To shake or not to shake: 13C-based evidence on anaerobic methane oxidation in paddy soil

The anaerobic oxidation of methane (AOM) removes most of the biologically produced methane (CH4) from marine ecosystems before it enters the atmosphere and thus mitigates greenhouse gas emissions. As compared to marine environments, surprisingly little is known about the role of AOM in terrestrial ecosystems. Particularly, how AOM controls the CH4 budget of paddy soils is unexplored, partly reflecting analytical difficulties in analyzing CH4 turnover. To date, the most commonly used method to study AOM in soils is in vitro incubation of microcosms with CH4 injection into the headspace with/without shaking of slurry. Shaking, however, introduces various errors and disturbances. Here we measured AOM in rice paddy soil using a new alternative approach that introduced 13C-labelled CH4 directly into soil slurry via a silicone tube without shaking. The results were compared to those obtained by the classical approaches (i.e., with and without tubes and/or shaking). In all batches, 13C enrichment of CO2 after 13CH4 injection clearly confirmed the occurrence of AOM in paddy soil. The cumulative AOM during 84 days reached 0.16–0.24 μg C g−1 dry soil without shaking, but it was 33–80% lower with shaking. Unexpectedly, the effect of silicone tubes on AOM was insignificant either with or without shaking, suggesting that the CH4 concentration in water (slurry) was not the main limiting factor for AOM. Without shaking, the controls without CH4 addition revealed a steady increase of CH4 in the headspace/tube, whereas the CH4 concentration in jars with shaking was constantly low during 59 days. This suggests that shaking inhibited methanogenesis. There was a strong linear correlation between the amount of CH4 oxidized and CH4 produced with shaking (R 2 = 0.91), whereas without shaking this relationship followed a power growth regression. Based on the current and reported AOM rates, rough upscale to paddy soils in China showed recycling of ca. 2.0 Tg C of CH4 each year, making AOM a crucial terrestrial CH4 sink.