In Situ Stable Isotope Probing

Microorganisms living in anoxic rice soils contribute 10 to 25 % of global meth-ane emissions. The most important carbon source for CH4 production is plant-derived carbon that enters soil as root exudates and debris. Pulse labeling of rice plants with 13CO2 resulted in incorporation of 13C into the ribosomal RNA of Rice Cluster I Archaea in the soil, indicating that this archaeal group plays a key role in CH4 production from plant-derived carbon. This group of micro-organisms has not yet been isolated but appears to be of global environmental importance. The translocation of plant photosynthates below ground and their subsequent decom-position by rhizospheric microorganisms are key to the terrestrial ecosystem carbon budget (1). It has been shown that 30 to 60 % of net photosynthesized carbon is allocated to roots, and as much as 40 to 90 % of this fraction enters soil in the forms of root exudates, sloughed-off cells, and decaying roots (2). In wetland soils and rice paddies, the below-ground carbon flow provides a major carbon source for meth-ane production (3, 4). However, little is known about the microbiota that are involved in the rhizosphere carbon cycle, and the soil micro-biota is considered a Bblack box [ in studies of soil carbon dynamics (5). A combination of stable isotope probing (SIP) and biomarker-based fingerprinting can be a powerful approach to investigate micro-bial species and in situ activity (6, 7). Bio-markers that have been used with SIP include nucleic acids (7–11) and phospholipid fatty acids (6, 12–14). Nucleic acids are particularly useful because they provide specific taxonomic information (15). Here we report on the appli-cation of RNA SIP in an intact rice-soil system to identify the active methane producers in the rhizosphere. Rice fields represent one of the most im-portant individual sources of the greenhouse gas CH 4 (16, 17). Below-ground carbon flo