Metabolism of position-labelled glucose in anoxic methanogenic paddy soil and lake sediment

Abstract Turnover times of radioactive glucose were shorter in paddy soil (4–16 min) than in Lake Constance sediment (18–62 min). In the paddy soil, 65–75% of the radioactive glucose was converted to soluble metabolites. In the sediment, only about 25% of the radioactive glucose was converted to soluble metabolites, the rest to particulate material. In anoxic paddy soil, the degradation pattern of position-labelled glucose was largely consistent with glucose degradation via the Embden-Meyerhof-Parnas (EMP) pathway followed by methanogenic acetate cleavage: CO₂ mainly originated from C-3,4, whereas CH₄ mainly originated from C-1 and C-6 of glucose. Acetate-carbon originated from C-1, C-2 and C-6 rather than from C-3,4 of glucose. In both paddy soil and Lake Constance sediment acetate and CO₂ were the most important early metabolites of radioactive glucose. Other early products included propionate, ethanol/butyrate, succinate, and lactate, but accounted each for less than 1–8% of the glucose utilized. The labelling of propionate by [3,4-¹⁴C]glucose suggests that it was mainly produced from glucose or lactate rather than from ethanol. Isopropanol and caproate were also detectable in paddy soil, but were not produced from radioactive glucose. Chloroform inhibited methanogenesis, inhibited the further degradation of radioactive acetate and resulted in the accumulation of H₂, however, did not inhibit glucose degradation. Since acetate was the main soluble fermentation product of glucose and was produced at a relatively high molar acetate: CO₂ ratio (2.5:1), homoacetogenesis appeared to be the most important glucose fermentation pathway.     

Hydrogen turnover by psychrotrophic homoacetogenic and mesophilic methanogenic bacteria in anoxic paddy soil and lake sediment

Abstract The effect of temperature on CH₄ production, turnover of dissolved H₂, and enrichment of H₂-utilizing anaerobic bacteria was studied in anoxic paddy soil and sediment of Lake Constance. When anoxic paddy soil was incubated under an atmosphere of H₂/CO₂, rates of CH₄ production increased 25°C, but decreased at temperatures lower than 20°C. Chloroform completely inhibited methano-genesis in anoxic paddy soil and lake sediment, but did not or only partially inhibit the turnover of dissolved H₂, especially at low incubation temperatures. Cultures with H₂ as energy source resulted in the enrichment of chemolithotrophic homoacetogenic bacteria whenever incubation temperatures were lower than 20°C. Hydrogenotrophic methanogens could only be enriched at 30°C from anoxic paddy soil. A homoacetogen     

Bicarbonate-dependent production and methanogenic consumption of acetate in anoxic paddy soil

Abstract Acetate turnover was measured in slurries of anoxic methanogenic paddy soil after addition of carrier-free [2-¹⁴C]-acetate. Acetate concentrations stayed fairly constant for about 1–2 days indicating steady state between production and consumption reactions. Depending on the experiment, acetate concentrations were between 100 and 3000 μM. Turnover rates were determined from the logarithmic decrease of [2-¹⁴C]-acetate or from the accumulation of acetate in the presence of chloroform resulting in similar values, i.e. 12–13 nmol h⁻¹g⁻¹d.w. soil at 17°C and 36–88 nmol h⁻¹g⁻¹d.w. at 30°C. Acetate consumption was completely inhibited by chloroform. The respiratory index (RI) was < 0.27. Hence, acetate was apparently consumed by methanogenic bacteria. About 80–90% of the CH₄ produced originated from the methyl group of acetate. The role of homoacetogenesis for acetate production was studied by measuring the incorporation of radioactive bicarbonate into acetate. In different experiments, CO₂ incorporation accounted for fractions of 1–60% of the acetate produced, about 10% being the most likely value for steady-state conditions. The fraction increased at high H₂ concentrations and decreased at high acetate concentrations. The rate of H₂ production that was required for chemolithotrophic acetate production from CO₂ was two orders of magnitude higher than the actually measured rate. Hence, most of the CO₂ incorporation into acetate was caused by electron donors other than H₂ (e.g., carbohydrates) and/or by exchange reactions.     

Effect of dilution on methanogenesis, hydrogen turnover and interspecies hydrogen transfer in anoxic paddy soil

Abstract Dilution of anoxic slurries of paddy soil resulted in a proportional decrease of the rates of total methanogenesis and the rate constants of H₂ turnover per gram soil. Dilution did not affect the fraction of H₂/CO₂-dependent methanogenesis which made up 22% of total CH₄ production. However, dilution resulted in a ten fold decrease of the H₂ steady state partial pressure from approximately 4 to 0.4 Pa indicating that H₂/CO₂-dependent methanogenesis was more or less independent of the H₂ pool. The rates of H₂ production calculated from the H₂ turnover rate constants and the H₂ steady state partial pressures accounted for only < 5% of H₂/CO₂-dependent methanogenesis in undiluted soil slurries and for even less after dilution. Upon dilution, the Gibbs free energy available for H₂/CO₂-dependent methanogenesis decreased from −28.4 to only −5.6 kJ per mol. The results indicate that methane was mainly produced from interspecies H₂ transfer within syntrophic bacterial associations and was not significantly affected by the outside H₂ pool.     

Fermentation pattern of methanogenic degradation of rice straw in anoxic paddy soil

The anaerobic degradation of different fractions of rice straw in anoxic paddy soil was investigated. Rice straw was divided up into stem, leaf sheath and leaf blade. The different straw fractions were mixed with paddy soil and incubated under anoxic conditions. Fermentation of straw components started immediately and resulted in transient accumulation of acetate, propionate, butyrate, isobutyrate, valerate, isovalerate and caproate with much higher concentrations in the presence than in the absence of straw. Also some unidentified compounds with UV absorption could be detected. The maximum concentrations of these compounds were different when using different straw fractions, suggesting differences in the degradation pathway of these straw fractions during the early phase of incubation, i.e. with Fe(III) and sulfate serving as oxidants. When concentrations of the intermediates decreased to background values, CH(4) production started. Rates of CH(4)unamended soil. During the methanogenic phase, the percentage contribution of fermentation products to CH(4) production was determined by inhibition with 2-bromoethanesulfonate (BES). Acetate (48-83%) and propionate (18-28%) were found to be the main intermediates of the carbon flow to CH(4), irrespective of the fraction of the rice straw or its absence. Mass balance calculations showed that 84-89% of CH(4) was formed via acetate in the various incubations. Radiotracer experiments showed that 11-27% of CH(4) was formed from H(2)/CO(2), thus confirming that acetate contributed 73-89% to methanogenesis. Our results show that the addition of rice straw and the fraction of the straw affected the fermentation pattern only in the early phase of degradation, but had no effect on the degradation pathway during the later methanogenic phase.     

Hydrogen metabolism and sulfate-dependent inhibition of methanogenesis in a eutrophic lake sediment (Lake Mendota)

Hydrogen metabolism was studied in anoxic sediments of the stratified Lake Mendota; using a method which allowed the measurement of in situ H₂ concentrations and the headspace-free analysis of turnover of dissolved H₂. Addition of sulfate resulted in partial but immediate inhibition of H₂-dependent methanogenesis. Sulfate addition did not result in an immediate decrease in the steady state concentration of dissolved H₂, nor did it significantly stimulate the rate constant of H₂ turnover. Sulfate-induced decrease in dissolved H₂ was only observed after prolonged incubation or when endogenous H₂ production was stimulated by added glucose. The turnover of the in situ H₂ accounted for only 14% of the H₂-dependent methanogenesis from bicarbonate. While rates of methanogenesis increased during the season, rates of H₂ turnover decreased, accounting for only 2% of the H₂-dependent methanogenesis at the end of summer stratification. These observations indicate that increasing proportions of CH₄ were formed from H₂ being directly transferred in syntrophic methanogenic associations. The rapid inhibition of H₂-dependent methanogenesis by exogenous sulfate may be explained at least partially by assuming methanogenic associations in which syntrophic sulfate reducers change their metabolism from fermentative H₂ production to sulfate reduction.     

应用16S rDNA克隆文库技术研究长期稻草还田对水稻土细菌多样性的影响

利用中国科学院桃源农业生态试验站施肥制度长期定位试验田对照(CK)和稻草还田(OM)施肥处理的土壤样品,应用16S rDNA克隆文库技术直接提取土壤微生物总DNA,分别构建细菌16S rDNA克隆文库,并进行序列测定和分析。结果表明,与对照(CK)相比,稻草还田(OM)后土壤细菌群落结构发生了显著改变,土壤细菌多样性和均匀度均有所降低。对照(CK)和稻草还田(OM)两个施肥处理的优势种群均为变形菌,酸杆菌次之;稻草还田减少了变形菌、疣微菌、绿弯菌和绿菌的分布,而增加了硝化螺旋菌的分布。16S rDNA系统发育分析则表明,稻草还田对酸杆菌群落结构影响最大,其次是疣微菌和δ-变形菌。     

Inhibition of methanogens increases photo-dependent nitrogenase activities in anoxic paddy soil amended with rice straw

The interaction between phototrophic dinitrogen fixers and methanogens was examined in soil slurries amended with rice straw using 2-bromoethanesulfonic acid (BES), a specific methanogenic inhibitor. Slurries incubated in light increased phototrophic nitrogenase activity (acetylene reducing activity), and showed growth of phototrophic purple bacteria and reduction of CH(4) emission, indicating outcompetition of purple bacteria with methanogens in photic zones. Adding BES effectively inhibited methane production and markedly increased phototrophic acetylene reducing activity accompanied with acetate accumulation, but did not affect populations of purple bacteria in the slurries. More acetate accumulated in the inhibited slurries incubated in dark. We suggest that increased availability of organic substrates for purple bacteria after stopping methanogenic consumption by BES caused the increased phototrophic acetylene reducing activity. These results indicate that, after purple bacteria grow enough, performance of their N(2) fixation may be limited by substrate availability, which methanogenesis may profoundly influence.     

长期不同施肥措施下岩溶水稻土可培养细菌群落变化及其主要影响因素

【背景】施肥是目前提高作物产量的较优策略,不同的施肥措施在不同程度上影响土壤肥力和微生物群落结构。【目的】探究岩溶水稻土理化性质变化与细菌群落变化的对应关系,进而反映不同施肥措施对土壤可培养细菌群落的影响。最后选出最优施肥方案,为后续的合理施肥工作提供依据。【方法】对岩溶水稻土进行不施肥、常规施肥、常规施肥加绿肥3种施肥处理,通过对土壤理化性质、可培养细菌群落丰度及多样性变化的研究,探究在不同施肥措施下对岩溶水稻土壤细菌群落的影响。【结果】对比不施肥处理,常规施肥处理下土壤pH值和有机碳含量下降,结合大量研究结果证明,无机肥或氮肥的长期过量施加使土壤pH值下降,常规施肥加绿肥有利于有机碳的积累。分离纯化共得到164株菌,分别来自Actinobacteria、Bacteroidetes、Firmicutes和Proteobacteria。属水平上常规施肥配施绿肥较常规施肥组优势菌属Sphingomonas、Lysobacter的相对丰度增加。细菌群落多样性增加,出现Paenibacillus、Streptomyces和Pseudomonas等特有功能菌属。优势菌属Sphingopyxis、Lysobacter、Paenibacillus、Bosea、Streptomyces、Pseudomonas和Bacillus与TN存在显著正相关,在常规施肥加绿肥处理土壤中增加。【结论】常规施肥加绿肥处理下,固氮、溶磷等功能菌丰度增加,增加土壤肥力,保持土壤养分的可利用性,对作物的增产起重要作用。岩溶水稻土常规施肥配施绿肥处理的效果优于不施肥和常规施肥处理。     

Characteristics of boron accumulation by fly ash application in paddy soil

Fly ash has a high content of plant available silicate which is strongly needed for rice cultivation in Korea. One concern for plants grown on soils amended with fly ash is boron (B) toxicity because most of the fresh fly ash contains considerable B. This study was conducted in paddy soil to determine B uptake by rice and characteristics of B accumulation in soil after fly ash application (0, 40, 80, and 120 Mg fly ash ha⁻¹). In all fly ash treatments, B content in rice leaves and available B in soil at all growing stage were higher than those of control, but were not exceeded a toxicity levels. Boron occluded in amorphous Fe and Al oxides comprised ca. 20–39% of total B and was not affected by fly ash application. Most of the B was accumulated by fly ash application as a residual B which is plant-unavailable form, comprised >60% of the total B in soil. Thus, fly ash can be a good soil amendment for rice production without B toxicity.     

Changes in soil greenhouse gas fluxes by land use change from primary forest

Primary forest conversion is a worldwide serious problem associated with human disturbance and climate change. Land use change from primary forest to plantation, grassland or agricultural land may lead to profound alteration in the emission of soil greenhouse gases (GHG). Here, we conducted a global meta‐analysis concerning the effects of primary forest conversion on soil GHG emissions and explored the potential mechanisms from 101 studies. Our results showed that conversion of primary forest significantly decreased soil CO₂ efflux and increased soil CH₄ efflux, but had no effect on soil N₂O efflux. However, the effect of primary forest conversion on soil GHG emissions was not consistent across different types of land use change. For example, soil CO₂ efflux did not respond to the conversion from primary forest to grassland. Soil N₂O efflux showed a prominent increase within the initial stage after conversion of primary forest and then decreased over time while the responses of soil CO₂ and CH₄ effluxes were consistently negative or positive across different elapsed time intervals. Moreover, either within or across all types of primary forest conversion, the response of soil CO₂ efflux was mainly moderated by changes in soil microbial biomass carbon and root biomass while the responses of soil N₂O and CH₄ effluxes were related to the changes in soil nitrate and soil aeration‐related factors (soil water content and bulk density), respectively. Collectively, our findings highlight the significant effects of primary forest conversion on soil GHG emissions, enhance our knowledge on the potential mechanisms driving these effects and improve future models of soil GHG emissions after land use change from primary forest.