A metagenomic window into carbon metabolism at 3 km depth in Precambrian continental crust

Abstract

Subsurface microbial communities comprise a significant fraction of the global prokaryotic biomass; however, the carbon metabolisms that support the deep biosphere have been relatively unexplored. In order to determine the predominant carbon metabolisms within a 3-km deep fracture fluid system accessed via the Tau Tona gold mine (Witwatersrand Basin, South Africa), metagenomic and thermodynamic analyses were combined. Within our system of study, the energy-conserving reductive acetyl-CoA (Wood-Ljungdahl) pathway was found to be the most abundant carbon fixation pathway identified in the metagenome. Carbon monoxide dehydrogenase genes that have the potential to participate in (1) both autotrophic and heterotrophic metabolisms through the reversible oxidization of CO and subsequent transfer of electrons for sulfate reduction, (2) direct utilization of H2 and (3) methanogenesis were identified. The most abundant members of the metagenome belonged to Euryarchaeota (22%) and Firmicutes (57%)-by far, the highest relative abundance of Euryarchaeota yet reported from deep fracture fluids in South Africa and one of only five Firmicutes-dominated deep fracture fluids identified in the region. Importantly, by combining the metagenomics data and thermodynamic modeling of this study with previously published isotopic and community composition data from the South African subsurface, we are able to demonstrate that Firmicutes-dominated communities are associated with a particular hydrogeologic environment, specifically the older, more saline and more reducing waters.

Figure 1

Carbon cycle-specific genes. The six carbon fixation pathways (3-hydtroxypropionate bicycle, Calvin cycle, dicarboxylate-hydroxybutyrate cycle, hydroxypropionate-hydroxybutyrate cycle, reductive acetyl-CoA cycle and reductive citric acid cycle) and other important C-cycling genes are listed on the x axis. The intensity of each block corresponds to the percent relative abundance (see legend) of each class (y axis) within a given individual gene/pathway (x axis). Gray squares indicate that no sequences were observed meeting the given criteria. At the top of each column is a bar chart that indicates the total abundance of each gene/pathway in the metagenome. The KEGG accessions for the genes used to annotate the six carbon fixation pathways are provided in Supplementary Table S4.

Figure 2

Relative abundance of single copy protein encoding genes (SC-PEGs). The phylum (y axis) level abundances (x axis) of SC-PEGs are displayed. The percent relative abundance of each phylum is shown at the end of each bar. The following phyla are included in the ‘Other' designation: Korarchaeota, Acidobacteria, Chlamydiae, Fusobacteria, Fibrobacteres, Tenericutes and Cyanobacteria.

Figure 3

Community composition relative to geochemistry. Top: the δ¹⁸O (‰) versus δ²H (‰) of the six Proteobacteria-dominated sites included in Magnabosco et al. (2014) are shown in blue circles. Firmicutes-dominated sites (MP104, Dr938H3, Dr546bh1) are green squares and TT107 is shown as a red star. The black line indicates the GWML (Craig, 1961). Bottom: the δ¹⁸O (‰) versus δ²H (‰) versus Eh (mV) are shown using the same color scheme as (top). *The δ¹⁸O and δ²H for the Firmicutes-dominated Ev818H6 site were not measured but the geochemical similarity with other boreholes located on Ev818 suggests that the water is located far from the GWML (Davidson et al., 2011). The Eh of Ev818H6 was measured to be −128 mV (Davidson et al., 2011).

Figure 4

The normalized abundances of PEGs within the ‘carbon monoxide induced hydrogenase' RAST category are shown. For each PEG (bars), the normalized abundance of that PEG in a given subsurface metagenomes is shown (dots). The size and color of the dot represents the percent relative abundance of Firmicutes identified in the metagenome. The TT107 metagenome (this study) is represented by a large, black circle (45% relative abundance of Firmicutes in the unassembled metagenome).