Isolation of Ultra-Small Opitutaceae-Affiliated Verrucomicrobia from a Methane-Fed Bioreactor

The bacterial phylum <i>Verrucomicrobiota</i> accommodates free-living and symbiotic microorganisms, which inhabit a wide range of environments and specialize in polysaccharide degradation. Due to difficulties in cultivation, much of the currently available knowledge about these bacteria originated from cultivation-independent studies. A phylogenetic clade defined by the free-living bacterium from oilsands tailings pond, <i>Oleiharenicola alkalitolerans</i>, and the symbiont of the tunicate <i>Lissoclinum</i> sp., <i>Candidatus</i> Didemniditutus mandelae, is a poorly studied verrucomicrobial group. This clade includes two dozen methagenome-assembled genomes (MAGs) retrieved from aquatic and soil habitats all over the world. A new member of this clade, strain Vm1, was isolated from a methane-fed laboratory bioreactor with a <i>Methylococcus</i>-dominated methane-oxidizing consortium and characterized in this study. Strain Vm1 was represented by ultra-small, motile cocci with a mean diameter of 0.4 µm that grew in oxic and micro-oxic conditions at temperatures between 20 and 42 °C. Stable development of strain Vm1 in a co-culture with <i>Methylococcus</i> was due to the ability to utilize organic acids excreted by the methanotroph and its exopolysaccharides. The finished genome of strain Vm1 was 4.8 Mb in size and contained about 4200 predicted protein-coding sequences, including a wide repertoire of CAZyme-encoding genes. Among these CAZymes, two proteins presumably responsible for xylan and arabinan degradation, were encoded in several MAGs of Vm1-related free-living verrucomicrobia, thus offering an insight into the reasons behind wide distribution of these bacteria in the environment. Apparently, many representatives of the <i>Oleiharenicola</i>-<i>Candidatus</i> Didemniditutus clade may occur in nature in trophic associations with methanotrophic bacteria, thus participating in the cycling of methane-derived carbon.