Methane‐derived carbon flow through microbial communities in arctic lake sediments

Aerobic methane (CH₄) oxidation mitigates CH₄ release and is a significant pathway for carbon and energy flow into aquatic food webs. Arctic lakes are responsible for an increasing proportion of global CH₄ emissions, but CH₄ assimilation into the aquatic food web in arctic lakes is poorly understood. Using stable isotope probing (SIP) based on phospholipid fatty acids (PLFA‐SIP) and DNA (DNA‐SIP), we tracked carbon flow quantitatively from CH₄ into sediment microorganisms from an arctic lake with an active CH₄ seepage. When 0.025 mmol CH₄ g^?1 wet sediment was oxidized, approximately 15.8–32.8% of the CH₄‐derived carbon had been incorporated into microorganisms. This CH₄‐derived carbon equated to up to 5.7% of total primary production estimates for Alaskan arctic lakes. Type I methanotrophs, including Methylomonas, Methylobacter and unclassified Methylococcaceae, were most active at CH₄ oxidation in this arctic lake. With increasing distance from the active CH₄ seepage, a greater diversity of bacteria incorporated CH₄‐derived carbon. Actinomycetes were the most quantitatively important microorganisms involved in secondary feeding on CH₄‐derived carbon. These results showed that CH₄ flows through methanotrophs into the broader microbial community and that type I methanotrophs, methylotrophs and actinomycetes are important organisms involved in using CH₄‐derived carbon in arctic freshwater ecosystems.     

Does dissolved organic carbon regulate biological methane oxidation in semiarid soils?

In humid ecosystems, the rate of methane (CH₄) oxidation by soil‐dwelling methane‐oxidizing bacteria (MOB) is controlled by soil texture and soil water holding capacity, both of which limit the diffusion of atmospheric CH₄ into the soil. However, it remains unclear whether these same mechanisms control CH₄ oxidation in more arid soils. This study was designed to measure the proximate controls of potential CH₄ oxidation in semiarid soils during different seasons. Using a unique and well‐constrained 3‐million‐year‐old semiarid substrate age gradient, we were able to hold state factors constant while exploring the relationship between seasonal potential CH₄ oxidation rates and soil texture, soil water holding capacity, and dissolved organic carbon (DOC). We measured unexpectedly higher rates of potential CH₄ oxidation in the wet season than the dry season. Although other studies have attributed low CH₄ oxidation rates in dry soils to desiccation of MOB, we present several lines of evidence that this may be inaccurate. We found that soil DOC concentration explained CH₄ oxidation rates better than soil physical factors that regulate the diffusion of CH₄ from the atmosphere into the soil. We show evidence that MOB facultatively incorporated isotopically labeled glucose into their cells, and MOB utilized glucose in a pattern among our study sites that was similar to wet‐season CH₄ oxidation rates. This evidence suggests that DOC, which is utilized by MOB in other environments with varying effects on CH₄ oxidation rates, may be an important regulator of CH₄ oxidation rates in semiarid soils. Our collective understanding of the facultative use of DOC by MOB is still in its infancy, but our results suggest it may be an important factor controlling CH₄ oxidation in soils from dry ecosystems.     

Activity, distribution, and abundance of methane-oxidizing bacteria in the near surface soils of onshore oil and gas fields

Methane-oxidizing bacteria (MOB) have long been used as an important biological indicator for oil and gas prospecting, but the ecological characteristics of MOB in hydrocarbon microseep systems are still poorly understood. In this study, the activity, distribution, and abundance of aerobic methanotrophic communities in the surface soils underlying an oil and gas field were investigated using biogeochemical and molecular ecological techniques. Measurements of potential methane oxidation rates and pmoA gene copy numbers showed that soils inside an oil and gas field are hot spots of methane oxidation and MOB abundance. Correspondingly, terminal restriction fragment length polymorphism analyses in combination with cloning and sequencing of pmoA genes also revealed considerable differences in the methanotrophic community composition between oil and gas fields and the surrounding soils. Principal component analysis ordination furthermore indicated a coincidence between elevated CH₄ oxidation activity and the methanotrophic community structure with type I methanotrophic Methylococcus and Methylobacter, in particular, as indicator species of oil and gas fields. Collectively, our results show that trace methane migrated from oil and gas reservoirs can considerably influence not only the quantity but also the structure of the methanotrophic community.     

Diversity of methanotrophs in urea-fertilized tropical rice agroecosystem

Laboratory experiments were conducted to study the population size, diversity and methane oxidation potential of methanotrophs in tropical rice agroecosystem under the influence of N-fertilizer. Results indicate that the diversity of methane oxidizing bacteria (MOB) is altered in fertilizer treated soils compared to untreated control. Nevertheless, Type I MOB still dominated in the fertilized soils whereas the diversity of Type II methanotrophs decreases. Control soils have higher MOB population and CH₄ oxidation capacity than fertilized soils. Rhizospheric soil is more populated than non-rhizospheric soil in both unfertilized and fertilized conditions. Variation in K_m and V_max of methane oxidation in soils appears to be due to variation in methanotrophic community. Experimental results indicate that methanotrophic community differs both quantitatively and qualitatively in unfertilized and fertilized soils.     

Methane-Oxidizing Bacteria in a Finnish Raised Mire Complex: Effects of Site Fertility and Drainage

Methane-oxidizing bacteria (MOB) are the only biological sinks for methane (CH₄). Drainage of peatlands is known to decrease overall CH₄ emission, but the effect on MOB is unknown. The objective of this work was to characterize the MOB community and activity in two ecohydrologically different pristine peatland ecosystems, a fen and a bog, and their counterparts that were drained in 1961. Oligotrophic fens are groundwater-fed peatlands, but ombrotrophic bogs receive additional water and nutrients only from rainwater. The sites were sampled in August 2003 down to 10 cm below the water table (WT), and cores were divided into 10-cm subsamples. CH₄ oxidation was measured by gas chromatography (GC) to characterize MOB activity. The MOB community structure was characterized by polymerase chain reaction–denaturing gradient gel electrophoresis (DGGE) and sequencing methods using partial pmoA and mmoX genes. The highest CH₄ oxidation rates were measured from the subsamples 20–30 and 30–40 cm above WT at the pristine oligotrophic fen (12.7 and 10.5 μmol CH₄ dm⁻³ h⁻¹, respectively), but the rates decreased to almost zero in the vicinity of WT. In the pristine ombrotrophic bog, the highest oxidation rate at 0–10 cm was lower than in the fen (8.10 μmol CH₄ dm⁻³ h⁻¹), but in contrast to the fen, oxidation rates of 4.5 μmol CH₄ dm⁻³ h⁻¹ were observed at WT and 10 cm below WT. Drainage reduced the CH₄ oxidation rates to maximum values of 1.67 and 5.77 μmol CH₄ dm⁻³ h⁻¹ at 30–40 and 20–30 cm of the fen and bog site, respectively. From the total of 13 pmoA-derived DGGE bands found in the study, 11, 3, 6, and 2 were observed in the pristine fen and bog and their drained counterparts, respectively. According to the nonmetric multidimensional scaling of the DGGE banding pattern, the MOB community of the pristine fen differed from the other sites. The majority of partial pmoA sequences belonged to type I MOB, whereas the partial mmoX bands that were observed only in the bog sites formed a distinct group relating more to type II MOB. This study indicates that fen and bog ecosystems differ in MOB activity and community structure, and both these factors are affected by drainage.     

Identification of functionally active aerobic methanotrophs in sediments from an arctic lake using stable isotope probing

Arctic lakes are a significant source of the greenhouse gas methane (CH₄), but the role that methane oxidizing bacteria (methanotrophs) play in limiting the overall CH₄ flux is poorly understood. Here, we used stable isotope probing (SIP) techniques to identify the metabolically active aerobic methanotrophs in upper sediments (0–1 cm) from an arctic lake in northern Alaska sampled during ice‐free summer conditions. The highest CH₄ oxidation potential was observed in the upper sediment (0–1 cm depth) with 1.59 µmol g wet weight^?1 day^?1 compared with the deeper sediment samples (1–3 cm, 3–5 cm and 5–10 cm), which exhibited CH₄ oxidation potentials below 0.4 µmol g wet weight^?1 day^?1. Both type I and type II methanotrophs were directly detected in the upper sediment total communities using targeted primer sets based on 16S rRNA genes. Sequencing of 16S rRNA genes and functional genes (pmoA and mxaF) in the ¹³C‐DNA from the upper sediment indicated that type I methanotrophs, mainly Methylobacter, Methylosoma, Methylomonas and Methylovulum miyakonense, dominated the assimilation of CH₄. Methylotrophs, including the genera Methylophilus and/or Methylotenera, were also abundant in the ¹³C‐DNA. Our results show that a diverse microbial consortium acquired carbon from CH₄ in the sediments of this arctic lake.     

CH4 production and oxidation processes in a boreal fen ecosystem after long‐term water table drawdown

Fens, which extend over vast areas in the Northern hemisphere, are sources of the greenhouse gas CH₄. Climate change scenarios predict a lowering water table (WT) in mires. To study the effect of WT drawdown on CH₄ dynamics in a fen ecosystem, we took advantage of a WT drawdown gradient near a ground water extraction plant. Methane fluxes and CH₄ production and oxidation potentials were related to microbial communities responsible for the processes in four mire locations (wet, semiwet, semidry, and dry). Principal component analyses performed on the vegetation, pH, CH₄, and WT results clearly separated the four sampling locations in the gradient. Long‐term lowering of WT was associated with decreased coverage of Sphagnum and aerenchymatic plants, decreased CH₄ field emissions and CH₄ production potential. Based on mcrA terminal restriction fragment length polymorphism the methanogen community structure correlated best with the methane production and coverage of aerenchymatic plants along the gradient. Methanosarcinaceae and Methanocellales were found at the pristine wet end of the gradient, whereas the Fen cluster characterized the dry end. The methane‐oxidizing bacterial community consisted exclusively of Methylocystis bacteria, but interestingly of five different alleles (T, S, R, M, and O) of the particulate methane monooxygenase marker gene pmoA. The M allele was dominant in the wet locations, and the occurrence of alleles O, S, and T increased with drainage. The occurrence of the R allele that characterized the upper peat layer correlated with CH₄ oxidation potential. These results advance our understanding of mire dynamics after long‐term WT drawdown and of the microbiological bases of methane emissions from mires.     

Weak phylogenetic signal in physiological traits of methane‐oxidizing bacteria

The presence of phylogenetic signal is assumed to be ubiquitous. However, for microorganisms, this may not be true given that they display high physiological flexibility and have fast regeneration. This may result in fundamentally different patterns of resemblance, that is, in variable strength of phylogenetic signal. However, in microbiological inferences, trait similarities and therewith microbial interactions with its environment are mostly assumed to follow evolutionary relatedness. Here, we tested whether indeed a straightforward relationship between relatedness and physiological traits exists for aerobic methane‐oxidizing bacteria (MOB). We generated a comprehensive data set that included 30 MOB strains with quantitative physiological trait information. Phylogenetic trees were built from the 16S rRNA gene, a common phylogenetic marker, and the pmoA gene which encodes a subunit of the key enzyme involved in the first step of methane oxidation. We used a Blomberg's K from comparative biology to quantify the strength of phylogenetic signal of physiological traits. Phylogenetic signal was strongest for physiological traits associated with optimal growth pH and temperature indicating that adaptations to habitat are very strongly conserved in MOB. However, those physiological traits that are associated with kinetics of methane oxidation had only weak phylogenetic signals and were more pronounced with the pmoA than with the 16S rRNA gene phylogeny. In conclusion, our results give evidence that approaches based solely on taxonomical information will not yield further advancement on microbial eco‐evolutionary interactions with its environment. This is a novel insight on the connection between function and phylogeny within microbes and adds new understanding on the evolution of physiological traits across microbes, plants and animals.     

Remarkable Recovery and Colonization Behaviour of Methane Oxidizing Bacteria in Soil After Disturbance Is Controlled by Methane Source Only

Little is understood about the relationship between microbial assemblage history, the composition and function of specific functional guilds and the ecosystem functions they provide. To learn more about this relationship we used methane oxidizing bacteria (MOB) as model organisms and performed soil microcosm experiments comprised of identical soil substrates, hosting distinct overall microbial diversities (i.e., full, reduced and zero total microbial and MOB diversities). After inoculation with undisturbed soil, the recovery of MOB activity, MOB diversity and total bacterial diversity were followed over 3 months by methane oxidation potential measurements and analyses targeting pmoA and 16S rRNA genes. Measurement of methane oxidation potential demonstrated different recovery rates across the different treatments. Despite different starting microbial diversities, the recovery and succession of the MOB communities followed a similar pattern across the different treatment microcosms. In this study we found that edaphic parameters were the dominant factor shaping microbial communities over time and that the starting microbial community played only a minor role in shaping MOB microbial community     

Molecular Analyses of Novel Methanotrophic Communities in Forest Soil That Oxidize Atmospheric Methane

Forest and other upland soils are important sinks for atmospheric CH₄, consuming 20 to 60 Tg of CH₄ per year. Consumption of atmospheric CH₄ by soil is a microbiological process. However, little is known about the methanotrophic bacterial community in forest soils. We measured vertical profiles of atmospheric CH₄ oxidation rates in a German forest soil and characterized the methanotrophic populations by PCR and denaturing gradient gel electrophoresis (DGGE) with primer sets targeting the pmoA gene, coding for the α subunit of the particulate methane monooxygenase, and the small-subunit rRNA gene (SSU rDNA) of all life. The forest soil was a sink for atmospheric CH₄ in situ and in vitro at all times. In winter, atmospheric CH₄ was oxidized in a well-defined subsurface soil layer (6 to 14 cm deep), whereas in summer, the complete soil core was active (0 cm to 26 cm deep). The content of total extractable DNA was about 10-fold higher in summer than in winter. It decreased with soil depth (0 to 28 cm deep) from about 40 to 1 μg DNA per g (dry weight) of soil. The PCR product concentration of SSU rDNA of all life was constant both in winter and in summer. However, the PCR product concentration of pmoA changed with depth and season. pmoA was detected only in soil layers with active CH₄ oxidation, i.e., 6 to 16 cm deep in winter and throughout the soil core in summer. The same methanotrophic populations were present in winter and summer. Layers with high CH₄ consumption rates also exhibited more bands of pmoA in DGGE, indicating that high CH₄ oxidation activity was positively correlated with the number of methanotrophic populations present. The pmoA sequences derived from excised DGGE bands were only distantly related to those of known methanotrophs, indicating the existence of unknown methanotrophs involved in atmospheric CH₄ consumption.     

Active Methanotrophs in Two Contrasting North American Peatland Ecosystems Revealed Using DNA-SIP

The active methanotroph community was investigated in two contrasting North American peatlands, a nutrient-rich sedge fen and nutrient-poor Sphagnum bog using in vitro incubations and ¹³C-DNA stable-isotope probing (SIP) to measure methane (CH₄) oxidation rates and label active microbes followed by fingerprinting and sequencing of bacterial and archaeal 16S rDNA and methane monooxygenase (pmoA and mmoX) genes. Rates of CH₄ oxidation were slightly, but significantly, faster in the bog and methanotrophs belonged to the class Alphaproteobacteria and were similar to other methanotrophs of the genera Methylocystis, Methylosinus, and Methylocapsa or Methylocella detected in, or isolated from, European bogs. The fen had a greater phylogenetic diversity of organisms that had assimilated ¹³C, including methanotrophs from both the Alpha- and Gammaproteobacteria classes and other potentially non-methanotrophic organisms that were similar to bacteria detected in a UK and Finnish fen. Based on similarities between bacteria in our sites and those in Europe, including Russia, we conclude that site physicochemical characteristics rather than biogeography controlled the phylogenetic diversity of active methanotrophs and that differences in phylogenetic diversity between the bog and fen did not relate to measured CH₄ oxidation rates. A single crenarchaeon in the bog site appeared to be assimilating ¹³C in 16S rDNA; however, its phylogenetic similarity to other CO₂-utilizing archaea probably indicates that this organism is not directly involved in CH₄ oxidation in peat.     

Effect of Afforestation and Reforestation of Pastures on the Activity and Population Dynamics of Methanotrophic Bacteria

We investigated the effect of afforestation and reforestation of pastures on methane oxidation and the methanotrophic communities in soils from three different New Zealand sites. Methane oxidation was measured in soils from two pine (Pinus radiata) forests and one shrubland (mainly Kunzea ericoides var. ericoides) and three adjacent permanent pastures. The methane oxidation rate was consistently higher in the pine forest or shrubland soils than in the adjacent pasture soils. A combination of phospholipid fatty acid (PLFA) and stable isotope probing (SIP) analyses of these soils revealed that different methanotrophic communities were active in soils under the different vegetations. The C₁₈ PLFAs (signature of type II methanotrophs) predominated under pine and shrublands, and C₁₆ PLFAs (type I methanotrophs) predominated under pastures. Analysis of the methanotrophs by molecular methods revealed further differences in methanotrophic community structure under the different vegetation types. Cloning and sequencing and terminal-restriction fragment length polymorphism analysis of the particulate methane oxygenase gene (pmoA) from different samples confirmed the PLFA-SIP results that methanotrophic bacteria related to type II methanotrophs were dominant in pine forest and shrubland, and type I methanotrophs (related to Methylococcus capsulatus) were dominant in all pasture soils. We report that afforestation and reforestation of pastures caused changes in methane oxidation by altering the community structure of methanotrophic bacteria in these soils.     

Interactions between Thaumarchaea,Nitrospira and methanotrophs modulate autotrophic nitrification in volcanic grassland soil

Ammonium/ammonia is the sole energy substrate of ammonia oxidizers, and is also an essential nitrogen source for other microorganisms. Ammonia oxidizers therefore must compete with other soil microorganisms such as methane-oxidizing bacteria (MOB) in terrestrial ecosystems when ammonium concentrations are limiting. Here we report on the interactions between nitrifying communities dominated by ammonia-oxidizing archaea (AOA) and Nitrospira-like nitrite-oxidizing bacteria (NOB), and communities of MOB in controlled microcosm experiments with two levels of ammonium and methane availability. We observed strong stimulatory effects of elevated ammonium concentration on the processes of nitrification and methane oxidation as well as on the abundances of autotrophically growing nitrifiers. However, the key players in nitrification and methane oxidation, identified by stable-isotope labeling using ¹³CO₂ and ¹³CH₄, were the same under both ammonium levels, namely type 1.1a AOA, sublineage I and II Nitrospira-like NOB and Methylomicrobium-/Methylosarcina-like MOB, respectively. Ammonia-oxidizing bacteria were nearly absent, and ammonia oxidation could almost exclusively be attributed to AOA. Interestingly, although AOA functional gene abundance increased 10-fold during incubation, there was very limited evidence of autotrophic growth, suggesting a partly mixotrophic lifestyle. Furthermore, autotrophic growth of AOA and NOB was inhibited by active MOB at both ammonium levels. Our results suggest the existence of a previously overlooked competition for nitrogen between nitrifiers and methane oxidizers in soil, thus linking two of the most important biogeochemical cycles in nature.     

Spatiotemporal variations in an assemblage of closely related planktonic aerobic methanotrophs

1. The assemblage of aerobic methane‐oxidising bacteria (MOB) was investigated in different seasons in the water column of a stratified freshwater lake. Species composition was analysed by performing denaturing gradient gel electrophoresis (DGGE) of the 16S rRNA genes and cloning analysis of the pmoA gene, which encodes the α‐subunit of particulate methane monooxygenase. The relative abundance of MOB to total bacteria was deduced from the copy number of the pmoA gene and 16S rRNA gene using real‐time polymerase chain reaction. 2. The profiles of the DGGE banding patterns changed with water depth, and these changes correlated with oxygen concentration and water temperature. The sequences of the DGGE bands obtained were all associated with the genus Methylobacter. During the analysis of pmoA gene, all clones sequenced were that of the Methylobacter/Methylosarcina group. The relative abundances of pmoA gene peaked around the oxycline, and small peaks of pmoA gene were also observed near the surface when peaks of methane were observed at the corresponding depth. 3. Profiles of the DGGE banding patterns suggested that ecophysiological characteristics differ among members of the genus Methylobacter; this indicates the importance of investigating the MOB assemblage at the species level or lower. Planktonic MOB seemed to be abundant around oxycline.     

Methanotrophic Activity,Abundance, and Diversity in Forested Swamp Pools: Spatiotemporal Dynamics and Influences on Methane Fluxes

Methane oxidation (methanotrophy) in the water column and sediments of forested swamp pools likely control seasonal and annual emission of CH₄ from these systems, but the methanotrophic microbial communities, their activities, locations, and overall impact, is poorly understood. Several techniques including ¹⁴CH₄ oxidation assays, culture-based most probable number (MPN) estimates of methane-oxidizing bacteria (MOB) and protozoan abundance, MOB strain isolation and characterization, and PCR techniques were used to investigate methanotrophy at a forested swamp near Ithaca, New York. The greatest methanotrophic activity and largest numbers of MOB occurred predominantly at the low oxygen sediment/water interface in the water column. Seasonally, methanotrophic activity was very dynamic, ranging from 0.1 to 61.9 μ moles CH₄ d^?1 g^?1 dry sediment, and correlated most strongly with dissolved inorganic carbon (r = 0.896). Incorporation of methanotrophic variables (abundance and activity) into existing CH₄ flux regression models improved model fit, particularly during mid summer when CH₄ fluxes were most dynamic. Annually integrated methane flux and methanotrophic activity measurements indicate that differences in methanotrophic activity at the sediment/water interface likely accounted for differences in the annual CH₄ emission from the field site. Direct isolations of MOB resulted in the repeated isolation of organisms most closely related to Methylomonas methanica S1. A single acidophilic, type II MOB related to Methylocella palustris K was also isolated. Using a PCR-based MPN method and 16S rRNA genome copy number from isolates and control strains, type I and type II MOB were enumerated and revealed type I dominance of the sediment-associated MOB community.     

Colonization of rice roots with methanogenic archaea controls photosynthesis‐derived methane emission

The methane emitted from rice fields originates to a large part (up to 60%) from plant photosynthesis and is formed on the rice roots by methanogenic archaea. To investigate to which extent root colonization controls methane (CH₄) emission, we pulse‐labeled rice microcosms with ¹³CO₂ to determine the rates of ¹³CH₄ emission exclusively derived from photosynthates. We also measured emission of total CH₄ (¹²⁺¹³CH₄), which was largely produced in the soil. The total abundances of archaea and methanogens on the roots and in the soil were analysed by quantitative polymerase chain reaction of the archaeal 16S rRNA gene and the mcrA gene coding for a subunit of the methyl coenzyme M reductase respectively. The composition of archaeal and methanogenic communities was determined with terminal restriction fragment length polymorphism (T‐RFLP). During the vegetative growth stages, emission rates of ¹³CH₄ linearly increased with the abundance of methanogenic archaea on the roots and then decreased during the last plant growth stage. Rates of ¹³CH₄ emission and the abundance of methanogenic archaea were lower when the rice was grown in quartz‐vermiculite with only 10% rice soil. Rates of total CH₄ emission were not systematically related to the abundance of methanogenic archaea in soil plus roots. The composition of the archaeal communities was similar under all conditions; however, the analysis of mcrA genes indicated that the methanogens differed between the soil and root. Our results support the hypothesis that rates of photosynthesis‐driven CH₄ emission are limited by the abundance of methanogens on the roots.     

Molecular characterization of a microbial consortium involved in methane oxidation coupled to denitrification under micro‐aerobic conditions

Methane can be used as an alternative carbon source in biological denitrification because it is nontoxic, widely available and relatively inexpensive. A microbial consortium involved in methane oxidation coupled to denitrification (MOD) was enriched with nitrite and nitrate as electron acceptors under micro‐aerobic conditions. The 16S rRNA gene combined with pmoA phylogeny of methanotrophs and nirK phylogeny of denitrifiers were analysed to reveal the dominant microbial populations and functional microorganisms. Real‐time quantitative polymerase chain reaction results showed high numbers of methanotrophs and denitrifiers in the enriched consortium. The 16S rRNA gene clone library revealed that Methylococcaceae and Methylophilaceae were the dominant populations in the MOD ecosystem. Phylogenetic analyses of pmoA gene clone libraries indicated that all methanotrophs belonged to Methylococcaceae, a type I methanotroph employing the ribulose monophosphate pathway for methane oxidation. Methylotrophic denitrifiers of the Methylophilaceae that can utilize organic intermediates (i.e. formaldehyde, citrate and acetate) released from the methanotrophs played a vital role in aerobic denitrification. This study is the first report to confirm micro‐aerobic denitrification and to make phylogenetic and functional assignments for some members of the microbial assemblages involved in MOD.     

Abundance, Activity, and Community Structure of Pelagic Methane-Oxidizing Bacteria in Temperate Lakes

The abundance and activity of methane-oxidizing bacteria (MOB) in the water column were investigated in three lakes with different contents of nutrients and humic substances. The abundance of MOB was determined by analysis of group-specific phospholipid fatty acids from type I and type II MOB, and in situ activity was measured with a ¹⁴CH₄ transformation method. The fatty acid analyses indicated that type I MOB most similar to species of Methylomonas, Methylomicrobium, and Methylosarcina made a substantial contribution (up to 41%) to the total bacterial biomass, whereas fatty acids from type II MOB generally had very low concentrations. The MOB biomass and oxidation activity were positively correlated and were highest in the hypo- and metalimnion during summer stratification, whereas under ice during winter, maxima occurred close to the sediments. The methanotroph biomass-specific oxidation rate (V) ranged from 0.001 to 2.77 mg CH₄-C mg⁻¹ C day⁻¹ and was positively correlated with methane concentration, suggesting that methane supply largely determined the activity and biomass distribution of MOB. Our results demonstrate that type I MOB often are a large component of pelagic bacterial communities in temperate lakes. They represent a potentially important pathway for reentry of carbon and energy into pelagic food webs that would otherwise be lost as evasion of CH₄.