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PubMed1981

The influence of pentose on breath methane.

In a population study the concentration of methane in the expired breath of methane-producing subjects was significantly related to the dietary intake of noncellulosic polysaccharide pentose (p less than 0.01) and to the intake of lignin (p less than 0.05). The single dose administration of complex polysaccharide sources rich in pentoses to fasting subjects did not result in an increase in methane production when measured over 5 h. The single dose administration of D(+) xylose and L(+) arabinose led to a significant increase in methane excretion in methane producers.

Adolescent↗

PubMed1993

Interaction between methane-producing status and diet on serum acetate concentration in humans.

About half the population excretes methane in the breath. To see if methane producing status influenced serum acetate, we studied six methane producers and six nonproducers on three separate days. For 36 h they ate a polysaccharide-free diet alone, or with 20 g of unabsorbed sugar lactulose, or 20 g of fermentable fiber, guar, in random order. The mean fasting serum acetate concentration on the three test days in producers was higher than in nonproducers, 84 +/- 5 vs. 69 +/- 5 mumol/L (P < 0.05). Compared to the control diet, both lactulose and guar raised serum acetate concentration significantly in both groups of subjects. However, there was a significant interaction between methane producing status and diet. After lactulose consumption, postprandial serum acetate was similar in both groups of subjects, but guar consumption had a significantly greater effect in producers than nonproducers (98 +/- 8 vs. 73 +/- 5 mumol/L; P < 0.05). We conclude that methane producing status may influence serum acetate concentrations in humans, depending upon the type of fermentable carbohydrate in the diet.

Acetates↗

PubMed2000

Different substrates and methane producing status affect short-chain fatty acid profiles produced by In vitro fermentation of human feces.

Five different substrates, i.e., lactulose, rhamnose, cornstarch, guar and ileostomy effluent, were used to determine whether methane producing status alters the production of short-chain fatty acids (SCFA) in methane producers (MP; n = 6) and nonproducers (MNP; n = 5). Fecal samples from MP and MNP were fermented with the five substrates using an in vitro fermentation method. Subjects with a mean breath methane concentration > 0.045 micromol/L above ambient air were classified as MP. Fermentation was stopped and samples were obtained at 3, 5 and 24 h. An HPLC method was used to measure the SCFA, acetate, propionate, isobutyrate, butyrate, valerate and isocaproate. A significant interaction between methane producing status and time for acetate production from lactulose was observed. There were no differences in fermentation of the four remaining substrates between MP and MNP, but there were significant differences among substrates in the two groups combined. Acetate production from lactulose was significantly greater than from the four other substrates, whereas that from ileostomy effluent was significantly less than the four other substrates. The amount of propionate produced from rhamnose was significantly higher than from the other substrates. The amount of butyrate produced from lactulose and cornstarch was significantly higher than from the other substrates. We conclude that differences exist in the fermentation patterns of lactulose, rhamnose, cornstarch, guar and ileostomy effluent. Methane producing status may influence fermentation patterns only of substrates that are largely fermented to acetate and not others.

Adult↗

PubMed1985

The development of methane production in childhood and adolescence.

Methane concentration in the expired air was analyzed in 393 infants, children, and adolescents. There was no methane production below the age of 3 years. In the age group 3-4 years, 6.4% of the children produced methane. The percentages were 14.3-18.2% in children aged 7-14. A rise in methane production to 39.4-45.9% was recorded from age 14 to 18, as compared with 49.4% in adults in Israel. The reasons for these changes in methane production with age are not known. Alterations in the colonic flora or the production of a specific substrate(s) are the main possibilities. In our opinion, the latter possibility deserves further investigation.

Adolescent↗

PubMed2000

Methylocella palustris gen. nov., sp. nov., a new methane-oxidizing acidophilic bacterium from peat bogs, representing a novel subtype of serine-pathway methanotrophs.

A new genus, Methylocella, and a new species, Methylocella palustris, are proposed for three strains of methane-oxidizing bacteria isolated from acidic Sphagnum peat bogs. These bacteria are aerobic, Gram-negative, colourless, non-motile, straight and curved rods that utilize the serine pathway for carbon assimilation, multiply by normal cell division and contain intracellular poly-beta-hydroxybutyrate granules (one at each pole). These strains use methane and methanol as sole sources of carbon and energy and are moderately acidophilic organisms with growth between pH 4.5 and pH 7.0, the optimum being at pH 5.0-5.5. The temperature range for growth is 10-28 degrees C with the optimum at 15-20 degrees C. The intracytoplasmic membrane system is different from those of type I and II methanotrophs. Cells contain an extensive periplasmic space and a vesicular membrane system connected to the cytoplasmic membrane. The strains grew only on media with a low salt content (0.2-0.5 g l(-1)). All three strains were found to possess soluble methane monooxygenase and are able to fix atmospheric nitrogen via an oxygen-sensitive nitrogenase. No products were observed in a PCR with particulate methane monooxygenase-targeted primers; hybridization with a pmoA probe was also negative. The major phospholipid fatty acids are 18:1 acids. The G+C content of the DNA is 61.2 mol%. The three strains share identical 16S rRNA gene sequences and represent a novel lineage of methane-oxidizing bacteria within the alpha-subclass of the class Proteobacteria and are only moderately related to type II methanotrophs of the Methylocystis-Methylosinus group. The three strains are most closely related to the acidophilic heterotrophic bacterium Beijerinckia indica subsp. indica (96.5% 16S rDNA sequence similarity). Collectively, these strains comprise a new species and genus Methylocella palustris gen. nov., sp. nov.; strain KT (= ATCC 700799T) is the type strain.

Base Composition↗

PubMed1994

Further evidence for multiple pathways in soluble methane-monooxygenase-catalysed oxidations from the measurement of deuterium kinetic isotope effects.

The data from the deuterium isotope experiments in this study show that the primary kinetic isotope effect for methane oxidation catalysed by soluble methane monooxygenase from Methylococcus capsulatus (Bath) is very small, < 2. In contrast, the primary kinetic isotope effect for -CH3 group oxidation in toluene is large, > 7. A mechanistic pathway in which a substrate radical is formed from hydrogen atom abstraction by a ferryl species is believed to operate for CH4, the toluene -CH3 group and similar alkanes. Direct oxygen atom addition, rather than H atom abstraction, is indicated for aromatic ring oxidations in benzene and toluene and for styrene oxide formation from styrene. Thus, more than one mechanistic pathway appears to operate in soluble methane-monooxygenase-catalysed reactions and, in some cases, the pathway chosen may be dictated by the substrate. In the soluble methane-monooxygenase-catalysed oxidation of toluene the rates of: (a) substrate dissociation from the enzyme-substrate complex, (b) product formation and (c) product release (benzyl alcohol and p-cresol) from the enzyme-product complex are comparable in magnitude. Therefore all three of these steps are partially rate-determining in the soluble methane monooxygenase catalytic cycle for toluene oxidation.

Benzene↗

PubMed2004

Inhibition of ruminal microbial methane production by beta-cyclodextrin iodopropane, malate and their combination in vitro.

The objective of this study was to evaluate the effects of different concentrations of l-malate (0, 5, 10 and 20 mm), 2-iodopropane-beta-cyclodextrin complex (CD-IP) (0, 0.1, 0.2 and 0.4 mm) and a combination of malate (10 and 20 mm) plus CD-IP (0.2 and 0.4 mm) on methane production from corn starch. Ruminal fluid was collected from dairy cows, mixed with phosphate buffer (1 : 2) and incubated (30 ml) anaerobically at 38 degrees C for 6 h with or without additives. Fermentation of corn starch in the presence of malate resulted in an increase (p < 0.05) in pH of the medium, total volatile fatty acid (VFA), total gas production and molar proportion of propionate. Acetate and ammonia-N concentration were unchanged. Methane production was decreased (p < 0.05) (15.5 to 20.4%). Addition of CD-IP in corn starch resulted in an increase (p < 0.05) in total VFA and molar proportion of propionate. Acetate, pH and ammonia-N concentration of the medium were decreased (p < 0.05). Total gas production was unchanged. Methane production was decreased (p < 0.05) (25.2 to 97.1%) and hydrogen production was increased (p < 0.05). Addition of l-malate to CD-IP resulted in an increase (p < 0.05) in total VFA, total gas production and molar proportion of propionate. Acetate and ammonia-N concentration were decreased (p < 0.05). No effects were observed on medium pH. Methane production was decreased (p < 0.05) (49.5 to 97.1%). Hydrogen production was also decreased (p < 0.05) (54.5 to 64.1%) compared with those of CD-IP alone. Therefore, these additives may be used as supplements to inhibit methane production as well as to improve rumen fermentation and animal performance.

Animals↗

PubMed2006

Microbiological investigation of methane- and hydrocarbon-discharging mud volcanoes in the Carpathian Mountains, Romania.

Paclele Mici is a terrestrial mud volcano field located in the Carpathian Mountains (Romania), where thermal alteration of sedimentary organic compounds leads to methane, higher hydrocarbons and other petroleum compounds that are continuously released into the environment. The hydrocarbons represent potential substrates for microorganisms. We studied lipid biomarkers, stable isotope ratios, the effect of substrate (methane, other organic compounds) addition and 16S rRNA genes to gain insights into the hitherto unknown microbial community at this site. Quantitative real-time polymerase chain reaction analysis demonstrated that bacteria were much more abundant than archaea. Phylogenetic analyses of 16S rDNA clone sequences indicated the presence of bacterial and archaeal lineages generally associated with the methane cycle (methanogens, aerobic and anaerobic methanotrophs), the sulfur cycle (sulfate reducers), and groups linked to the anaerobic degradation of alkanes or aromatic hydrocarbons. The presence of sulfate reducers, methanogens and methanotrophs in this habitat was also confirmed by concurrent surveys of lipid biomarkers and their isotopic signatures. Incubation experiments with several common and complex substrates revealed the potential of the indigenous microbial community for sulfate reduction, methanogenesis and aerobic methanotrophy. Additionally, consistently to the detection of methane-oxidizing archaea (ANME) and 13C-depleted archaeal lipids, a weak but significant activity of anaerobic methane oxidation was measured by radiotracer techniques and in vitro. This survey is the first to report the presence and activity of ANME in a terrestrial environment.

Archaea↗

PubMed2007

In vitro cell growth of marine archaeal-bacterial consortia during anaerobic oxidation of methane with sulfate.

Anoxic sediment from a methane hydrate area (Hydrate Ridge, north-east Pacific; water depth 780 m) was incubated in a long-term laboratory experiment with semi-continuous supply of pressurized [1.4 MPa (14 atm)] methane and sulfate to attempt in vitro propagation of the indigenous consortia of archaea (ANME-2) and bacteria (DSS, Desulfosarcina/Desulfococcus cluster) to which anaerobic oxidation of methane (AOM) with sulfate has been attributed. During 24 months of incubation, the rate of AOM (measured as methane-dependent sulfide formation) increased from 20 to 230 micromol day(-1) (g sediment dry weight)(-1) and the number of aggregates (determined by microscopic counts) from 0.5 x 10(8) to 5.7 x 10(8) (g sediment dry weight)(-1). Fluorescence in situ hybridization targeting 16S rRNA of both partners showed that the newly grown consortia contained central archaeal clusters and peripheral bacterial layers, both with the same morphology and phylogenetic affiliation as in the original sediment. The development of the AOM rate and the total consortia biovolume over time indicated that the consortia grew with a doubling time of approximately 7 months (growth rate 0.003 day(-1)) under the given conditions. The molar growth yield of AOM was approximately 0.6 g cell dry weight (mol CH(4) oxidized)(-1); according to this, only 1% of the consumed methane is channelled into synthesis of consortia biomass. Concentrations of biomarker lipids previously attributed to ANME-2 archaea (e.g. sn-2-hydroxyarchaeol, archaeol, crocetane, pentamethylicosatriene) and Desulfosarcina-like bacteria [e.g. hexadecenoic-11 acid (16:1omega5c), 11,12-methylene-hexadecanoic acid (cy17:0omega5,6)] strongly increased over time (some of them over-proportionally to consortia biovolume), suggesting that they are useful biomarkers to detect active anaerobic methanotrophic consortia in sediments.

Anaerobiosis↗

PubMed2006

Methane and sulfate profiles within the subsurface of a tidal flat are reflected by the distribution of sulfate-reducing bacteria and methanogenic archaea.

The anoxic layers of marine sediments are dominated by sulfate reduction and methanogenesis as the main terminal oxidation processes. The aim of this study was to analyze the vertical succession of microbial populations involved in these processes along the first 4.5 m of a tidal-flat sediment. Therefore, a quantitative PCR approach was applied using primers targeting the domains of Bacteria and Archaea, and key functional genes for sulfate reduction (dsrA) and methanogenesis (mcrA). The sampling site was characterized by an unusual sulfate peak at 250 cm depth resulting in separate sulfate-methane transition zones. Methane and sulfate profiles were diametrically opposed, with a methane maximum in the sulfate-depleted zone showing high numbers of archaea and methanogens. The methane-sulfate interfaces harbored elevated numbers of sulfate reducers, and revealed a slight increase in mcrA and archaeal 16S rRNA genes, suggesting sulfate-dependent anaerobic oxidation of methane. A diversity analysis of both functional genes by PCR-denaturing gradient gel electrophoresis revealed a vertical succession of subpopulations that were governed by geochemical and sedimentologic conditions. Along the upper 200 cm, sulfate-reducing populations appeared quite uniform and were dominated by the Deltaproteobacteria. In the layers beneath, an apparent increase in diversity and a shift to the Firmicutes as the predominant group was observed.

Anaerobiosis↗

PubMed1986

Seasonal variations in the stable carbon isotopic signature of biogenic methane in a coastal sediment.

Systematic seasonal variations in the stable carbon isotopic signature of methane gas occur in the anoxic sediments of Cape Lookout Bight, a lagoonal basin on North Carolina's Outer Banks. Values for the carbon isotope ratio (delta 13C) of methane range from -57.3 per mil during summer to -68.5 per mil during winter in gas bubbles with an average methane content of 95%. The variations are hypothesized to result from changes in the pathways of microbial methane production and cycling of key substrates including acetate and hydrogen. The use of stable isotopic signatures to investigate the global methane cycle through mass balance calculations, involving various sediment and soil biogenic sources, appears to require seasonally averaged data from individual sites.

Anaerobiosis↗

PubMed1977

Thermophilic methane production from cattle waste.

Methane production from waste of cattle fed a finishing diet was investigated, using four 3-liter-working volume anaerobic digestors at 60 degrees C. At 55 degrees C a start-up culture, in which waste was the only source of bacteria, was generated within 8 days and readily adapted to 60 degrees C, where efficiency of methanogenesis was greater. Increasing the temperature from 60 to 65 degrees C tended to drastically lower efficiency. When feed concentrations of volatile solids (VS, organic matter) were increased in steps of 2% after holding for 1 months at a given concentration, the maximum concentrations for efficient fermentation were 8.2, 10.0, 11.6, and 11.6% for the retention times (RT) of 3, 6, 9, and 12 days, respectively. The VS destructions for these and lower feed concentrations were 31 to 37, 36 to 40, 47 to 49 and 51 to 53% for the 3-, 6-, 9-, and 12-day RT digestors, respectively, and the corresponding methane production rates were about 0.16, 0.18, 0.20, and 0.22 liters/day per g of VS in the feed. Gas contained 52 to 57% methane. At the above RT and feed concentrations, alkalinity rose to 5,000 to 7,700 mg of CaCo3 per liter (pH to 7.5 to 7.8), NH3 plus NH4+ to 64 to 90 mM, and total volatile acids to 850 to 2,050 mg/liter as acetate. The 3-day RT digestor was quite stable up to 8.2% feed VS and at this feed concentration produced methane at the very high rate of 4.5 liters/day per liter of digestor. Increasing the percentage of feed VS beyond those values indicated above resulted in greatly decreased organic matter destruction and methane production, variable decrease in pH, and increased alkalinity, ammonia, and total volatile acid concentrations, with propionate being the first to accumulate in large amounts. In a second experiment with another lot of waste, the results were similar. These studies indicate that loading rates can be much higher than those previously thought useful for maximizing methanogenesis from cattle waste.

Anaerobiosis↗

PubMed1978

Changes in proportions of acetate and carbon dioxide used as methane precursors during the anaerobic digestion of bovine waste.

In an anaerobic digestor which was fed daily with bovine waste, during the early stages after feeding (4 to 7 h) acetate (via the methyl group) accounted for almost 90% of the methane produced. As time after feeding increased, acetate declined as a precursor so that in the 12- to 14-h and 21- to 23-h periods, after feeding the methyl group accounted for 80 and 73% of the methane produced, respectively. Measurements of methane production from CO2 reduction showed that in the 2- to 12-h period after feeding, CO2 accounted for 14% of the methane produced, whereas in the 12- to 24-h period it accounted for 27-5%. These results show that the percentages of methane accounted for by acetate and CO2 vary with time after feeding the digestor.

Acetates↗

PubMed1997

Transformation yields of chlorinated ethenes by a methanotrophic mixed culture expressing particulate methane monooxygenase.

Transformation yields for the aerobic cometabolic degradation of five chlorinated ethenes were determined by using a methanotrophic mixed culture expressing particulate methane monooxygenase (pMMO). Transformation yields (expressed as moles of chlorinated ethene degraded per mole of methane consumed) were 0.57, 0.25, 0.058, 0.0019, and 0.00022 for trans-1,2-dichloroethylene (t-DCE), vinyl chloride (VC), cis-1,2-dichloroethylene (c-DCE), trichloroethylene (TCE), and 1,1-dichloroethylene (1,1-DCE), respectively. Degradation of t-DCE and VC was observed only in the presence of formate or methane, sources of reducing energy necessary for cometabolism. The t-DCE and VC transformation yields represented 35 and 15%, respectively, of the theoretical maximum yields, based on reducing-energy availability from methane dissimilation to carbon dioxide, exclusive of all other processes that require reducing energy. The yields for t-DCE and VC were 20 times greater than the yields reported by others for cells expressing soluble methane monooxygenase (sMMO). Transformation yields for c-DCE, TCE, and 1,1-DCE were similar to or less than those for cultures expressing sMMO. Although methanotrophic biotreatment systems have typically been designed to incorporate cultures expressing sMMO, these results suggest that pMMO expression may be highly advantageous for degradation of t-DCE or VC. It may also be much easier to maintain pMMO expression in treatment systems, because pMMO is expressed by all methanotrophs whereas sMMO is expressed only by type II methanotrophs under copper-limited conditions.

Bacteria↗

PubMed2000

Biomarker evidence for widespread anaerobic methane oxidation in Mediterranean sediments by a consortium of methanogenic archaea and bacteria. The Medinaut Shipboard Scientific Party.

Although abundant geochemical data indicate that anaerobic methane oxidation occurs in marine sediments, the linkage to specific microorganisms remains unclear. In order to examine processes of methane consumption and oxidation, sediment samples from mud volcanoes at two distinct sites on the Mediterranean Ridge were collected via the submersible Nautile. Geochemical data strongly indicate that methane is oxidized under anaerobic conditions, and compound-specific carbon isotope analyses indicate that this reaction is facilitated by a consortium of archaea and bacteria. Specifically, these methane-rich sediments contain high abundances of methanogen-specific biomarkers that are significantly depleted in (13)C (delta(13)C values are as low as -95 per thousand). Biomarkers inferred to derive from sulfate-reducing bacteria and other heterotrophic bacteria are similarly depleted. Consistent with previous work, such depletion can be explained by consumption of (13)C-depleted methane by methanogens operating in reverse and as part a consortium of organisms in which sulfate serves as the terminal electron acceptor. Moreover, our results indicate that this process is widespread in Mediterranean mud volcanoes and in some localized settings is the predominant microbiological process.

Anaerobiosis↗

PubMed2000

Response of atmospheric methane consumption by maine forest soils to exogenous aluminum salts.

Atmospheric methane consumption by Maine forest soils was inhibited by additions of environmentally relevant levels of aluminum. Aluminum chloride was more inhibitory than nitrate or sulfate salts, but its effect was comparable to that of a chelated form of aluminum. Inhibition could be explained in part by the lower soil pH values which resulted from aluminum addition. However, significantly greater inhibition by aluminum than by mineral acids at equivalent soil pH values indicated that inhibition also resulted from direct effects of aluminum per se. The extent of inhibition by exogenous aluminum increased with increasing methane concentration for soils incubated in vitro. At methane concentrations of >10 ppm, inhibition could be observed when aluminum chloride was added at concentrations as low as 10 nmol g (fresh weight) of soil(-1). These results suggest that widespread acidification of soils and aluminum mobilization due to acid precipitation may exacerbate inhibition of atmospheric methane consumption due to changes in other parameters and increase the contribution of methane to global warming.

Aluminum Compounds↗

PubMed2005

Cooccurrence of aerobic and anaerobic methane oxidation in the water column of Lake Plusssee.

Dissolved methane was investigated in the water column of eutrophic Lake Plusssee and compared to temperature, oxygen, and sulfide profiles. Methane concentrations and delta-13C signatures indicated a zone of aerobic methane oxidation and additionally a zone of anaerobic methane oxidation in the anoxic water body. The latter coincided with a peak in hydrogen sulfide concentration. High cell numbers of aerobic and anaerobic methane-oxidizing microorganisms were detected by fluorescence in situ hybridization (FISH) or the more sensitive catalyst-amplified reporter deposition-FISH, respectively, in these layers.

Aerobiosis↗

PubMed1995

Organization and growth phase-dependent transcription of methane genes in two regions of the Methanobacterium thermoautotrophicum genome.

Two regions of the Methanobacterium thermoautotrophicum genome containing genes that encode enzymes involved in methanogenesis (methane genes) have been cloned and sequenced to determine the extent of methane gene clustering and conservation. One region from the M. thermoautotrophicum strains delta H and Winter, extending approximately 13.5 kb upstream from the adjacent mvhDGAB and mrtBDGA operons that encode the methyl-viologen-reducing hydrogenase (MVH) and the methyl coenzyme M reductase II (MRII), respectively, was sequenced, and 76% sequence identity and very similar gene organizations were demonstrated. Five closely linked open reading frames were located immediately upstream of the mvh operon and were designated flpECBDA. The flpCBD genes encode amino acid sequences that are 31, 47, and 65% identical to the primary sequences of the alpha and beta subunits of formate dehydrogenase and the delta subunit of MVH, respectively. Located immediately upstream of the flp genes was the mth gene, which encodes the H2-dependent methylene-tetrahydromethanopterin dehydrogenase (MTH). In contrast to this mth-flp-mvh-mrt cluster of methane genes, a separate approximately 5.4-kb genomic fragment cloned from M. thermoautotrophicum delta H contained only one methane gene, the mtd gene, which encodes the 8-hydroxy-5-deazaflavin (H2F420)-dependent methylene-tetrahydromethanopterin dehydrogenase (MTD). Northern (RNA) blot experiments demonstrated that mth was transcribed only at early growth stages in fermentor-grown cultures of M. thermoautotrophicum delta H, whereas mtd was transcribed at later growth stages and in the stationary phase. Very similar transcription patterns have been observed by T.D. Pihl, S. Sharma, and J. N. Reeve (J. Bacteriol. 176:6384-6391, 1994) for the MRI- and MRII-encoding operons, mrtBDGA and mcrBDCGA, im M. thermoautotrophicum deltaH, suggesting coordinated regulation of methane gene expression. In contrast to the growth phase-dependent transcription of the mth/mrt and mtd/mcr genes, transcription of the mvhDGAB and frhADGB operons, which encode the two (NiFe) hydrogenases in M. thermoautotrophicum deltaH, was found to occur at all growth stages.

Amino Acid Sequence↗