Phylogenetic Diversity and Single-Cell Genome Analysis of “Melainabacteria”, a Non-Photosynthetic Cyanobacterial Group, in the Termite Gut

Termite guts harbor diverse yet-uncultured bacteria, including a non-photosynthetic cyanobacterial group, the class “Melainabacteria”. We herein reported the phylogenetic diversity of “Melainabacteria” in the guts of diverse termites and conducted a single-cell genome analysis of a melainabacterium obtained from the gut of the termite Termes propinquus. We performed amplicon sequencing of 16S rRNA genes from the guts of 60 termite and eight cockroach species, and detected melainabacterial sequences in 48 out of the 68 insect species, albeit with low abundances (0.02–1.90%). Most of the melainabacterial sequences obtained were assigned to the order “Gastranaerophilales” and appeared to form clusters unique to termites and cockroaches. A single-cell genome of a melainabacterium, designated phylotype Tpq-Mel-01, was obtained using a fluorescence-activated cell sorter and whole genome amplification. The genome shared basic features with other melainabacterial genomes previously reconstructed from the metagenomes of human and koala feces. The bacterium had a small genome (~1.6 Mb) and possessed fermentative pathways possibly using sugars and chitobiose as carbon and energy sources, while the pathways for photosynthesis and carbon fixation were not found. The genome contained genes for flagellar components and chemotaxis; therefore, the bacterium is likely motile. A fluorescence in situ hybridization analysis showed that the cells of Tpq-Mel-01 and/or its close relatives are short rods with the dimensions of 1.1±0.2 μm by 0.5±0.1 μm; for these bacteria, we propose the novel species, “Candidatus Gastranaerophilus termiticola”. Our results provide fundamental information on “Melainabacteria” in the termite gut and expand our knowledge on this underrepresented, non-photosynthetic cyanobacterial group.