Chemolithotrophy in the continental deep subsurface: Sanford Underground Research Facility (SURF), USA

The deep subsurface is an enormous repository of microbial life. However, the metabolic capabilities of these microorganisms and the degree to which they are dependent on surface processes are largely unknown. Due to the logistical difficulty of sampling and inherent heterogeneity, the microbial populations of the terrestrial subsurface are poorly characterized. In an effort to better understand the biogeochemistry of deep terrestrial habitats, we evaluate the energetic yield of chemolithotrophic metabolisms and microbial diversity in the Sanford Underground Research Facility (SURF) in the former Homestake Gold Mine, SD, USA. Geochemical data, energetic modeling, and DNA sequencing were combined with principle component analysis to describe this deep (down to 8100 ft below surface), terrestrial environment. SURF provides access into an iron-rich Paleoproterozoic metasedimentary deposit that contains deeply circulating groundwater. Geochemical analyses of subsurface fluids reveal enormous geochemical diversity ranging widely in salinity, oxidation state (ORP 330 to −328 mV), and concentrations of redox sensitive species (e.g., Fe(2+) from near 0 to 6.2 mg/L and Σ S(2-) from 7 to 2778μg/L). As a direct result of this compositional buffet, Gibbs energy calculations reveal an abundance of energy for microorganisms from the oxidation of sulfur, iron, nitrogen, methane, and manganese. Pyrotag DNA sequencing reveals diverse communities of chemolithoautotrophs, thermophiles, aerobic and anaerobic heterotrophs, and numerous uncultivated clades. Extrapolated across the mine footprint, these data suggest a complex spatial mosaic of subsurface primary productivity that is in good agreement with predicted energy yields. Notably, we report Gibbs energy normalized both per mole of reaction and per kg fluid (energy density) and find the later to be more consistent with observed physiologies and environmental conditions. Further application of this approach will significantly expand our understanding of the deep terrestrial biosphere

Genomic Description of ‘Candidatus Abyssubacteria,’ a Novel Subsurface Lineage Within the Candidate Phylum Hydrogenedentes

The subsurface biosphere is a massive repository of fixed carbon, harboring approximately 90% of Earth’s microbial biomass. These microbial communities drive transformations central to Earth’s biogeochemical cycles. However, there is still much we do not understand about how complex subterranean microbial communities survive and how they interact with these cycles. Recent metagenomic investigation of deeply circulating terrestrial subsurface fluids revealed the presence of several novel lineages of bacteria. In one particular example, phylogenomic analyses do not converge on any one previously identified taxon; here we describe the first full genomic sequences of a new bacterial lineage within the candidate phylum Hydrogenedentes, ‘Candidatus Abyssubacteria.’ A global survey revealed that members of this proposed lineage are widely distributed in both marine and terrestrial subsurface environments, but their physiological and ecological roles have remained unexplored. Two high quality metagenome assembled genomes (SURF_5: 97%, 4%; SURF_17: 91% and 4% completeness and contamination, respectively) were reconstructed from fluids collected 1.5 kilometers below surface in the former Homestake gold mine—now the Sanford Underground Research Facility (SURF)—in Lead, South Dakota, United States. Metabolic reconstruction suggests versatile metabolic capability, including possible nitrogen reduction, sulfite oxidation, sulfate reduction and homoacetogenesis. This first glimpse into the metabolic capabilities of these cosmopolitan bacteria suggests that they are involved in key geochemical processes, including sulfur, nitrogen, and carbon cycling, and that they are adapted to survival in the dark, often anoxic, subsurface biosphere

A FASTA file containing 44 SSU rRNA genes with length >300 base pairs, including 40 extracted from the 77 genomes from metagenomes

A FASTA file containing 44 SSU rRNA genes with length >300 base pairs, including 40 extracted from the 77 genomes from metagenomes, plus 4 additional SSU rRNA genes identified in preliminary (i.e. non-reported) GFM

Annotation- Pfam and KO- files for all 20 Chloroflexi genomes

text files with complete annotations for each genome, individuall

Horizontally transferred gene files

Amino acid sequence files and alignments used to build trees in manuscript figures

Energy and carbon metabolisms in a deep terrestrial subsurface fluid microbial community.

The terrestrial deep subsurface is a huge repository of microbial biomass, but in relation to its size and physical heterogeneity, few sites have been investigated in detail. Here, we applied a culture-independent metagenomic approach to characterize the microbial community composition in deep (1500 meters below surface) terrestrial fluids. Samples were collected from a former gold mine in Lead, South Dakota, USA, now Sanford Underground Research Facility (SURF). We reconstructed 74 genomes from metagenomes (MAGs), enabling the identification of common metabolic pathways. Sulfate and nitrate/nitrite reduction were the most common putative energy metabolisms. Complete pathways for autotrophic carbon fixation were found in more than half of the MAGs, with the reductive acetyl-CoA pathway by far the most common. Nearly 40% (29 of 74) of the recovered MAGs belong to bacterial phyla without any cultivated members-microbial dark matter. Three of our MAGs constitute two novel phyla previously only identified in 16 S rRNA gene surveys. The uniqueness of this data set-its physical depth in the terrestrial subsurface, the relative abundance and completeness of microbial dark matter genomes and the overall diversity of this physically deep, dark, community-make it an invaluable addition to our knowledge of deep subsurface microbial ecology