Methanotrophic Research Articles

Agro-technologies for greenhouse gases mitigation in flooded rice fields for promoting climate smart agriculture

We investigated methane (CH4) and nitrous oxide (N2O), two important greenhouse gases (GHGs) emissions using the closed chamber method from a flooded rice field in Brahmaputra valley of Assam, northeast part of India. We tried to understand the factors responsible for the emission and identify appropriate agro-technologies for their mitigation. Various factors like water level, drainage management, soil organic carbon management, crop management, fertilizer amendment, cultivar type etc. affect the GHG production and emission from the flooded rice soil. In this study, six treatments were employed, namely, farmer's practice (FP), recommended fertilizer dosage (RDF), direct seeded rice (DSR), intermittent wetting drying (IWD), use of efficient methanotrophs (MTH), and use of ammonium sulfate as a nitrogen source for real-time nitrogen management using leaf color chart, (AS). GHG flux was measured through the static closed chamber technique. Soil temperature, pH, and redox potential (Eh) and other soil physico-chemical and biological properties that have a potential role in GHG emission were also assessed. The lowest CH4 flux was observed in IWD treatment. The highest CH4 but lowest N2O flux was observed in RDF thus portraying a tradeoff relationship among these two GHGs. The highest N2O flux was observed in AS. Changes in Eh strongly altered CH4 and N2O emissions. The CH4 flux for the growing season varied from 62.5 to 86.3 kg ha−1 with an average of 72.4 kg ha−1. The average N2O flux was 0.89 kg ha−1 with values fluctuating between 0.72 – and 1.08 kg ha−1. The findings of this study could assist in understanding the factors affecting the source, production, and sink of these two important GHGs. IWD, along with judicious N-based fertilizer use, could provide significant respite from GHG emissions in rice-based agriculture. These climate-smart strategies not only reduce emissions but also have the potential to improve yield.

METHANE OXIDATION AND BIOGEOCHEMICAL PARAMETERS OF GRAY FOREST SOIL AS INFLUENCES BY FALLOW-TO-WOOD SUCCESSION

The succession of microbial community structure and function is a central ecological topic, as microbes drive the Earths biogeochemical cycles and are responsible for the regulation of the gaseous composition of the atmosphere, the processes of soils formation and their resistance to natural and anthropogenic factors. To elucidate the response and mechanistic underpinnings of soil methane-oxidizing microbial community structure and metabolic potential relevant to natural forest succession on gray forest soils of Moscow region from farming through the fallows of different ages (5-25 years) to a forest biocenosis was studied. Highest methane oxidation was observed from older restored sites its recovery with restoration age. Molecular analyses indicate the presence of aerobic type I and type II groups of methanotrophs in arable and meadow sites and only type II methanotophs in forested sites. Collectively, these data indicate shifts in microbial populations associated with methane oxidation in the context of soil restoration age and provide significant insights into the response of these microbial populations to severe disturbance and recovery.

Addendum: The rise and fall of methanotrophy following a deepwater oil-well blowout

Addendum: The rise and fall of methanotrophy following a deepwater oil-well blowout

Highly efficient methane assimilation through Embden-Meyerhof-Parnas pathway in Methylomicrobium alcaliphilum 20Z

In order to understand metabolic functions essential for methane assimilation, we investigate dribulose monophosphate pathway and adjacent pathways in gammaproteobacterial Methylomicrobium alcaliphilum 20Z by using combined approaches of RNA-seq, LC-MS, and 13C-labeled techniques. The absolute quantification of metabolome showed that the concentrations of intermediates, such as glucose-6-phosphate and 2-dehydro-3-deoxy-phosphogluconate, involved in Entner-Doudoroff (EDD) pathway were (150.95 +/- 28.75) micromol/L and below the limit of detection of mass spectrometry. In contrast, fructose-1, 6-bisphosphate, glyceraldehyde-3-phosphate/dihydroxyacetone and phosphoenolpyruvate in Embden-Meyerhof-Parnas (EMP) pathway had significantly higher concentrations with (1 142.02 +/- 302.88) micromol/L, (1 866.76 +/- 388.55) micromol/L and (3 067.57 +/- 898.13) micromol/L, respectively. 13C-labeling experiment further indicated that the enrichment of [3-13C1]-pyruvate involved in EMP pathway was 4-6 fold higher than [1,13C1]-pyruvate in EDD pathway in a dynamic course. Moreover, gene expression profile showed that the expression levels of genes in EMP pathway (e.g. fbaA, tpiA, gap and pykA) were 2 479.2, 2 493.9, 2 274.6 and 1 846.0, respectively, but gene expressionlevels in EDD pathway (e.g. pgi, eda and edd) were only 263.8, 341.2 and 225.4, respectively. Overall our current results demonstrated that EMP pathway was the main route for methane assimilation in M. alcaliphilum 20Z. This discovery challenged our understanding of methane assimilation pathway in gammaproteobacterial methanotrophic bacteria, and further provided an important insight for efficient methane biocatalysis in the future.

Methanotrophy and methylotrophy

Methanotrophy and methylotrophy

Poly-3-hydroxybutyrate production from methanol by Methylosinus trichosporium IMV3011 in the non-sterilized fed-batch fermentation

The methanotrophic bacteria have promising applications in the production of a biopolymer, poly-3-hydroxybutyrate (PHB). The fed-batch fermentation strategies were studied for PHB accumulation by methanol-grown Methylosinus trichosporium IMV3011 under the non-sterilized conditions. The biomass of 2.91 g dry wt/L and PHB accumulation of 47.6% (w/w) were reached in the fed-batch fermentation. In order to increase PHB content, some organic acids were added into the culture as inhibitors of tricarboxylic acid cycle (TCA). It was found that the malic acid should be optimum for PHB accumulation. The biomass and PHB yield would increase to 3.32 g dry wt/L and 58.5% (w/w), respectively by adding 0.2 g/L malic acid. Other key intermediates in the metabolism such as citric acid, succinic acid and acetyl-CoA, also influenced the cell growth and PHB accumulation to a certain extent. The PHB produced in the fermentation process was of steady quality with molecular weights up to 1.7 × 106 Da. Key words: Biosynthesis, poly-3-hydroxybutyrate, methanotrophic bacteria, methanol, fed-batch fermentation.

Advances of Intact Polar Membrane Lipids as Chemical Biomarkers for Extant Microorganisms in Marine Sediments

Intact polar membrane lipids(IPLs)can serve as chemical biomarkers of living cells that can provide information on the extant microbial community structure and in situ microbial processes.IPLs analysis is a culture independent,rapid and non-selective procedure in contrast to biological methods.The compositions of cell membrane intact polar membrane lipids of bacteria and Achaea and their applications in the researches of biogeochemistry and microbial ecology are summarized in this paper.The recent progresses of IPLs applications are reviewed,with emphasis on those microorganisms that play a specific role in biogeochemical cycle,such as anaerobic ammonium-oxidizing bacteria,anaerobic methanotrophic Achaea,ammonia-oxidizing Achaea and marine Crenarcheota with tetraether lipids,or those special ecological systems,such as hydrothermal vent,cold seep and marine deep biosphere.The advances of analytical method for microbial IPLs in marine sediments are briefly introduced,and their application prospects are proposed.

Nickel-alkyl bond formation in the active site of methyl-coenzyme m reductase.

Methyl-coenzyme M reductase (MCR) catalyzes the methane-forming step in methanogenic archaea and most probably also the methane-oxidizing step in methanotrophic archaea. The enzyme contains coenzyme F(430) as prosthetic group. F(430) is a nickel porphinoid that has to be in the reduced Ni(I) state for the enzyme to be active. The presently discussed catalytic mechanisms of MCR can in principle be divided into two basic models. In one model the key intermediate features a methyl-Ni(III) species being either formed in a nucleophilic substitution reaction or in an oxidative addition reaction. In the other model first the thioether sulfur of methyl-coenzyme M binds to the Ni(I), which subsequently results in the release of the methyl group as methyl radical leaving behind a Ni(II)-sulfur bond. The experimental evidence for and against a methyl-nickel intermediate is reviewed.

Methane oxidation in slurry storage surface crusts.

Livestock manure is a significant source of atmospheric methane (CH4), especially during liquid storage. In liquid manure (slurry) storages a surface crust may form naturally, or an artificial surface crust can be established. We investigated whether there is a potential for CH4 oxidation in this environment. Surface crust materials were sampled from experimental storages with cattle slurry (with natural crust) or anaerobically digested cattle slurry (with straw layer) that had been stored with or without a wooden cover. Extracts of surface crust material were incubated with 5.6% CH4 in the headspace, and methanotrophic activity was demonstrated in all four treatments following a 4- to 10-d lag phase. Subsequent incubation of field-moist surface crust material with 350 microL L(-1) CH4 also showed CH4 oxidation, indicating a potential for CH4 removal under practical storage conditions. There was no CH4 oxidation activity during incubation of autoclaved samples. Methane oxidation rates were 0.1 to 0.5 mg kg(-1) organic matter (OM) h(-1), which is comparable with the activity in wetlands and rice paddies. Partial drying increased CH4 oxidation to 0.2 to 1.4 mg kg(-1) OM h(-1), probably as a result of improved diffusivity within the surface crust. Rewetting reversed the stimulation of methanotrophic activity in some treatments, but not in others, possibly due to a decline in CH4 production in anaerobic volumes, or to growth of methanotrophs during incubation. This study presents direct evidence for methanotrophic activity in slurry storages. Measures to ensure crust formation with or without a solid cover appear to be a cost-effective greenhouse gas mitigation option.

Kinetic Study of Trichloroethylene Biodegradation by Methylosinus trichosporium OB3b PP358 lmmobilized in a Fibrous-Bed Bioreactor

Biodegradation of trichloroethylene (TCE) by resting cells of methanotrophic Methylosinus trichosporium OB3b PP358, which constitutively expresses soluble methane monooxygenase (sMMO), was studied in a fibrous-bed bioreactor operated in the recycle batch mode. Cells were grown on methane as the substrate with aeration, and then used to degrade TCE through the cometabolism with sMMO in the absence of methane. Complete biodegradation of TCE was verified with the TCE and chloride ion mass balance. In general, concentration of up to ~12 mg/l, and followed the first-order reaction kinetics. TCE transformation was inhibited in the presence of methane or methanol. Without the energy source and after being exposed to TCE for an extended period, cells gradually lost most of their capability in degrading TCE, which was attributed to reduced sMMO enzyme activity due to lack of NADH and cell death caused by TCE toxicity and oxygen starvation. However, the reactor was able to recover its TCE degradation ability after rejuvenating and regrowing cells with methane and air. Compared to free cell and other immobilized cell systems, the cells immobilized in the fibrous-bed bioreactor not only showed a much higher TCE degradation rate (up to 84.77 mg/(1 day) or ~32 times of that from free cells), but also had a better tolerance to TCE (22.6 mg/l or ~11 times higher than that with free cells). With periodical rejuvenation, the bioreactor could be used for long-term treatment of TCE-contaminated water.

The effect of oxygen on methanol oxidation by an obligate methanotrophic bacterium studied by in vivo (13)C nuclear magnetic resonance spectroscopy.

13C NMR was used to study the effect of oxygen on methanol oxidation by a type II methanotrophic bacterium isolated from a bioreactor in which methane was used as electron donor for denitrification. Under high (35-25%) oxygen conditions the first step of methanol oxidation to formaldehyde was much faster than the following conversions to formate and carbon dioxide. Due to this the accumulation of formaldehyde led to a poisoning of the cells. A more balanced conversion of (13)C-labelled methanol to carbon dioxide was observed at low (1-5%) oxygen concentrations. In this case, formaldehyde was slowly converted to formate and carbon dioxide. Formaldehyde did not accumulate to inhibitory levels. The oxygen-dependent formation of formaldehyde and formate from methanol is discussed kinetically and thermodynamically.

Characterization of Microbial Consortia in Paddy Rice Soil by Phospholipid Analysis.

Microbial biomass and community structure in paddy rice soil during the vegetation period of rice were estimated by analysis of their phospholipid fatty acids (PLFA), hydroxy fatty acids of lipopolysaccharides (LPS-HYFA), and phospholipid ether lipids (PLEL) directly extracted from the soil. A clear change in the composition of the community structure at different sampling periods was observed, indicated by the principal component analysis of the PLFA. A dramatic decline of ester-linked PLFA was observed in the soil samples taken at the second sampling time. In contrast to the ester-linked PLFA, the non-ester-linked PLFA composition did not change. The hydroxy fatty acids of lipopolysaccharides as well as ether lipids decreased consecutively during the observation period. Total microbial abundance was estimated to be (4.1-7.3) x 10(9) cells g(-1) soil (dry weight). About 44% account for aerobic and 32% for facultative anaerobic bacteria, and 24% for archaea, on average. According to the profile and patterns of PLFA in the soil sample, it may be suggested that the paddy soil at the August sampling period contained more abundant facultative anaerobic bacteria (ca. 36%) and archaea (ca. 37%), but the total microbial biomass was significantly lower than in the remaining sampling periods. As the plant approached maturity, the microbial community structure in the soil changed to contain more abundant Gram-negative bacteria and methanotrophs.