Little research has been conducted on microbial diversity deep under the Earth’s surface. In this study, the microbial communities of three deep terrestrial subsurface aquifers were investigated. Temporal community data over 6 years revealed that the phylogenetic structure and community dynamics were highly dependent on the degree of isolation from the earth surface biomes. The microbial community at the shallow site was the most dynamic and was dominated by the sulfur-oxidizing genera Sulfurovum or Sulfurimonas at all-time points. The microbial community in the meteoric water filled intermediate aquifer (water turnover approximately every 5 years) was less variable and was dominated by candidate phylum OD1. Metagenomic analysis of this water demonstrated the occurrence of key genes for nitrogen and carbon fixation, sulfate reduction, sulfide oxidation and fermentation. The deepest water mass (5000 year old waters) had the lowest taxon richness and surprisingly contained Cyanobacteria. The high relative abundance of phylogenetic groups associated with nitrogen and sulfur cycling, as well as fermentation implied that these processes were important in these systems. We conclude that the microbial community patterns appear to be shaped by the availability of energy and nutrient sources via connectivity to the surface or from deep geological processes.

The deep biosphere is one of the least understood ecosystems on Earth. Although most microbiological studies in this system have focused on prokaryotes and neglected microeukaryotes, recent discoveries have revealed existence of fossil and active fungi in marine sediments and sub-seafloor basalts, with proposed importance for the subsurface energy cycle. However, studies of fungi in deep continental crystalline rocks are surprisingly few. Consequently, the characteristics and processes of fungi and fungus-prokaryote interactions in this vast environment remain enigmatic. Here we report the first findings of partly organically preserved and partly mineralized fungi at great depth in fractured crystalline rock (−740 m). Based on environmental parameters and mineralogy the fungi are interpreted as anaerobic. Synchrotron-based techniques and stable isotope microanalysis confirm a coupling between the fungi and sulfate reducing bacteria. The cryptoendolithic fungi have significantly weathered neighboring zeolite crystals and thus have implications for storage of toxic wastes using zeolite barriers.

Precipitation of exceptionally ¹³C-depleted authigenic carbonate is a result of, and thus a tracer for, sulphate-dependent anaerobic methane oxidation, particularly in marine sediments. Although these carbonates typically are less depleted in ¹³C than in the source methane, because of incorporation of C also from other sources, they are far more depleted in ¹³C (δ¹³C as light as −69‰ V-PDB) than in carbonates formed where no methane is involved. Here we show that oxidation of biogenic methane in carbon-poor deep groundwater in fractured granitoid rocks has resulted in fracture-wall precipitation of the most extremely ¹³C-depleted carbonates ever reported, δ¹³C down to −125‰ V-PDB. A microbial consortium of sulphate reducers and methane oxidizers has been involved, as revealed by biomarker signatures in the carbonates and S-isotope compositions of co-genetic sulphide. Methane formed at shallow depths has been oxidized at several hundred metres depth at the transition to a deep-seated sulphate-rich saline water. This process is so far an unrecognized terrestrial sink of methane.

Oxidation and reduction of iron can occur through abiotic (chemical) and biotic (microbial) processes. Abiotic iron oxidation is a function of pH and O₂ concentration. Biotic iron oxidation is carried out by a diverse group of bacteria, using O₂ or NO₃ as terminal electron acceptors. At circumneutral pH, both processes occur at similar rates and compete with each other. Abiotic iron reduction is catalyzed by iron-sulfur minerals or different types of organic compounds, whereas biotic iron reduction is carried out by a diverse group of microorganisms, often using chemical agents to dissolve solid iron minerals. We used iron oxidizing microbial mats to assess the potential impact of microbial activity on the deposition of banded iron formations (BIF). The mats were collected during several years from experimental tanks connected to groundwater aquifers with different Fe²⁺ concentrations. To separate between biotic and abiotic iron oxidation, live and killed mats were incubated with ⁵⁷Fe²⁺. Separate analyses of the water and solid phase revealed that the iron oxidation and reduction rates per mL of solid matter (biomass and iron precipitates) were 0.4-73 mmol L⁻¹ d⁻¹ and 30–280 mmol L⁻¹ d⁻¹, respectively. No significant differences in iron oxidation rates were observed between the live and killed samples. The iron reduction rates, however, were higher in the live samples in mats from 3 out of 4 environments. We suggest that in natural systems, in the presence of organic matter, biotic and abiotic iron oxidation and reduction are not separable processes. Fe²⁺ will be biotically and abiotically oxidized as well as bind to exposed charged groups of organic substances. Either way, this iron may serve as a nucleation matrix for further abiotic iron precipitation. The oxidized iron is then susceptible to iron reduction, which can likewise be a direct metabolic or an abiotic process. Nevertheless, it is important to note the significance of organic matter, since both the abiotic oxidation and reduction of iron are often mediated by substrates of biological origin.

Fractured rocks of impact craters may be suitable hosts for deep microbial communities on Earth and potentially other terrestrial planets, yet direct evidence remains elusive. Here, we present a study of the largest crater of Europe, the Devonian Siljan structure, showing that impact structures can be important unexplored hosts for long-term deep microbial activity. Secondary carbonate minerals dated to 80 ± 5 to 22 ± 3 million years, and thus postdating the impact by more than 300 million years, have isotopic signatures revealing both microbial methanogenesis and anaerobic oxidation of methane in the bedrock. Hydrocarbons mobilized from matured shale source rocks were utilized by subsurface microorganisms, leading to accumulation of microbial methane mixed with a thermogenic and possibly a minor abiotic gas fraction beneath a sedimentary cap rock at the crater rim. These new insights into crater hosted gas accumulation and microbial activity have implications for understanding the astrobiological consequences of impacts.