Characterization of a halotolerant acetoclastic methanogen highly enriched from marine sediment and its application in removal of acetate.

A marine sediment collected from Hiroshima Bay was cultured in artificial seawater, containing 0.51 M NaCl and 60 mM acetate and was found to exhibit active methane production at 37°C. Following four successive serial dilutions of cultures in medium containing 0.51 M NaCl, 60 mM acetate, and antibiotics, the well-acclimated methanogen was found to exhibit growth over a range of NaCl concentration (between 0 M and 2.06 M). The specific growth rates of the highly enriched methanogen, termed strain HA, in the absence of NaCl and in the presence of 1.54 M NaCl were estimated to be 0.037 h(-1) and 0.027 h(-1), respectively. The pH and temperature for optimum growth were determined to be 7.0-8.8 and 37°C, respectively. Although cells that had morphology similar to Methanosaeta sp. became dominant in the culture, methane production was still detected in the medium containing 0.51 M NaCl and other substrates such as methanol, formate, and methylamine, indicating contamination with other methanogens. The phylogenetic tree based on 16S rRNA gene sequences revealed that the strain HA was closely related to Methanosaeta harundinacea 6Ac and 8Ac(T), with sequence similarity of 98% and 97%, respectively. The continuous removal of acetate with upflow anaerobic filter reactor for industrial use of strain HA determined a methane production rate of 70 mM/d under condition of 0.51 M NaCl and successful methane production even under 1.54 M NaCl.