Choline and N,N-Dimethylethanolamine as Direct Substrates for Methanogens

Andrew J. Watkins (1) , Erwan Roussel (1) , Gordon Webster (1) , R. John Parkes (1) , Henrik Sass (1)

1 School of Earth and Ocean Sciences [Cardiff]

Choline (N,N,N-trimethylethanolamine), which is widely distributed in membrane lipids and is a component of sediment biota, has been shown to be utilized anaerobically by mixed prokaryote cultures to produce methane but not by pure cultures of meth-anogens. Here, we show that five recently isolated Methanococcoides strains from a range of sediments (Aarhus Bay, Denmark; Severn Estuary mudflats at Portishead, United Kingdom; Darwin Mud Volcano, Gulf of Cadiz; Napoli mud volcano, eastern Mediterranean) can directly utilize choline for methanogenesis producing ethanolamine, which is not further metabolized. Di-and monomethylethanolamine are metabolic intermediates that temporarily accumulate. Consistent with this, dimethylethanol-amine was shown to be another new growth substrate, but monomethylethanolamine was not. The specific methanogen inhibi-tor 2-bromoethanesulfonate (BES) inhibited methane production from choline. When choline and trimethylamine are provided together, diauxic growth occurs, with trimethylamine being utilized first, and then after a lag (ϳ7 days) choline is metabolized. Three type strains of Methanococcoides (M. methylutens, M. burtonii, and M. alaskense), in contrast, did not utilize choline. However, two of them (M. methylutens and M. burtonii) did metabolize dimethylethanolamine. These results extend the known substrates that can be directly utilized by some methanogens, giving them the advantage that they would not be reliant on bacterial syntrophs for their substrate supply. I n sediments, terminal oxidation processes show a vertical sequence which follows the decreasing redox potentials of the respective electron acceptors. Aerobic respiration is present at the sediment surface, followed by nitrate and metal reduction, sulfate reduction, and methanogenesis (19). A more positive redox potential allows a more efficient use of the electron donor and consequently a lower threshold concentration for electron donors (7). For example, sulfate reducers can efficiently use hydrogen or acetate at lower concentration and therefore can easily outcompete methanogens (15, 18). There are, however, exceptions to this vertical zonation. Methanogenic archaea have been detected in significant numbers in the upper sediment layers by cultivation-based (8) and molecular methods (26, 27, 34) as well as directly by in situ methanogenic activity measurements (6, 14, 20, 22). The occurrence and activity of methanogens in these layers despite the presence of relatively high sulfate concentrations has been explained by the use of noncompetitive substrates consumed exclusively by methanogens (13, 21, 35). These are typically C 1 compounds, such as methanol and methylated nitrogen (methyl-amine, dimethylamine, and trimethylamine), or sulfur compounds (methanethiol and dimethyl sulfide). Other N-methyl-ated amines bearing a larger side chain, such as ethanolamine in N-methylethanolamine and choline (N,N,N-trimethylethano-lamine), have not been shown to support growth of methanogen pure cultures (9, 11, 29); in contrast, they can be degraded by sulfate reducers (2, 9, 30). This is supported by environmental studies which showed a stimulation of methanogenic activity upon choline or glycine betaine addition but also a simultaneous increase in sulfate reduction, suggesting a two-step degradation pathway involving a sulfate reducer (or fermenter) releasing tri-methylamine that serves as a substrate for methanogenesis (9, 11, 13). In the present study, we demonstrate the methanogenic utilization of choline and N,N-dimethylethanolamine by pure cultures of methanogens affiliated with the genus Methanococcoides. The potential implications of this novel methanogenic pathway will be discussed.

Domaines: Biodiversité et Ecologie; Milieux et Changements globaux; Ecosystèmes; Interactions entre organismes; Bactériologie

Andrew J. Watkins, Erwan Roussel, Gordon Webster, R. John Parkes, Henrik Sass. Choline and N,N-Dimethylethanolamine as Direct Substrates for Methanogens. Applied and Environmental Microbiology, 2012, 78 (23), pp.8298-8303. ⟨10.1128/AEM.01941-12⟩. ⟨hal-01145265⟩