Abstract
BackgroundThere is a dearth of sequenced and closed microbial genomes from environments that exceed > 500 m below level terrestrial surface. Coupled with even fewer cultured isolates, study and understanding of how life endures in the extreme oligotrophic subsurface environments is greatly hindered. Using a de novo hybrid assembly of Illumina and Oxford Nanopore sequences we produced a circular genome with corresponding methylome profile of the recently characterized thermophilic, anaerobic, and fumarate-respiring subsurface bacterium, Thermanaerosceptrum fracticalcis, strain DRI-13T to understand how this microorganism survives the deep subsurface.ResultsThe hybrid assembly produced a single circular genome of 3.8 Mb in length with an overall GC content of 45%. Out of the total 4022 annotated genes, 3884 are protein coding, 87 are RNA encoding genes, and the remaining 51 genes were associated with regulatory features of the genome including riboswitches and T-box leader sequences. Approximately 24% of the protein coding genes were hypothetical. Analysis of strain DRI-13T genome revealed: 1) energy conservation by bifurcation hydrogenase when growing on fumarate, 2) four novel bacterial prophages, 3) methylation profile including 76.4% N6-methyladenine and 3.81% 5-methylcytosine corresponding to novel DNA methyltransferase motifs. As well a cluster of 45 genes of unknown protein families that have enriched DNA mCpG proximal to the transcription start sites, and 4) discovery of a putative core of bacteriophage exclusion (BREX) genes surrounded by hypothetical proteins, with predicted functions as helicases, nucleases, and exonucleases.ConclusionsThe de novo hybrid assembly of strain DRI-13T genome has provided a more contiguous and accurate view of the subsurface bacterium T. fracticalcis, strain DRI-13T. This genome analysis reveals a physiological focus supporting syntrophy, non-homologous double stranded DNA repair, mobility/adherence/chemotaxis, unique methylome profile/recognized motifs, and a BREX defense system. The key to microbial subsurface survival may not rest on genetic diversity, but rather through specific syntrophy niches and novel methylation strategies.
Highlights
There is a dearth of sequenced and closed microbial genomes from environments that exceed > 500 m below level terrestrial surface
SPAdes-Hybrid resulted in a fragmented assembly with high numbers of contigs but was able to produce a large contig of the expected length; it contained gaps filled with ambiguous bases, which could not be resolved
In this study, we used a hybrid genome assembly of Thermanaerosceptrum fracticalcis strain DRI-13T we could increase the detail of metabolism, respiration, DNA repair, ABC transporters, and cellular defense and introduce the first view of the methylome of this subsurface bacterium
Summary
There is a dearth of sequenced and closed microbial genomes from environments that exceed > 500 m below level terrestrial surface. Using a de novo hybrid assembly of Illumina and Oxford Nanopore sequences we produced a circular genome with corresponding methylome profile of the recently characterized thermophilic, anaerobic, and fumarate-respiring subsurface bacterium, Thermanaerosceptrum fracticalcis, strain DRI-13T to understand how this microorganism survives the deep subsurface. Previous studies predict that subsurface life is primarily limited by the availability of energy and nutrients from the environment [6]. Extensive commensal relationships do not adequately explain a subsurface ecosystem steady state lasting thousand if not millions of years [14]. This mystery is further deepened when considering how microorganisms must manage energy expenditures to survive, defend against viral predation, DNA repair, avert entropy, and compete with other microorganisms for resources [15]
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