is proposed for strain 1GB^T. The type strain is 1GB^T ( = NBRC 109676^T = KCTC 32430^T).

A novel oligotrophic bacterium, designated strain CCA6, was isolated from leaf soil collected in Japan. Cells of the strain were found to be a Gram‐negative, non‐sporulating, motile, rod‐shaped bacterium. Strain CCA6 grew at 10–45°C (optimum 20°C) and pH 4.5–10.0 (optimum pH 5.0). The strain was capable of growth in poor‐nutrient (oligotrophic) medium, and growth was unaffected by high‐nutrient medium. The major fatty acid and predominant quinone system were C_16:0 and ubiquinone‐8. Phylogenetic analysis based on 16S rRNA gene sequences indicated strain CCA6 presented as a member of the family Enterobacteriaceae. Multilocus sequence analysis (MLSA) based on fragments of the atpD, gyrB, infB, and rpoB gene sequences was performed to further identify strain CCA6. The MLSA showed clear branching of strain CCA6 with respect to Enterobacter type strains. The complete genome of strain CCA6 consisted of 4,476,585 bp with a G+C content of 54.3% and comprising 4,372 predicted coding sequences. The genome average nucleotide identity values between strain CCA6 and the closest related Enterobacter type strain were <88.02%. Based on its phenotypic, chemotaxonomic and phylogenetic features, strain CCA6 (=HUT 8142^T =KCTC 62525^T) can be considered as a novel species within the genus Enterobacter with the proposed name Enterobacter oligotrophica.

A Gram-negative, non-spore-forming, rod-shaped bacterial strain, AR20T, was isolated from anodic biofilms of an acetate-fed microbial fuel cell in Japan and subjected to a polyphasic taxonomic study. Strain AR20T grew optimally at pH 7.0−8.0 and 25°C. It contained Q-8 as the predominant ubiquinone and C16:0, summed feature 3 (C16:1ω7c and/or iso-C15:02OH), and C18:1ω7c as the major fatty acids. The DNA G+C content was 67.1 mol%. A neighbor-joining phylogenetic tree revealed that strain AR20T clustered with three type strains of the genus Hydrogenophaga (H. flava, H. bisanensis and H. pseudoflava). Strain AR20T exhibited 16S rRNA gene sequence similarity values of 95.8−97.7% to the type strains of the genus Hydrogenophaga. On the basis of phenotypic, chemotaxonomic and phylogenetic data, strain AR20T is considered a novel species of the genus Hydrogenophaga, for which the name Hydrogenophaga electricum sp. nov. is proposed. The type strain is AR20T (= KCTC 32195T = NBRC 109341T).