Dependent Methanogenesis Research Articles

Methane formation and consumption by sediments in a cross-channel profile of a small river impoundment

Rivers are a natural source of methane (CH4) into the atmosphere and may contribute significantly to total CH4 emissions. Even though the details of sources of CH4 in rivers are not fully understood, weirs have been recognized as a hotspot of CH4 emissions. In this study, we investigated CH4 production and consumption in air-exposed river sediments along a cross-channel transect located upstream of a weir. Stable carbon isotopes were used for determination of individual methanogenic pathways. In order to understand the relationship between physicochemical and biological processes, additional parameters such as organic matter, grain median size, and carbon and nitrogen content were characterized as well. Generally, samples from the surface sediment layer (0-10 cm) had higher CH4 production than sediments from the deeper layer (10-20 cm) during the incubation experiments. Sediments near the bank zones and in the mid-channel were characterized by the highest organic carbon content (6.9 %) as well the highest methanogenic activity (2.5 mmol g-1 DW d-1). The CH4 production was predominated by H2/CO2 dependent methanogenesis in the surface sediment layer (0-10 cm), while the proportion of acetoclastic and hydrogenotrophic methanogenesis in the deeper sediment layer (10-20 cm) was balanced. The CH4 oxidation potential of sediments showed the same spatial pattern as observed for the CH4 production. Our results showed high spatial variability of sediment CH4 production and oxidation in the cross-channel profile upstream of the weir, whereas the highest CH4 dynamics were observed in the littoral zones. This variability was closely linked with the carbon and nitrogen content in the sediment samples.

Methanogenic Pathway and Archaeal Community Structure in the Sediment of Eutrophic Lake Dagow: Effect of Temperature

Methanogenic degradation of organic matter is an important microbial process in lake sediments. Temperature may affect not only the rate but also the pathway of CH4 production by changing the activity and the abundance of individual microorganisms. Therefore, we studied the function and structure of a methanogenic community in anoxic sediment of Lake Dagow, a eutrophic lake in north-eastern Germany. Incubation of sediment samples (in situ 7.5 degrees C) at increasing temperatures (4, 10, 15, 25, 30 degrees C) resulted in increasing production rates of CH4 and CO2 and in increasing steady-state concentrations of H2. Thermodynamic conditions for H2/CO2 -dependent methanogenesis were only exergonic at 25 and 30 degrees C. Inhibition of methanogenesis with chloroform resulted in the accumulation of methanogenic precursors, i.e., acetate, propionate, and isobutyrate. Mass balance calculations indicated that less CH4 was formed via H2 at 4 degrees C than at 30 degrees C. Conversion of 14CO2 to 14CH4 also showed that H2/CO2 -dependent methanogenesis contributed less to total CH4 production at 4 degrees C than at 30 degrees C. [2-14 C]Acetate turnover rates at 4 degrees C accounted for a higher percentage of total CH4 production than at 30 degrees C. Collectively, these results showed a higher contribution of H2-dependent methanogenesis and a lower contribution of acetate-dependent methanogenesis at high versus low temperature. The archaeal community was characterized by cloning, sequencing, and phylogenetic analysis of the 16S rRNA genes retrieved from the sediment. Sequences were affiliated with Methanosaetaceae, Methanomicrobiaceae, and three deeply branching euryarchaeotal clusters, i.e., group III, Rice cluster V, and a novel euryarchaeotal cluster, the LDS cluster. Terminal restriction fragment length polymorphism (T-RFLP) analysis showed that 16S rRNA genes affiliated to Methanosaetaceae (20-30%), Methanomicrobiaceae (35-55%), and group III (10-25%) contributed most to the archaeal community. Incubation of the sediment at different temperatures (4-30 degrees C) did not result in a systematic change of the archaeal community composition, indicating that change of temperature primarily affected the activity rather than the structure of the methanogenic community.

Seasonal variation in pathways of CH4 production and in CH4 oxidation in rice fields determined by stable carbon isotopes and specific inhibitors

AbstractFlooded rice fields, which are an important source of the atmospheric methane, have become a model system for the study of interactions between various microbial processes. We used a combination of stable carbon isotope measurements and application of specific inhibitors in order to investigate the importance of various methanogenic pathways and of CH4 oxidation for controlling CH4 emission. The fraction of CH4 produced from acetate and H2/CO2 was calculated from the isotopic signatures of acetate, carbon dioxide (CO2) and methane (CH4) measured in porewater, gas bubbles, in the aerenchyma of the plants and/or in incubation experiments. The calculated ratio between both pathways reflected well the ratio determined by application of methyl fluoride (CH3F) as specific inhibitor of acetate‐dependent methanogenesis. Only at the end of the season, the theoretical ratio of acetate: H2 = 2 : 1 was reached, whereas at the beginning H2/CO2‐dependent methanogenesis dominated. The isotope discrimination was different between rooted surface soil and unrooted deep soil. Root‐associated CH4 production was mainly driven by H2/CO2. Porewater CH4 was found to be a poor proxy for produced CH4.The fraction of CH4 oxidised was calculated from the isotopic signature of CH4 produced in vitro compared to CH4 emitted in situ, corrected for the fractionation during the passage from the aerenchyma to the atmosphere. Isotope mass balances and in situ inhibition experiments with difluoromethane (CH2F2) as specific inhibitor of methanotrophic bacteria agreed that CH4 oxidation was quantitatively important at the beginning of the season, but decreased later. The seasonal pattern was consistent with the change of potential CH4 oxidation rates measured in vitro. At the end of the season, isotope techniques detected an increase of oxidation activity that was too small to be measured with the flux‐based inhibitor technique. If porewater CH4 was used as a proxy of produced CH4, neither magnitude nor seasonal pattern of in situ CH4 oxidation could be reproduced. An oxidation signal was also found in the isotopic signature of CH4 from gas bubbles that were released by natural ebullition. In contrast, bubbles stirred up from the bulk soil had preserved the isotopic signature of the originally produced CH4.

Evidence for anaerobic syntrophic acetate oxidation during methane production in the profundal sediment of subtropical Lake Kinneret (Israel).

Methane production was measured in samples of the profundal sediment from Lake Kinneret. Production rates of CH(4) were higher at 30 degrees C than at the in situ temperature of 15 degrees C and were higher in the top 5 cm layer than below. Turnover of [2-(14)C]-acetate resulted in the production of (14)CH(4) and (14)CO(2) with turnover times of < 42 min. However, < 30% of the added radioactivity was converted to gaseous products, indicating that only part of the acetate pool was microbially available. The calculated acetate turnover rates were sufficient to account for total CH(4) production, indicating that CH(4) was produced exclusively from acetate. This conclusion was confirmed by inhibition of methanogens with chloroform, which resulted in an almost stoichiometric accumulation of acetate. However, a large percentage (30-60%) of [2-(14)C]-acetate was converted to (14)CO(2), despite lack of reducible sulphate or other oxidants in the sediment. Anoxic preincubation of the sediment did not result in reduced production of (14)CO(2). Therefore, part of the acetate must have been oxidized rather than methanogenically cleaved. Conversion of [(14)C]-bicarbonate to (14)CH(4) indicated that 30-50% of total CH(4) production originated from reduction of CO(2). To reconcile the relatively high contribution of H(2)/CO(2)-dependent methanogenesis with the relatively high oxidative conversion of acetate, we assume that part of the acetate was used syntrophically by consortia of acetate-oxidizing bacteria and H(2)/CO(2)-using methanogens. This conclusion is supported by favourable thermodynamic conditions for syntrophic acetate oxidation under in situ conditions and complete inhibition of [2-(14)C]-acetate turnover at high H(2) partial pressures. Further evidence to support this conclusion comes from the analysis of the structure of the archaeal community. Terminal restriction fragment length polymorphism (T-RFLP) and partial sequence analysis of the SSU rRNA genes amplified from DNA extracts of the sediment showed Methanomicrobiaceae as the dominant methanogenic group, whereas acetoclastic methanogens could not be detected.

Microbial processes influencing methane emission from rice fields

SummaryIrrigated rice fields are an important source of atmospheric methane. In order to improve our understanding of the controlling processes, we measured in situ CH4 emission and CH4 oxidation in an Italian rice field in 1998 and 1999, and studied CH4 production in soil and root samples. The CH4 emission rates were correlated with diurnal temperature variations and showed pronounced seasonal and interannual variations. The contribution of CH4 oxidation to total CH4 flux, determined by specific inhibition with difluoromethane, decreased from 40% at the beginning to zero at the end of the season. The stable carbon isotopic composition of the emitted CH4 also decreased. The CH4‐oxidizing bacteria probably became limited by nitrogen as indicated by the seasonal decrease of NH4+. Thus, CH4 oxidation had little effect on CH4 emission. Methane production on rice roots was relatively constant over the season. Methane production in soil slowly increased after flooding and was highest in the middle of the season. Pore water concentrations of CH4 showed a similar seasonal pattern. In 1999, CH4 production increased later in the season and reached lower rates than in 1998. An additional drainage in 1999 resulted in higher ferric iron concentrations, higher soil redox potentials and lower acetate concentrations. As a result, acetate‐utilizing methanogens were probably out‐competed by iron‐reducers so that a larger percentage of [2–14C]acetate was converted to 14CO2 instead of 14CH4. The residual CH4 production was relatively low and was mainly due to H2/CO2‐dependent methanogenesis. Experiments with radioactive bicarbonate and with methyl fluoride as specific inhibitor showed that the theoretical ratio of 7:3 of methanogenesis from acetate vs. H2/CO2 was only reached later in the season when total CH4 production was at the maximum. In conclusion, our results give a mechanistic explanation for the intraseasonal and interannual differences in CH4 emission.

Methane production in eutrophic Lake Plußsee: seasonal change, temperature effect and metabolic processes in the profundal sediment

Rates of CH4 production in the profundal sediment of Lake Plusssee were higher during the summer than during fall, and fairly well explained the CH 4 concentration that accumulated in the hypolymnion during summer stratification. Oxidation of CH 4 at the oxycline played only a marginal role in the seasonal CH 4 cycle. Although the sediment temperature (4 °C) did not change with season, CH 4 production rates could be stimulated by incubation at increased temperature (25 °C). At both 4 °C and 25 °C, thermodynamic conditions in the sediment allowed exergonic production of CH 4 from either acetate, H 2 /CO 2 or methanol. Inhibition of methanogenesis by 2-bromoethane sulfonic acid (BES) resulted in the accumulation of acetate, but was only detectable at 25 °C. Inhibition of acetoclastic methanogenesis by methyl fluoride resulted in partial inhibition of CH 4 and was only detectable at 25 °C, Incorporation of radioactive bicarbonate into CH 4 was also only detectable at 25 °C and then accounted for about 25-42 % of total CH 4 production. [2- 14 C]acetate was converted to 14 CH 4 with turnover times of about 24 and 97h at 25 °C and 4 °C, respectively, but accounted for only 4-9 % of total CH 4 production. The turnover times of radioactive methanol were even larger (about 96 at 25 °C and 153 h at 4 °C) and accounted for <1 % of total CH 4 production. Hence, these processes could account for a larger percentage of CH 4 production only if taking place in sediment microniches that did not fully equilibrate with the pore water. Production of CH 4 at 25 °C was stimulated by addition of pectin, acetate, methanol or H 2 , but at 4 °C, it was only stimulated by H 2 . The stable carbon isotopic composition of CH 4 and CO 2 in the water column and in incubated sediment slurries indicated that CH 4 production was dominated by acetoclastic methanogenesis, with H 2 /CO 2 -dependent methanogenesis contributing more at 25 °C than 4 °C. In conclusion, the data are consistent with the assumption that CH 4 production at in-situ temperature (4 °C) was due to acetoclastic methanogenesis tightly coupled to acetate production, whereas it was due to both acetoclastic and H 2 /CO 2 -dependent methanogenesis at 25 °C. This assumption is also consistent with the observation that the sediment fermented added glucose or pectin to acetate as a major product.

Methanogenic archaea and CO2-dependent methanogenesis on washed rice roots.

Washed excised roots of rice (Oryza sativa) immediately started to produce CH4 when they were incubated in phosphate buffer under anoxic conditions (N2 atmosphere), with initial rates varying between 2 and 70nmolh(-1)g(-1) dry weight of root material (mean +/- SE: 20.3 +/- 5.9 nmol h(-1) g(-1) dry weight; n = 18). Production of CH4 continued for at least 500 h, with rates usually decreasing slowly. CH4 production was not significantly affected by methyl fluoride, an inhibitor of acetoclastic methanogenesis. Less than 0.5% of added [2-14C]-acetate was converted to 14CH4, and conversion of 14CO2 to 14CH4 indicated that CH4 was almost exclusively produced from CO2. Occasionally, however, especially when the roots were incubated without additional buffer, CH4 production started to accelerate after about 200h reaching rates of > 100 nmol h(-1) g(-1) dry weight. Methyl fluoride inhibited methanogenesis by more than 20% only in these cases, and the conversion of 14CO2 to 14CH4 decreased. These results indicate that CO2-dependent rather than acetoclastic methanogenesis was primarily responsible for CH4 production in anoxically incubated rice roots. Determination of most probable numbers of methanogens on washed roots showed highest numbers (10(6)g(-1) dry roots) on H2 and ethanol, i.e. substrates that support CH4 production from CO2. Numbers on acetate (10(5) g(-1) dry roots) and methanol (10(4)g(-1) dry roots) were lower. Methanogenic consortia enriched on H2 and ethanol were characterized phylogenetically by comparative sequence analysis of archaeal small-subunit (SSU) ribosomal RNA-encoding genes (rDNA). These sequences showed a high similarity to SSU rDNA clones that had been obtained previously by direct extraction of total DNA from washed rice roots. The SSU rDNA sequences recovered from the H2/CO2-using consortium either belonged to a novel lineage of methanogens that grouped within the phylogenetic radiation of the Methanosarcinales and Methanomicrobiales or were affiliated with Methanobacterium bryantii. SSU rDNA sequences retrieved from the ethanol-using consortium either grouped within the genus Methanosarcina or belonged to another novel lineage within the phylogenetic radiation of the Methanosarcinales and Methanomicrobiales. Cultured organisms belonging to either of the two novel lineages have not been reported yet.

Presence of a Na+/H+ antiporter in Methanobacterium thermoautotrophicum and its role in Na+ dependent methanogenesis

Methane formation from H2/CO2 by methanogenic bacteria is dependent on Na+ ions. In this communication it is shown with Methanobacterium thermoautotrophicum that a Na+/H+ antiporter plays a role in methane formation from H2 and CO2 and in the regulation of the ΔpH. This is based on the following findings: