The effect of bituminous coal on methanogenic mixed cultures and pure cultures of Methanococcus and Methanosarcina

In numerous global coal-bearing basins, methane, possessing a biogenic stable isotope composition, is spatially and temporally associated with groundwater recharge. However, beyond groundwater inoculating the subsurface with microorganisms, the biological mechanisms that control the distribution of biogenic methane are poorly understood. In this study, we examined the interactions between bituminous coal and a) methanogenic microbial communities sourced from the goaf of a Bowen Basin underground coal mine and b) pure cultures of methanogens. When coal mine microbial consortium was amended with acetate and a low concentration of coal (1 g in 25 mL medium), methane production was stimulated compared to the addition of acetate alone, though the presence of coal did not affect the methane production from H2/CO2. To test whether methanogens benefited directly from the addition of coal, 1 g of either quartz sand or bituminous coal was added to a pure culture of Methanococcus maripaludis, a hydrogenotrophic methanogen isolated from the coal mine and grown on H2/CO2, and to a pure culture of Methanosarcina barkeri, an acetoclastic methanogen grown on acetate. In this experiment coal was not included as an energy source for microbial growth but to test interactions between coal and methanogens. The presence of coal in the medium did not affect methane production by Ms. barkeri but slightly inhibited methane production by M. maripaludis at the start of the growth phase. Scanning electron microscopy revealed that M. maripaludis cells were attached to both sand grains and coal particles, with preferential attachment to rough surfaces, such as cracks within broken coal pieces and clay-rich areas of coal. When the experiment with M. maripaludis and Ms. barkeri was repeated with a 1:1 volumetric ratio of bituminous coal to medium, which more accurately reflects the environmental conditions of a coal seam, methane production by M. maripaludis and Ms. barkeri was completely inhibited. In addition, M. maripaludis cell numbers declined after inoculation. This suggests that at a high coal: fluid ratio, methanogenesis is inhibited by some component of the coal, e.g., bitumens in the coal or coal-sourced hydrocarbons dissolved into the medium. Based on these laboratory results, we propose that in coal seams, the dilution of inhibitory compounds may be one mechanism by which groundwater recharge promotes biogenic methane production.