A Novel Slowly Evolving Lineage of the <i>Desulforudis</i> Clade From the Deep Subsurface

Endemic to the deep subsurface biosphere sulphate-reducing 'Desulforudis audaxviator' has been called a living microbial fossil due to the high nucleotide sequence identity of its genomes across continents. Evolutionary stasis of this bacterium was established based on the analysis of metagenome assembled genomes, single cell genomes and a single axenic culture. The lack of high-quality reference genomes necessitates efforts to cultivate and isolate pure cultures that could shed light on the hypothetical slow evolution of Desulforudis-clade bacteria deep underground. Molecular signatures demonstrated the presence of Desulforudis-like phylotypes in subsurface environments worldwide. Here we report the isolation of four novel strains of the Desulforudis-clade, all belonging to Desulfosceptrum tomskiensis gen. nov. sp. nov. Four strains of the new species were isolated from deep boreholes in Western Siberia, separated by hundreds of kilometres. Genome comparisons revealed minimal differences between these strains, with average nucleotide sequence identity (ANI) values above 99.9%, low number of SNPs, and near-identical CRISPRs. The bacterium, together with Desulforudis audaxviator BYF^T gen. nov. sp. nov., deposited in international culture collections, provides a bases for understanding the slow evolution of Bacillota endemic to the deep biosphere.