Hallelujah! The in situ nitrogen cycle within the subterranean crust

Abstract:

Subsurface microorganisms need C and N to maintain, grow and reproduce. However, the processing of N at the terrestrial deep subsurface has been understudied. Based on previous data generated from fracture water samples collected from mines in South Africa, including (i) stable isotope analyses of bulk N, N₂, NO₃^- and NH₃/NH⁴⁺, (ii) cloning and sequencing of NifH genes in DNA and RNA samples, and (iii) detection of Nar, Nif and NPD genes in several metagenomes, the nitrogen cycle occurred at the terrestrial deep subsurface sites has been proposed recently. We carried out an in-depth investigation of fracture water samples from a borehole located at 1.34 km depth in Beatrix Au mine where N and O isotope analyses of NO₃^- indicated enriched d¹⁵N and d¹⁸O values, consistent with fractionation during nitrate reduction. We identified the microorganisms responsible for the N-cycling and characterized the genes and proteins that were expressed. Metagenomic data provided a critical foundation for annotating the transcripts and proteomes. Preliminary results revealed the presence of transcripts and enzymes for denitrification (nitrate, nitrite and nitrous-oxide reduction) and N₂ fixation. b-Proteobacteria is the predominant group of denitrifying bacteria, such as Cadidatus Accumulibacter, Dechlorosoma, Thiobacillus, Azoarcus and Sulfuricella, as shown in the transcriptomic and proteomic data. Transcripts homologous to nitrogenase and its assembly proteins belong to b-Proteobacteria and the firmicutes Ca. Desulforudis audaxviator.