Dissecting the in Vivo Metabolic Potential of Two Human Gut Acetogens

Fermenting microbial communities generate hydrogen; its removal through the production of acetate, methane, or hydrogen sulfide modulates the efficiency of energy extraction from available nutrients in many ecosystems. We noted that pathway components for acetogenesis are more abundantly and consistently represented in the gut microbiomes of monozygotic twins and their mothers than components for methanogenesis or sulfate reduction and subsequently analyzed the metabolic potential of two sequenced human gut acetogens, Blautia hydrogenotrophica and Marvinbryantia formatexigens in vitro and in the intestines of gnotobiotic mice harboring a prominent saccharolytic bacterium. To do so, we developed a generally applicable method for multiplex sequencing of expressed microbial mRNAs (microbial RNA-Seq) and, together with mass spectrometry of metabolites, showed that these organisms have distinct patterns of substrate utilization. B. hydrogenotrophica targets aliphatic and aromatic amino acids. It increases the efficiency of fermentation by consuming reducing equivalents, thereby maintaining a high NAD⁺/NADH ratio and boosting acetate production. In contrast, M. formatexigens consumes oligosaccharides, does not impact the redox state of the gut, and boosts the yield of succinate. These findings have strategic implications for those who wish to manipulate the hydrogen economy of gut microbial communities in ways that modulate energy harvest.     

Isolation and characterization of Dehalobacterium formicoaceticum gen. nov. sp. nov., a strictly anaerobic bacterium utilizing dichloromethane as source of carbon and energy

A strictly anaerobic, dichloromethane-utilizing bacterium was isolated from a previously described dichloromethane-fermenting, two-component mixed culture. In a mineral medium with vitamins, the organism converted 5 mM dichloromethane within 7 days to formate plus acetate in a molar ratio of 2:1 and to biomass and traces of pyruvate. Of 50 potential substrates and combinations of substrates tested, only dichloromethane supported growth. The organism had a DNA G+C content of 42.7 mol%. From its phylogenetic position deduced from 16S rDNA analysis and from its unique substrate range, we conclude that the organism represents a new genus and a new species within the phylum of the gram-positive bacteria for which we propose the name Dehalobacterium formicoaceticum. Cell extracts were found to contain carbon monoxide dehydrogenase, methylene tetrahydrofolate dehydrogenase, formyl tetrahydrofolate synthetase, and hydrogenase activities, whereas activities of methenyl tetrahydrofolate cyclohydrolase and methylene tetrahydrofolate reductase were not detectable. Activity for dehalogenation of dichloromethane was lost on preparation of cell extracts, but was maintained in cell suspensions. Oxygen and reagents that react with thiol groups caused irreversible inhibition, and propyl iodide caused reversible inhibition of dehalogenation. Our observations suggest: 1) conversion of dichloromethane to methylene tetrahydrofolate, which gives rise to both formate and the methyl group of acetate, or 2) conversion of two molecules of dichloromethane to methylene tetrahydrofolate (which is oxidized to formate) and parallel reductive dehalogenation of one dichloromethane to the methyl group of the corrinoid-protein involved in acetate formation. Received: 11 March 1996 / Accepted 3 May 1996     

Ruminococcus hydrogenotrophicus sp. nov., a new H2/CO2-utilizing acetogenic bacterium isolated from human feces

A new H₂/CO₂-utilizing acetogenic bacterium was isolated from the feces of a non-methane-excreting human subject. The two strains S5a33 and S5a36 were strictly anaerobic, gram-positive, non-sporulating coccobacilli. The isolates grew autotrophically by metabolizing H₂/CO₂ to form acetate as sole metabolite and were also able to grow heterotrophically on a variety of organic compounds. The major end product of glucose and fructose fermentation was acetate; the strains also formed ethanol, lactate and, to a lesser extent, isobutyrate and isovalerate. The G+C content of DNA of strain S5a33 was 45.2 mol%. 16S rRNA gene sequencing demonstrated that the two acetogenic isolates were phylogenetically identical and represent a new subline within Clostridium cluster XIVa. Based on phenotypic and phylogenetic considerations, a new species, Ruminococcus hydrogenotrophicus, is proposed. The type strain of R. hydrogenotrophicus is S5a33 (DSM 10507). Furthermore, H₂/CO₂ acetogenesis appeared to be a common property of most of the species phylogenetically closely related to strain S5a33 (Clostridium coccoides, Ruminococcus hansenii, and Ruminococcus productus). Received: 11 April 1996 / Accepted: 11 June 1996     

Sodium bioenergetics in methanogens and acetogens

Abstract Recent investigations with Methanosarcina barkeri elucidated the role of sodium ions in the energy metabolism of methanogenic bacteria and provided evidence for a novel mechanism of energy transduction with Na⁺ as the coupling ion. During methanogenesis from methanol, an eletrochemical sodium gradient generated by a Na⁺/H⁺ antiporter is used as the driving force for the thermodynamically unfavourable oxidation of methanol to the formal redox level of formaldehyde. During methanogenesis from H₂+ CO₂, the reverse reaction, the reduction of formaldehyde to the level of methanol, is accompanied by a primary, electron transport-driven sodium extrusion. Acetogenesis from H₂+ CO₂ as carried out by Acetobacterium woodii is a sodium-dependent process and is accompanied by the generation of a transmembrane sodium gradient with the reduction of formaldehyde to the level of methanol as the sodium-dependent step.     

Re-examination of thee metabolic potentials of the acetogens Clostridium aceticum and Clostridium formicoaceticum: chemolithoautotrophic and aromatic-dependent growth

Both Clostridium formicoaceticum and Clostridium aceticum grew chemolithoautotrophically on carbon monoxide plus CO2 in defined medium in the absence of carbohydrates, amino acids, or other carbon and energy sources. Formate supported the growth of both organisms as well in both defined and undefined media (both of which also contained CO2). Hydrogen was stimulatory to the growth of C. formicoaceticum upon first transfer into H2-enriched formate medium; however, neither chemolithoautotrophic growth at the expense of H2 plus CO2 nor hydrogenase could be demonstrated with this acetogen. Consistent with recent findings with other acetogens, numerous aromatic compounds were utilized by C. aceticum and C. formicoaceticum: (i) aromatic methoxyl groups were O-demethylated; (ii) aromatic acrylates were reduced; and (iii) aromatic aldehydes were oxidized. These findings demonstrate that the metabolic potentials of these two acetogens are greater than previously recognized.