Real time monitoring of hydrogenotrophic methanogenesis under deep saline aquifers conditions

To investigate the microbial response to deep underground gas injection, specifically CO2 and H2, a new optically transparent high-pressure reactor was developed to monitor autotrophic microbial growth via in situ and ex situ characterization techniques. The main advantages rely on avoiding any decompression phases during the entire process, thanks to direct optical access. Here, we monitored the growth of the model methanogenic strain Methanothermococcus thermolithotrophicus by applying different H2/CO2 partial pressures at a total pressure of 100 bar, which is representative of the deep underground storage environment. These reactors can be used to investigate deep microbial strains under pressure conditions close to their natural environments, eliminating decompression biases. Additionally, we measured the methane production of the strain at the end of the incubation, with or without stirring. The results showed an increase in methane production with increasing CO2 and H2 partial pressures, up to p(H2/CO2, 80/20 mol%) = 15 bar for unstirred conditions and p(H2/CO2, 80/20 mol%) = 30 bar for stirred conditions, respectively. This difference in behavior depending on stirring conditions was clearly observed through biofilm formation in the absence of stirring, favoring methane production and extending the range of H2 / CO2 partial pressures under which the strain can grow.