Analysis of Microbial Activity Under a Supercritical CO{sub 2} Atmosphere

Because the extent and impact of microbial activity in deep saline aquifers during geologic sequestration is unknown, the objectives of this proposal were to: (1) characterize the growth requirements and optima of a biofilm-producing supercritical CO{sub 2}-tolerant microbial consortium (labeled MIT0212) isolated from hydrocarbons recovered from the Frio Ridge, TX carbon sequestration site; (2) evaluate the ability of this consortium to grow under simulated reservoir conditions associated with supercritical CO{sub 2} injection; (3) isolate and characterize individual microbial strains from this consortium; and (4) investigate the mechanisms of supercritical CO{sub 2} tolerance in isolated strains and the consortium through genome-enabled studies. Molecular analysis of genetic diversity in the consortium MIT0212 revealed a predominance of sequences closely related to species of the spore-forming genus Bacillus. Strain MIT0214 was isolated from this consortium and characterized by physiological profiling and genomic analysis. We have shown that the strain MIT0214 is an aerobic spore-former and capable of facultative anaerobic growth under both reducing N{sub 2} and CO{sub 2} atmospheres by fermentation and possibly anaerobic respiration. Strain MIT0214 is best adapted to anaerobic growth at pressures of 1 atm but is able to growth at elevated pressures After 1 week growth was observed at pressures as high as 27 atm (N{sub 2}) or 9 atm (CO{sub 2}) and after 26-30 days growth can be observed under supercritical CO{sub 2}. In addition, we have determined that spores of strain B. cereus MIT0214 are tolerant of both direct and indirect exposure to supercritical CO{sub 2}. Additional physiological characterization under aerobic conditions have revealed MIT0214 is able to grow from temperature of 21 to 45 °C and salinities 0.01 to 40 g/L NaCl with optimal growth occurring at 30°C and from 1 - 5 g NaCl/L. The genome sequence of B. cereus MIT0214 shared 89 to 91% of genes with other genome-sequenced strains with 93.3 to 97.8% nucleotide identity among shared genes. Comparison of the sequence of MIT0214 or a B. cereus strain isolated from an oil well in China to B. cereus isolates from surface environments revealed a higher proportion of genes involved in Cell wall and capsule biosynthesis and metabolism, metabolism of aromatic compounds, and stress response. Since Bacillus species, including B. cereus strains, have commonly been recovered from other “extreme” environments including the deep subsurface – the scCO{sub 2} tolerance of spores and growth under high pCO{sub 2} conditions is consistent with persistence in a subsurface environment after CO{sub 2} injection