Structure and function of methanotrophic communities in a landfill-cover soil

In landfill-cover soils, aerobic methane-oxidizing bacteria (MOB) convert CH(4) to CO(2), mitigating emissions of the greenhouse gas CH(4) to the atmosphere. We investigated overall MOB community structure and assessed spatial differences in MOB diversity, abundance and activity in a Swiss landfill-cover soil. Molecular cloning, terminal restriction-fragment length polymorphism (T-RFLP) and quantitative PCR of pmoA genes were applied to soil collected from 16 locations at three different depths to study MOB community structure, diversity and abundance; MOB activity was measured in the field using gas push-pull tests. The MOB community was highly diverse but dominated by Type Ia MOB, with novel pmoA sequences present. Type II MOB were detected mainly in deeper soil with lower nutrient and higher CH(4) concentrations. Substantial differences in MOB community structure were observed between one high- and one low-activity location. MOB abundance was highly variable across the site [4.0 × 10(4) to 1.1 × 10(7) (g soil dry weight)(-1)]. Potential CH(4) oxidation rates were high [1.8-58.2 mmol CH(4) (L soil air)(-1) day(-1) ] but showed significant lateral variation and were positively correlated with mean CH(4) concentrations (P < 0.01), MOB abundance (P < 0.05) and MOB diversity (weak correlation, P < 0.17). Our findings indicate that Methylosarcina and closely related MOB are key players and that MOB abundance and community structure are driving factors in CH(4) oxidation at this landfill.     

Molecular analyses of novel methanotrophic communities in forest soil that oxidize atmospheric methane

Forest and other upland soils are important sinks for atmospheric CH(4), consuming 20 to 60 Tg of CH(4) per year. Consumption of atmospheric CH(4) by soil is a microbiological process. However, little is known about the methanotrophic bacterial community in forest soils. We measured vertical profiles of atmospheric CH(4) 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 alpha 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 CH(4) in situ and in vitro at all times. In winter, atmospheric CH(4) 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 microg 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 CH(4) 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 CH(4) consumption rates also exhibited more bands of pmoA in DGGE, indicating that high CH(4) 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 CH(4) consumption.     

Aerobic and anaerobic methanotrophic communities in urban landscape wetland

Applied Microbiology and Biotechnology - Both aerobic methane-oxidizing bacteria (MOB) and nitrite-dependent anaerobic methane oxidation (n-damo) organisms can be important methane sinks in a...     

Potential activity of methane and ammonium oxidation by methanotrophic communities from the soda lakes of southern Transbaikal

Radioisotopic measurements of the methane consumption by mud samples taken from nine Southern Transbaikal soda lakes (pH 9.5–10.6) showed an intense oxidation of methane in the muds of Lakes Khuzhirta, Bulamai Nur, Gorbunka, and Suduntuiskii Torom, with the maximum oxidation rate in the mud of Lake Khuzhirta (33.2 nmol/(ml day)). The incorporation rate of the radioactive label from¹⁴CH₄ into¹⁴CO₂ was higher than into acid-stable metabolites. Optimum pH values for methane oxidation in water samples were 7–8, whereas mud samples exhibited two peaks of methane oxidation activity (at pH 8.15–9.4 and 5.8–6.0). The majority of samples could oxidize ammonium to nitrites; the oxidation was inhibited by methane. The PCR amplification analysis of samples revealed the presence of genes encoding soluble and paniculate methane monooxygenase and methanol dehydrogenase. Three alkaliphilic methanotrophic bacteria of morphotype I were isolated from mud samples in pure cultures, one of which, B5, was able to oxidize ammonium to nitrites at pH 7–11. The data obtained suggest that methanotrophs are widely spread in the soda lakes of Southern Transbaikal, where they can actively oxidize methane and ammonium.     

Response of methanotrophic communities to afforestation and reforestation in New Zealand

Methanotrophs use methane (CH₄) as a carbon source. They are particularly active in temperate forest soils. However, the rate of change of CH₄ oxidation in soil with afforestation or reforestation is poorly understood. Here, soil CH₄ oxidation was examined in New Zealand volcanic soils under regenerating native forests following burning, and in a mature native forest. Results were compared with data for pasture to pine land-use change at nearby sites. We show that following soil disturbance, as little as 47 years may be needed for development of a stable methanotrophic community similar to that in the undisturbed native forest soil. Corresponding soil CH₄-oxidation rates in the regenerating forest soil have the potential to reach those of the mature forest, but climo-edaphic fators appear limiting. The observed changes in CH₄-oxidation rate were directly linked to a prior shift in methanotrophic communities, which suggests microbial control of the terrestrial CH₄ flux and identifies the need to account for this response to afforestation and reforestation in global prediction of CH₄ emission.     

Activity and species composition of aerobic methanotrophic communities in tundra soils

The low-temperature, methane-oxidizing activities and species composition of methanotrophic communities in various tundra bog soils were investigated by radioisotopic and immunofluorescent methods. Methanotrophic bacteria carried out the methane oxidation process through all horizons of seasonally thawed layers down to permafrost. The highest activity of the process has been observed in the water surface layer of overmoistured soils and in water-logged moss covers. Up to 40% of¹⁴CH₄ added was converted into¹⁴CO₂, bacterial biomass, and organic exometabolites. By immunofluoresecent analysis it was demonstrated that the representatives of I+X (Methylomonas, Methylobacter, andMethylococcus) and II (Methylosinus, Methylocystis) methanotrophic groups occurred simultaneously in all samples at 61.6% and 38.4%, respectively. The number of methane-oxidizing bacteria in the ecosystems studied was 0.1–22.9×10⁶ cells per gram of soil. Methanotrophic organisms ranged from 1% to 23% of the total bacterial number.     

Analysis of methanotrophic communities in landfill biofilters using diagnostic microarray

Biofilters operated for the microbial oxidation of landfill methane at two sites in Northern Germany were analysed for the composition of their methanotrophic community by means of diagnostic microarray targeting the pmoA gene of methanotrophs. The gas emitted from site Francop (FR) contained the typical principal components (CH4, CO2, N2) only, while the gas at the second site Müggenburger Strasse (MU) was additionally charged with non-methane volatile organic compounds (NMVOCs). Methane oxidation activity measured at 22 degrees C varied between 7 and 103 microg CH4 (g dw)(-1) h(-1) at site FR and between 0.9 and 21 microg CH4 (g dw)(-1) h(-1) at site MU, depending on the depth considered. The calculated size of the active methanotrophic population varied between 3 x 10(9) and 5 x 10(11) cells (g dw)(-1) for biofilter FR and 4 x 10(8) to 1 x 10(10) cells (g dw)(-1) for biofilter MU. The methanotrophic community in both biofilters as well as the methanotrophs present in the landfill gas at site FR was strongly dominated by type II organisms, presumably as a result of high methane loads, low copper concentration and low nitrogen availability. Within each biofilter, community composition differed markedly with depth, reflecting either the different conditions of diffusive oxygen supply or the properties of the two layers of materials used in the filters or both. The two biofilter communities differed significantly. Type I methanotrophs were detected in biofilter FR but not in biofilter MU. The type II community in biofilter FR was dominated by Methylocystis species, whereas the biofilter at site MU hosted a high abundance of Methylosinus species while showing less overall methanotroph diversity. It is speculated that the differing composition of the type II population at site MU is driven by the presence of NMVOCs in the landfill gas fed to the biofilter, selecting for organisms capable of co-oxidative degradation of these compounds.     

Aerobic methanotrophic communities in the bottom sediments of Lake Baikal

The results of the first methodical investigation into the aerobic methanotrophic communities inhabiting the bottom sediments of Lake Baikal are reported. Use of the radioisotopic method revealed methane consumption in 12 10- to 50-cm-long sediment cores. The maximum methane consumption rates (495-737 microl/(dm3 day) were recorded in sediments in the regions of hydrothermal vents and oil and gas occurrence. Methane consumption was most active in the surface layers of the sediments (0-4 cm); it decreased with the sediment depth and became negligible or absent at depths below 20 cm. The number of methanotrophic bacteria usually ranged from 100 to 1000 cells/cm3 of sediment and reached 1 million cells/cm3 in the regions of oil and gas occurrence. The 17 enrichment cultures obtained were represented mainly by morphotype II methanotrophs. Phylogenetic analysis of the enrichment cultures in terms of the amino acid sequence of the alpha subunit of the membrane-bound methane monooxygenase revealed the predominance of methanotrophs of the genus Methylocystis. The results obtained suggest the presence of an active aerobic methanotrophic community in Lake Baikal.     

Mineralization of hexachlorocyclohexane in soil during solid-phase bioremediation

Soil containing hexachlorocyclohexane (HCH) was spiked with ¹⁴C--HCH and then subjected to bioremediation in bench-scale microcosms to determine the rate and extent of mineralization of the ¹⁴C-labeled HCH to ¹⁴CO₂. The soil was treated using two different DARAMEND amendments, D6386 and D6390. The amendments were previously found to enhance natural HCH bioremediation as determined by measuring the disappearance of parent compounds under either strictly oxic conditions (D6386), or cycled anoxic/oxic conditions (D6390). Within 80 days of the initiation of treatment, mineralization was observed in all of the strictly oxic microcosms. However, mineralization was negligible in the cycled anoxic/oxic microcosms throughout the 275-day study, even after cycling was ceased at 84 days and although significant removal (up to 51%) of indigenous -HCH (146 mg/kg) was detected by GC with electron capture detector. Of the amended, strictly oxic treatments, only one, in which 47% of the spiked ¹⁴C-HCH was recovered as ¹⁴CO₂, enhanced mineralization compared with an unamended treatment (in which 34% recovery was measured). Other oxic treatments involving higher amendment application rates or auxiliary carbon sources were inhibitory to mineralization. Thus, although HCH degradation occurs during the application of either oxic or cycled anoxic/oxic DARAMEND treatments, mineralization of -HCH may be inhibited depending on the amendment and treatment protocol.     

Long-term effects of mineral versus organic fertilizers on activity and structure of the methanotrophic community in agricultural soils

Agricultural practices, such as mineral nitrogen fertilization, have an impact on the soil's ability to oxidize methane, but little is known about the shifts in the methanotrophic community composition associated with these practices. Therefore, the long-term effect of both mineral (NH4NO3) and organic (manure and GFT-compost) fertilizer applications on the soil methanotrophic community activity and structure were investigated. Both high and low affinity methane oxidation rates were lower in the soil treated with mineral fertilizer compared to the other soils. An enhanced nitrate concentration was observed in the mineral fertilized soil but nitrate did not show a direct affect on the high affinity methane oxidation. In contrast, the low affinity methane oxidation was slowed down by increased nitrate concentrations, which suggests a direct effect of nitrate on low affinity methane oxidation. Denaturing gradient gel electrophoresis (DGGE) analysis of 16S rRNA gene fragments specific for methanotrophs revealed a distinct community between the mineral and organic fertilized soils as extra Type I methanotrophic bands (phylotypes) became visible in the organic fertilized soils. These phylotypes were not visible in the patterns of the added organic fertilizers suggesting an indirect effect of the organic fertilizers on the methanotrophic community. Additionally, a molecular analysis was performed after the low affinity methane oxidation test. The enhanced methane concentrations used in the test enriched certain low affinity methanotrophs in the organic fertilized soils but not in the mineral fertilized soil. Supporting the molecular and functional observations, fatty acids characteristic for methanotrophs were less abundant in the soil treated with mineral fertilizer compared to the soil treated with compost. In conclusion, the function and molecular and chemical composition of the methanotrophic community are all altered in soil fertilized with mineral fertilizer.     

Effects of O2 and CH4 on presence and activity of the indigenous methanotrophic community in rice field soil

The activity and distribution of methanotrophs in soil depend on the availability of CH4 and O2. Therefore, we investigated the activity and structure of the methanotrophic community in rice field soil under four factorial combinations of high and low CH4 and O2 concentrations. The methanotrophic population structure was resolved by denaturant gradient gel electrophoresis (DGGE) with different PCR primer sets targeting the 16S rRNA gene, and two functional genes coding for key enzymes in methanotrophs, i.e. the particulate methane monooxygenase (pmoA) and the methanol dehydrogenase (mxaF). Changes in the biomass of type I and II methanotrophic bacteria in the rice soil were determined by analysis of phospholipid-ester-linked fatty acid (PLFA) biomarkers. The relative contribution of type I and II methanotrophs to the measured methane oxidation activity was determined by labelling of soil samples with 14CH4 followed by analysis of [14C]-PLFAs. CH4 oxidation was repressed by high O2 (20.5%), and enhanced by low O2 (1%). Depending on the CH4 and O2 mixing ratios, different methanotrophic communities developed with a higher diversity at low than at high CH4 concentration as revealed by PCR-DGGE. However, a prevalence of type I or II populations was not detected. The [14C]-PLFA fingerprints, on the other hand, revealed that CH4 oxidation activity was dominated by type I methanotrophs in incubations with low CH4 mixing ratios (1000 p.p.m.v.) and during initiation of CH4 consumption regardless of O2 or CH4 mixing ratio. At high methane mixing ratios (10 000 p.p.m.v.), type I and II methanotrophs contributed equally to the measured CH4 metabolism. Collectively, type I methanotrophs responded fast and with pronounced shifts in population structure and dominated the activity under all four gas mixtures. Type II methanotrophs, on the other hand, although apparently more abundant, always present and showing a largely stable population structure, became active later and contributed to CH4 oxidation activity mainly under high CH4 mixing ratios.     

转基因水稻对土壤微生物群落结构及功能的影响

以非转基因水稻为对照,以变性梯度凝胶电泳(DGGE)和Biolog技术为手段,研究了2种转基因水稻对土壤微生物群落结构及功能的影响。结果显示:转基因水稻仅在生长发育旺盛期对土壤细菌数量有显著影响;且不同品种转基因水稻土壤微生物间的遗传距离大于转基因水稻与对照间土壤微生物的距离,即2个转基因水稻品种对土壤微生物群落遗传多样性的影响均不显著;在水稻抽穗期,2种转基因水稻与其对照的土壤微生物群落在72h时的平均光密度呈现显著差异,而到了成熟期则差异不显著。土壤微生物群落多样性指数和均匀度指数也表现出类似趋势。本试验证明,在水稻生长发育旺盛时期,Mclntosh指数(u)是一个有效区分转基因水稻和非转基因水稻土壤微生物群落多样性的指标。     

Quantification of mcrA by fluorescent PCR in methanogenic and methanotrophic microbial communities

A quantitative fluorogenic PCR method for detecting methanogenic and methanotrophic orders was established using a refined primer set for the methyl coenzyme M reductase subunit A gene (mcrA). The method developed was applied to several microbial communities in which diversity and abundance of methanogens or anaerobic methanotrophs (ANMEs) was identified by 16S rRNA gene clone analysis, and strong correlations between the copy numbers of mcrA with those of archaeal 16S rRNA genes in the communities were observed. The assay can be applied to detecting and assessing the abundance of methanogens and/or ANMEs in anoxic environments that could not be detected by 16S rRNA gene sequence analyses.     

Nitrification inhibition in a flooded soil by hexachlorocyclohexane and carbofuran

Summary The effect of a commercial granular formulation of hexachlorocyclohexane (HCH) on nitrification in a flooded soil was studied at 10 and 100 ppm a.i. The oxidation of the added ammonium to nitrate was inhibited significantly at 10 ppm and almost completely at 100 ppm, concomitant with a proportional decrease in the, populations of ammonium- and nitrite-oxidising autotrophic bacteria. Of special interest is the synergistic increase in the inhibition of nitrification by a combined application of HCH and carbofuran.     

The effect of predation on the structure and invasibility of assembled communities

Summary Fifty four microcosmic communities were assembled over 4 months from a 28-species source pool of phytoplankton using nine different invasion patterns each replicated six times. Three communities from each set of replicates then were invaded with a cladoceran that feeds on phytoplankton. All communities were then treated identically for an additional 4 months. In all nine invasion categories species richness was greater in predated communities. Predation opened communities to invasion by increasing the representation of infrequently sampled species at the expense of more common species. Invasion rate was four times more influential than predation and over eleven times more important than either invasion order or the timing pattern of interspecific arrivals in determining species richness in this system of communitites.     

Recovery of in-situ methanotrophic activity following acetylene inhibition

Methane (CH₄) is the second most important greenhouse gas after carbon dioxide (CO₂). To understand CH₄ cycling, quantitative information about microbial CH₄ oxidation in soils is essential. Field methods such as the gas push-pull test (GPPT) to quantify CH₄ oxidation are often used in combination with specific inhibitors, such as acetylene (C₂H₂). Acetylene irreversibly binds to the enzyme methane monooxygenase, but little is known about recovery of CH₄ oxidation activity after C₂H₂ inhibition in situ, which is important when performing several experiments at the same location. To assess recovery of CH₄ oxidation activity following C₂H₂ inhibition, we performed a series of GPPTs over 8 weeks at two different locations in the vadose zone above a petroleum hydrocarbon-contaminated aquifer in Studen, Switzerland. After 4 weeks a maximum recovery of 30% and 50% of the respective initial activity was reached, with a subsequent slight drop in activity at both locations. Likely, CH₄ oxidation activity and CH₄ concentrations were too low to allow for rapid recovery following C₂H₂ inhibition at the studied locations. Therefore, alternative competitive inhibitors have to be evaluated for application in conjunction with GPPTs, especially for sites with low activity.