Methanotrophic activity in Carboniferous coalbed rocks

Abstract

Carboniferous coalbed rocks originating from the Lublin Coal Basin (South-East Poland) were investigated for the presence and activity of aerobic methanotrophic bacteria. Laboratory studies of samples collected from 914 to 1004m below earth's surface and performed at 30°C, 100% water capacity (WHC) and 10% v/v headspace CH4 revealed microbial methane oxidation which ranged from 0.13 to 0.75μM CH4 g−1day−1. Methanotrophic activity (MA) was also investigated as a function of temperature (5–30°C), moisture content (25–200% WHC) and substrate concentration (1–30% v/v CH4). The highest MA was recorded at 30°C, 100% WHC and 20% CH4 v/v, indicating the mesophilic and microaerophilic character of methanotrophic bacteria (MB) present in the coalbed. Lack of detectable MA at 5 and 10°C shows narrow temperature tolerance of MB, which may result from a long-term isolation of the bacterial habitat and stability of thermal conditions in the coalbed.Cryo-SEM revealed presence in the investigated rocks of bacteria (c.a. 2–2.5μm long and 0.5–1μm wide) and spore-like structures (0.5–1μm in diameter). These observations were confirmed by fluorescence in situ hybridization with probes Mg705 (5′fluoresceine), Mg84 (5′Cy3) and Ma450 (5′Cy5), which showed that enrichments in NMS medium contain types I and II MB of similar size. Identification performed with the use of DNA amplification of 16S rRNA targeted group specific primers confirmed the presence of bacteria belonging to Methylosinus, Methylomicrobium, Methylocystis and Methylocaldum. Here we show that coalbed rocks constitute a habitat for diverse MB.