Peatlands are a major natural source of atmospheric methane (CH4). Emissions from Sphagnum-dominated mires are lower than those measured from other mire types. This observation may partly be due to methanotrophic (i.e., methane-consuming) bacteria associated with Sphagnum. Twenty-three of the 41 Sphagnum species in Finland can be found in the peatland at Lakkasuo. To better understand the Sphagnum-methanotroph system, we tested the following hypotheses: (1) all these Sphagnum species support methanotrophic bacteria; (2) water level is the key environmental determinant for differences in methanotrophy across habitats; (3) under dry conditions, Sphagnum species will not host methanotrophic bacteria; and (4) methanotrophs can move from one Sphagnum shoot to another in an aquatic environment. To address hypotheses 1 and 2, we measured the water table and CH4 oxidation for all Sphagnum species at Lakkasuo in 1-5 replicates for each species. Using this systematic approach, we included Sphagnum spp. with narrow and broad ecological tolerances. To estimate the potential contribution of CH4 to moss carbon, we measured the uptake of delta13C supplied as CH4 or as carbon dioxide dissolved in water. To test hypotheses 2-4, we transplanted inactive moss patches to active sites and measured their methanotroph communities before and after transplantation. All 23 Sphagnum species showed methanotrophic activity, confirming hypothesis 1. We found that water level was the key environmental factor regulating methanotrophy in Sphagnum (hypothesis 2). Mosses that previously exhibited no CH4 oxidation became active when transplanted to an environment in which the microbes in the control mosses were actively oxidizing CH4 (hypothesis 4). Newly active transplants possessed a Methylocystis signature also found in the control Sphagnum spp. Inactive transplants also supported a Methylocystis signature in common with active transplants and control mosses, which rejects hypothesis 3. Our results imply a loose symbiosis between Sphagnum spp. and methanotrophic bacteria that accounts for potentially 10-30% of Sphagnum carbon.

The main objectives of this study were to uncover the pathways used for methanogenesis in three different boreal peatland ecosystems and to describe the methanogenic populations involved. The mesotrophic fen had the lowest proportion of CH 4 produced from H 2 -CO 2 . The oligotrophic fen was the most hydrogenotrophic, followed by the ombrotrophic bog. Each site was characterized by a specific group of methanogenic sequences belonging to Methanosaeta spp. (mesotrophic fen), rice cluster-I (oligotrophic fen), and fen cluster (ombrotrophic bog).Northern peatlands are important emitters of the green house gas methane (CH 4 ) produced by methanogenic archaea (3,18,30). Methanogens utilize a limited number of substrates, the most important of which are acetate and H 2 -CO 2 (38). In peatlands, H 2 -CO 2 -dependent methanogenesis is thought to be the main pathway for CH 4 production (20,21,26,37), but in some minerotrophic peatlands (fens), acetoclastic methanogenesis is often predominant in upper peat layers (4,23,31). The diversity of methanogenic communities of fen (15, 17) and bog (2, 16, 35) ecosystems has recently been described, but data on the combined investigation of methanogenic pathways and methanogen populations are scarce. To the best of our knowledge, community studies have never been associated with the detection of methanogenic pathways in fen ecosystems, and only one study of an acidic bog ecosystem has been published (20). The aim of our study was to determine the precursors used for methanogenesis in three peatland ecosystems (a mesotrophic fen, an oligotrophic fen, and an ombrotrophic bog) and to describe the diversity of methane-producing archaea by using molecular methods that target the functional methyl coenzyme M reductase gene (mcrA).Replicate samples from depths with the highest potential CH 4 production rates were taken from a mesotrophic fen, an oligotrophic fen, and an ombrotrophic bog at the Lakkasuo mire complex in central Finland (61°48ЈN, 24°19ЈE) in August 2003. The fraction of CH 4 produced from H 2 -CO 2 was estimated at each site by a tracer experiment (8, 11) and by the inhibition of acetoclastic methanogenesis with CH 3 F (methyl fluoride) (6,14,22). The Gibbs free energy (⌬G) of H 2 -dependent methanogenesis was calculated (10, 12), and potential CH 4 production was measured as described earlier (17). Fatty acids and alcohols dissolved in the pore water were analyzed by high-pressure liquid chromatography (25). All experiments were conducted on triplicate samples from each peat ecosystem. DNA was extracted directly from peat samples (17), and a portion of the methanogen-specific mcrA was amplified with the primer pair ML (28). Clone libraries were subjected to restriction fragment length polymorphism (RFLP) analysis with the restriction enzymes MspI and TaqI, and representatives of the biggest RFLP groups were sequenced for phylogenetic analysis.The contributions of H 2 -CO 2 -dependent methanogenesis to total CH 4 production varied clearly among the three peatland ecosystems (...

Mires forming an ecohydrological gradient from nutrient-rich, groundwater-fed mesotrophic and oligotrophic fens to a nutrient-poor ombrotrophic bog were studied by comparing potential methane (CH(4)) production and methanogenic microbial communities. Methane production was measured from different depths of anoxic peat and methanogen communities were detected by detailed restriction fragment length polymorphism (RFLP) analysis of clone libraries, sequencing and phylogenetic analysis. Potential CH(4) production changed along the ecohydrological gradient with the fens displaying much higher production than the ombrotrophic bog. Methanogen diversity also decreased along the gradient. The two fens had very similar diversity of methanogenic methyl-coenzyme M reductase gene (mcrA), but in the upper layer of the bog the methanogen diversity was strikingly lower, and only one type of mcrA sequence was retrieved. It was related to the Fen cluster, a group of novel methanogenic sequences found earlier in Finnish mires. Bacterial 16S rDNA sequences from the fens fell into at least nine phyla, but only four phyla were retrieved from the bog. The most common bacterial groups were Deltaproteobacteria, Verrucomicrobia and Acidobacteria.

Methane (CH 4 ) emissions from boreal wetlands show considerable seasonal variation, including small winter emissions. We addressed the seasonality of CH 4 -producing microbes by comparing archaeal communities and the rates and temperature response of CH 4 production in a boreal fen at three key phases of growing season and in winter. Archaeal community analysis by terminal restriction fragment length polymorphism and cloning of 16S ribosomal DNA and reversetranscribed RNA revealed slight community shifts with season. The main archaeal groups remained the same throughout the year and were Methanosarcinaceae, Rice cluster II and Methanomicrobiales-associated Fen cluster. These methanogens and the crenarchaeal groups 1.1c and 1.3 were detected from DNA and RNA, but the family Methanosaetaceae was detected only from RNA. Differences between DNA-and RNA-based results suggested higher stability of DNA-derived communities and better representation of the active CH 4 producers in RNA. Methane production potential, measured as formation of CH 4 in anoxic laboratory incubations, showed prominent seasonality. The potential was strikingly highest in winter, possibly due to accumulation of methanogenic substrates, and maximal CH 4 production was observed at ca. 30 1C. Archaeal community size, determined by quantitative PCR, remained similar from winter to summer. Low production potential in late summer after a water level draw-down suggested diminished activity due to oxygen exposure. Our results indicated that archaeal community composition and size in the boreal fen varied only slightly despite the large fluctuations of methanogenic potential. Detection of mRNA of the methanogenic mcrA gene confirmed activity of methanogens in winter, accounting for previously reported winter CH 4 emissions. The ISME Journal (2008Journal ( ) 2, 1157Journal ( -1168 doi:10.1038/ismej.2008 published online 24 July 2008 Subject Category: microbial ecology and functional diversity of natural habitats

Wetlands, including peatlands, are the main source of natural methane emission. Well-defined fen microsites have different methane emissions rates, but it is not known whether the methane-producing Archaea communities vary at these sites. Possible horizontal variations of communities, in a natural oligotrophic fen, were analysed by characterizing the methanogens from two well-defined microsites: Eriophorum lawn and Hummock. Community structures were studied at two different layers of the fen, showing, respectively, high and low methane production. The structure of methanogen populations was determined using molecular techniques targeting the 16SrRNA gene and combined denaturing gradient gel electrophoresis (DGGE) and restriction fragment length polymorphism (RFLP) analysis. Results subjected to non-metric multidimensional scaling (MDS), diversity indices calculation and phylogenetic analysis revealed that upper layer communities changed with site while deeper layer communities remained the same. Phylogenetic analyses revealed six different clusters of sequences grouping with only two known orders of methanogens. Upper layers of Hummock were dominated by sequences clustering with members of Methanomicrobiales and sequences dominating the upper part of the Eriophorum lawn were related to members of the order Methanosarcinales. Novel methanogenic sequences were found at both sites at both depths. Vegetation characterizing the microsites probably influences the microbial communities in the layers of the fen where methane is produced.