Diversity and Possible Activity of Microorganisms in Underground Gas Storage Aquifers

— Underground natural gas storage facilities (UGS) have been recently proposed as sites to store “green” gas containing biogas, synthetic methane, and molecular hydrogen. The composition of the UGS microbial communities and the effect of H 2 on these communities are poorly understood. This work deals with determination of microbial diversity in the samples of reservoir water from the underground horizons of the Shchelkovo, Kasimov, and Kaluga UGS. Groundwater is an anaerobic habitat containing acetic and other lower fatty acids, methanol, and dissolved gases that may serve as substrates for microorganisms. Low abundance of cultivated aerobic organotrophic bacteria and anaerobic fermenting, sulfate-reducing, and methanogenic microorganisms in the studied samples was shown. High-throughput sequencing of the V4 region of the 16S rRNA gene revealed the presence of Firmicutes (2.4–53.6%), Bacteroidetes (1.0–39.7%), Alphaproteobacteria (0.9–9.6%), Actinobacteria (0.1–1.7%), Desulfobacteria (0.1–1.6%), Verrucomicrobia (0–3.4%), and Planctomycetes (0–1.3%) in the studied microbial communities. The share of archaeal sequences in the libraries did not exceed 1.5%. In the water sample from the Kaluga UGS, members of the genus Marinobacter predominated; methanogens of the genera Methanosphaera , Methanolobus , and Methanobrevibacter were found among the minor components. Methylotrophic bacteria of the genera Methylococcus and Methylobacterium‒Methylorubrum and anaerobic fermenting bacteria of the genus Bacteroides predominated in the reservoir water from the Shchelkovo and Kasimov UGSs. Using the iVikodak program, the potential ability of microbial communities to use methane, methanol, benzoate, and polycyclic aromatic hydrocarbons, as well as to participate in the transformations of sulfur and nitrogen compounds, was shown. In the underground communities, bacteria and archaea were found, potentially capable of using H 2 in their energy metabolism, including the processes of sulfate reduction, methanogenesis and acetogenesis. These results indicate that microbiological and geochemical monitoring is required during the operation of UGS, especially during the injection of hydrogen.