Summary auto-generated
Two strictly anaerobic bacterial strains (YUAN-3T and X-29) were isolated from molasses wastewater in a hydrogen-producing bioreactor. Both isolates are Gram-positive, non-spore-forming, motile rods that ferment various sugars to produce acetate, ethanol, hydrogen, and carbon dioxide. The strains grew optimally at 35°C and pH 4.5–5.0, with generation times of 4.2 hours. Phylogenetic analysis of 16S rRNA gene sequences revealed they belong to the Clostridium leptum subgroup but showed only 91.8% similarity to the closest known species (C. cellulosi), indicating a novel genus. The isolates possessed distinctive features including branched-chain fatty acids (predominantly iso-C16:0 and iso-C12:0), LL-DAP in cell walls, and DNA G+C contents of 47.8–49.0 mol%. Strain YUAN-3T exhibited autoaggregation and autofluorescence, while strain X-29 did not, though both shared identical physiological characteristics. Based on polyphasic evidence including phylogenetic distance, unique chemotaxonomic traits, and biochemical properties, the authors propose a new genus and species: Ethanoligenens harbinense, with YUAN-3T designated as the type strain.
Key findings
- Two novel strictly anaerobic bacterial strains isolated from molasses wastewater represent a new genus, Ethanoligenens harbinense, within the Clostridium leptum rRNA subgroup with <92% 16S rRNA similarity to known species
- Both strains fermented sugars to produce major fermentation products of ethanol, acetate, hydrogen, and carbon dioxide, with maximal H₂ yield of 2.81 mol/mol glucose for strain YUAN-3T
- The isolates possessed distinctive branched-chain fatty acid profiles (44.18% iso-C16:0 and 26.67% iso-C12:0) and LL-DAP cell wall composition, differing notably from related Clostridium species
- Strain YUAN-3T formed large autoaggregative granules and was autofluorescent, while strain X-29 remained non-aggregative, though both shared 100% 16S rRNA gene sequence similarity and identical physiological traits
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Abstract
Two strictly anaerobic bacterial strains (YUAN-3T and X-29) were isolated from anaerobic activated sludge of molasses wastewater in a continuous stirred-tank reactor. The strains were Gram-positive, non-spore-forming, mesophilic and motile. Cells were regular rods (0·40·8x1·58·0 µm) and occurred singly, in pairs and sometimes in chains of up to eight. Autoaggregative and autofluorescent growth of strain YUAN-3T and non-aggregative growth of strain X-29 were observed at 2044 °C and pH 3·59·0. Both strains hydrolysed gelatin and aesculin and fermented several kinds of mono-, di- and oligosaccharides. Fermentation end products formed from glucose were acetate, ethanol, hydrogen and carbon dioxide. The predominant cellular fatty acids were the branched-chain fatty acids iso-C16 : 0 (44·18 %) and iso-C12 : 0 (26·67 %). The DNA G+C contents of strains YUAN-3T and X-29 were 47·8 and 49·0 mol%, respectively. Phylogenetic analysis based on 16S rRNA gene sequences revealed that the isolates represent a novel phyletic sublineage within the Clostridium cellulosi rRNA cluster, with <92 % 16S rRNA gene sequence similarity to currently known species. On the basis of polyphasic evidence from this study, it is proposed that the unknown bacterium should be classified in a new genus as a novel species, Ethanoligenens harbinense gen. nov., sp. nov. The type strain of Ethanoligenens harbinense is YUAN-3T (=JCM 12961T=CGMCC 1.5033T).