Methanogenic Sediments Research Articles

Successional Changes in an Evolving Anaerobic Chlorophenol-Degrading Community Used To Infer Relationships between Population Structure and System-Level Processes

The response of a complex methanogenic sediment community to 2-chlorophenol (2-CP) was evaluated by monitoring the concentrations of this model contaminant and important metabolic intermediates and products and by using rRNA-targeted probes to track several microbial populations. Key relationships between the evolving population structure, formation of metabolic intermediates, and contaminant mineralization were identified. The nature of these relationships was intrinsically linked to the metabolism of benzoate, an intermediate that transiently accumulated during the mineralization of 2-CP. Before the onset of benzoate fermentation, reductive dehalogenation of 2-CP competed with methanogenesis for endogenous reducing equivalents. This suppressed H(2) levels, methane production, and archaeal small-subunit (SSU)-rRNA concentrations in the sediment community. The concentrations of bacterial SSU rRNA, including SSU rRNA derived from "Desulfovibrionaceae" populations, tracked with 2-CP levels, presumably reflecting changes in the activity of dehalogenating organisms. After the onset of benzoate fermentation, the abundance of Syntrophus-like SSU rRNA increased, presumably because these syntrophic organisms fermented benzoate to methanogenic substrates. Consequently, although the parent substrate 2-CP served as an electron acceptor, cleavage of its aromatic nucleus also influenced the sediment community by releasing the electron donors H(2) and acetate. Increased methane production and archaeal SSU-rRNA levels, which tracked with the Syntrophus-like SSU-rRNA concentrations, revealed that methanogenic populations in particular benefited from the input of reducing equivalents derived from 2-CP.

Reductive dehalogenation and conversion of 2-chlorophenol to 3-chlorobenzoate in a methanogenic sediment community: implications for predicting the environmental fate of chlorinated pollutants.

Biotransformation of 2-chlorophenol by a methanogenic sediment community resulted in the transient accumulation of phenol and benzoate. 3-Chlorobenzoate was a more persistent product of 2-chlorophenol metabolism. The anaerobic biotransformation of phenol to benzoate presumably occurred via para-carboxylation and dehydroxylation reactions, which may also explain the observed conversion of 2-chlorophenol to 3-chlorobenzoate.

Methane As a Product of Chloroethene Biodegradation under Methanogenic Conditions

Radiometric detection headspace analyses of microcosms containing bed sediments from two geographically distinct sites indicated that 10−39% of the radiolabeled carbon transformed during anaerobic biodegradation of [1,2-14C]trichloroethene (TCE) or [1,2-14C]vinyl chloride (VC) under methanogenic conditions was ultimately incorporated into 14CH4. The results demonstrate that, in addition to ethene, ethane, and CO2, CH4 can be a significant product of chloroethene biodegradation in some methanogenic sediments.

Enhanced anaerobic benzene degradation with the addition of sulfate

Potential mechanisms for stimulating anaerobic benzene degradation in methanogenic sediments from a petroleum‐contaminated aquifer were evaluated. In short‐term (<2 weeks) incubations, addition of sulfate slightly stimulated benzene degradation and caused a small decrease in the ratio of methane to carbon dioxide production from benzene. However, in longer‐term (>100 days) incubations, sulfate significantly stimulated benzene degradation with a complete shift to carbon dioxide as the end product of benzene degradation. The addition of Fe(III) and humic substances had short‐term and long‐term effects that were similar to the effects of the sulfate amendments. In studies in which anaerobic groundwater was pumped through columns of aquifer sediments, addition of sulfate to the groundwater significantly enhanced the removal of benzene from the groundwater. The stoichiometry of sulfate and benzene removal from the groundwater passing through the sediment columns was consistent with benzene oxidation to carbon dioxide with sulfate serving as the primary electron acceptor. These results demonstrate for the first time that addition of sulfate may be an effective strategy for enhancing anaerobic benzene removal in some petroleum‐contaminated aquifers.

Rapid Benzene Degradation in Methanogenic Sediments from a Petroleum-Contaminated Aquifer

In methanogenic sediments from a petroleum-contaminated aquifer, [14C]benzene was converted to 14CH4 and 14CO2 without an apparent lag. Phenol, acetate, and propionate were intermediates in benzene mineralization. These results suggest that alternative electron acceptors need not be available for there to be significant natural attenuation of benzene in some petroleum-contaminated aquifers.

Evidence for Isotopic Exchange during Metabolism of Stable-Isotope-Labeled Formate in a Methanogenic Sediment

The disappearance of stable-isotope-labeled formate in freshwater sediment from Lake Vechten was studied by gas chromatography-mass spectrometry. Both deuterium- and (sup13)C-labeled formate were removed from pore water at a high rate (400 (plusmn) 35 [standard deviation] and 1,700 (plusmn) 600 (mu)M h(sup-1), respectively), with concomitant formation of nonlabeled formate. The removal rate of labeled and nonlabeled formate amounted to 240 (plusmn) 15 and 400 (plusmn) 40 (mu)M h(sup-1), respectively. The formation of nonlabeled formate is explained by isotopic exchange due to the activity of formate dehydrogenase rather than by the turnover of formate.

Significance of Ether Lipids Derived from Methanogens in Sediments from Modern and Ancient Basins: ABSTRACT

High amounts of biphytanyl carbon skeletons with 0-3 cyclopentyl rings and low amounts of 2, 6, 10, 14, 18-pentamethyleicosane (PME) were detected after treatment with hydrogen iodide of fractions of several sediment extracts. These compounds are biosynthesized by archaebacteria only. Since the sediments were deposited in normal marine environments, these lipids are derived from methanogenic bacteria and more specifically Methanosarcina barkeri. In recent sediments from in and below the extensive oxygen-minimum zone in the Arabian Sea high amounts were detected in surface sediment samples where oxygen is still present. Our data suggest that these ether-bound compounds may be indicators for methane-production in the upper part of the water column and possibly an extensive oxygen-minimum zone. The concentrations varied strongly (2 orders of magnitude) in the Miocene Monterey Formation suggesting significant variations in water column methanogenic activity. The stable carbon isotopic compositions of the acyclic and cyclic biphytane carbon skeletons are several per mil heavier than those of compounds derived from photoautotrophic organisms whereas the stable carbon isotopic compositions of PME are several per mil lighter. This indicates the presence of different species of methanogens utilizing different carbon substrates.

Dissolved hydrogen concentrations in sulfate-reducing and methanogenic sediments

Dissolved hydrogen concentrations in sulfate-reducing and methanogenic sediments

Field and laboratory studies of methane oxidation in an anoxic marine sediment: Evidence for a methanogen‐sulfate reducer consortium

Field and laboratory studies of anoxic sediments from Cape Lookout Bight, North Carolina, suggest that anaerobic methane oxidation is mediated by a consortium of methanogenic and sulfate‐reducing bacteria. A seasonal survey of methane oxidation and CO2 reduction rates indicates that methane production was confined to sulfate‐depleted sediments at all times of year, while methane oxidation occurred in two modes. In the summer, methane oxidation was confined to sulfate‐depleted sediments and occurred at rates lower than those of CO2 reduction. In the winter, net methane oxidation occurred in an interval at the base of the sulfate‐containing zone. Sediment incubation experiments suggest both methanogens and sulfate reducers were responsible for the observed methane oxidation. In one incubation experiment both modes of oxidation were partially inhibited by 2‐bromoethanesulfonic acid (a specific inhibitor of methanogens). This evidence, along with the apparent confinement of methane oxidation to sulfate‐depleted sediments in the summer, indicates that methanogenic bacteria are involved in methane oxidation. In a second incubation experiment, net methane oxidation was induced by adding sulfate to homogenized methanogenic sediments, suggesting that sulfate reducers also play a role in the process. We hypothesize that methanogens oxidize methane and produce hydrogen via a reversal of CO2 reduction. The hydrogen is efficiently removed and maintained at low concentrations by sulfate reducers. Pore water H2 concentrations in the sediment incubation experiments (while net methane oxidation was occurring) were low enough that methanogenic bacteria could derive sufficient energy for growth from the oxidation of methane. The methanogen‐sulfate reducer consortium is consistent not only with the results of this study, but may also be a feasible mechanism for previously documented anaerobic methane oxidation in both freshwater and marine environments.

Toxic effects of pollutants on the mineralization of 4‐chlorophenol and benzoate in methanogenic river sediment

AbstractThe toxic effects of pollutants on the mineralization of 2 μg/L [U‐14C]‐4‐chlorophenol and benzoate were studied in microcosms with methanogenic sediment from a little harbor in the Rhine River In contrast with studies using a high sub‐strate concentration, no lag time was observed and the half‐lives for 4‐chlorophenol and benzoate were 1.6 and 0.55 h, respectively The effect of increasing additions of benzene, chloroform, 1,2‐dichloroethane, pentachlorophenol, and zinc on each mineralization reaction was measured. Toxicity data were fitted with a logistic dose‐effect curve. The IC10 is defined as the concentration of a toxicant inhibiting the mineralization rate for 10% The IC10 concentrations of benzene, chloroform, 1,2‐dichloroethane, pentachlorophenol, and zinc on the benzoate mineralization were 150, 0 04, 71, 6, and 842 mg/kg sediment dry weight, respectively. This latter value includes the background concentration of 800 mg Zn/kg sediment The mineralization of 4‐chlorophenol and benzoate showed similarities in the sensitivity to these toxicants. 4‐Chlorophenol can be degraded via benzoate, which might explain the similarities in sensitivity of both mineralization reactions. Chloroform proved to be extremely toxic to anaerobic mineralization reactions, which might be attributed to the formation of very toxic and reactive intermediates formed during the slow anaerobic degradation of the chloroform in anaerobic sediments Sediment quality criteria derived solely from standard toxicity tests using aerobic organisms may lead to complete inhibition of several important microbial processes in anaerobic sediments

Thermodynamics of methanogenic intermediary metabolism in littoral sediment of Lake Constance

In anoxic methanogenic sediments organic matter is degraded to CH4 and CO2 via intermediary metabolites. When CH4 production in slurries of littoral sediment was inhibited by chloroform, acetate accumulated with a rate (2.26 μM h−1) similar to the turnover rate (2.09 μM h−1) of [2−14C]acetate. Addition of chloroform resulted also in accumulation of propionate > 2-propanol > caproate > valerate > H2. Accumulation of H2 was small but sufficient to thermodynamically inhibit consumption of caproate and valerate by H+-reducing bacteria. Consumption stopped when the available Gibbs free energy had increased from about −16 to about −9 kJ mol−1 H2 produced. 2-Propanol increased probably mainly because of the accumulation of acetate with the available δG increasing from about −13 to −3 kJ mol−1 of 2-propanol consumed. Propionate accumulation, however, could not be explained by thermodynamic inhibition of propionate consumption since the Gibbs free energy of this reaction was generally very low (δG ≈ −3 kJ mol−1). Bacterial enrichment cultures on cellulose resulted in the production of similar metabolites as observed during the accumulation experiments. Assuming that propionat, 2-propanol, caproate and valerate were converted via acetate and H2 to CH4, their accumulation rates plus that of acetate accounted for 134% of the rate of CH4 production. Carbon flow through acetate accounted for 80–87% of the total carbon flow to CH4. This relatively high percentage may be due to the relative importance of either homoacetogenesis or of acetate-rich organic matter (e.g., chitin) in littoral sediment of Lake Constance.

Methane transport and oxidation in the unsaturated zone of aSphagnumpeatland

Efflux rates and oxidation rates of methane (CH4) were measured in a northernSphagnumbog, Thoreau's Bog in Concord, Massachusetts, by using a gradient methodology which does not change in situ conditions. A remote‐sampling technique was devised to obtain undisturbed CH4profiles as a function of depth in the peat; effective diffusion coefficients in peat were estimated both physically and by using propane as a tracer. By combining these techniques we estimated the average late summer CH44 flux from deep, anaerobic methanogenic sediments to the unsaturated zone to be 3.5 × 10−11mol cm−2s−1(± 1.0 × 10−10mol cm−2s−1), while the CH4flux from this unsaturated zone to the atmosphere was 3.7 × 10−12mol cm−2s−1(± 5.0 × 10−12mol cm−2s−1). Therefore a large fraction of the CH4flux was consumed before it reached the atmosphere. Most CH4consumption, presumably by oxidation, occurred between the water table, located 12 to 15 cm below the bog surface, and about 6 cm below the bog surface. In this region, CH4concentrations and oxidation rates were unevenly distributed, probably following patterns of upward transport of CH4by bubbles via fissures and tubes in the saturated zone. Between the surface of the bog and 6‐cm depth, CH4concentrations were more uniformly distributed, most likely because of greater horizontal mixing in this depth range. Analysis of CH4distributions in unsaturated peat is a straightforward and practical technique to measure both net CH4efflux and CH4oxidation with minimal disturbance to the peat structure and gas exchange conditions.

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

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

Isolation and partial characterization of aClostridium species transforming para-hydroxybenzoate and 3,4-dihydroxybenzoate and producing phenols as the final transformation products

Organisms present in methanogenic freshwater lake sediments from the vicinity of Athens, Georgia, were adapted to mineralize 2,4-dichlorophenol. Repeated addition of 0.5 to 2.7 mmol/liter of phenol, and later of 0.5-6.2 mmol/liter p-hydroxybenzoate (p-OHB), to such enrichments led to the conversion of p-OHB to phenol at a rate of up to 100 mmol p-OHB per liter per day. Subsequently, a spore-forming, obligately anaerobic bacterium, strain JW/Z-1, was isolated which transformed p-OHB to phenol and 3,4-dihydroxybenzoate (3,4-OHB) to catechol (1,2-dihydroxybenzene) stoichiometrically without further metabolism of the phenols. The strain did not transform benzoate, 4-chlorophenol, 2,4-dichlorophenol, 4-chlorobenzoate, o- and m-hydroxybenzoate, 2,4- and 3,5-dihydroxybenzoate, 2,3,4- and 3,4,5-trihydroxybenzoate, or 4-aminobenzoate. Yeast extract was required for growth of strain JW/Z-1 and only high concentrations of casein hydrolysate or tryptone could substitute it, to some extent. Except for sodium acetate, and some amino acids together with a 20-fold increased concentration of vitamins, no single carbohydrate or defined organic compound has been found to support growth of this strain in the presence (or in the absence) of 0.2 to 0.5% (w/v) yeast extract. The fermentation products during growth on yeast extract indicated that the metabolism of amino acid degradation was the major source for growth. The decarboxylating activity was inducible by p-OHB for the decarboxylation of p-OHB, and at a lower rate for 3,4-OHB, and by 3,4-OHB only for 3,4-OHB, suggesting that two different enzyme systems exist. The addition of the aromatic amino acids phenol or benzoate did not induce the decarboxylation activity in cultures growing with yeast extract. Growth was observed at temperatures ranging from 12-41°C (Topt, 33-34°C) and at pH-values ranging from 6.0-10.0 (pHopt, 7.2-8.2). The shortest doubling time observed for strain JW/Z-1 was 3.2 hours.

In situ stimulation of bacterial sulfate reduction in sulfate-limited freshwater lake sediments

By adding sulfate in the form of solid gypsum, it was possible to transform in situ a predominantly methanogenic sediment ecosystem into a sulfate-reducing one. The concentrations of sulfate, sulfide, methane, acetate, propionate, soluble iron, and manganese were determined in the porewater before and after the transition. Although sulfate was no longer limiting, acetate and propionate continued to accumulate and reached much higher concentrations than under sulfate-limited conditions. Metabolic activities of fermenting bacteria and of sulfate reducers, which belong to the group that incompletely oxidizes organic material, might be responsible for the increased production of volatile fatty acids. The elevated concentrations of soluble Fe(II)2+ and Mn(II)2+ observed in the porewater stem from iron and manganese compounds which may be reduced chemically by hydrogen sulfide and other microbially produced reducing agents or directly through increased activities of the iron and manganese reducing bacteria. In the horizon with high sulfate-reducing activities the methane concentrations in the porewater were lower than in non-stimulated sediment regions. The shape of the concentration depth profile indicates methane consumption through sulfate reducing processes. The in situ experiment demonstrates the response of a natural microbial ecosystem to fluctuations in the environmental conditions.

In situ stimulation of bacterial sulfate reduction in sulfate-limited freshwater lake sediments

By adding sulfate in the form of solid gypsum, it was possible to transform in situ a predominantly methanogenic sediment ecosystem into a sulfate-reducing one. The concentrations of sulfate, sulfide, methane, acetate, propionate, soluble iron, and manganese were determined in the porewater before and after the transition. Although sulfate was no longer limiting, acetate and propionate continued to accumulate and reached much higher concentrations than under sulfate-limited conditions. Metabolic activities of fermenting bacteria and of sulfate reducers, which belong to the group that incompletely oxidizes organic material, might be responsible for the increased production of volatile fatty acids. The elevated concentrations of soluble Fe(II) 2+ and Mn(II) 2+ observed in the porewater stem from iron and manganese compounds which may be reduced chemically by hydrogen sulfide and other microbially produced reducing agents or directly through increased activities of the iron and manganese reducing bacteria. In the horizon with high sulfate-reducing activities the methane concentrations in the porewater were lower than in non-stimulated sediment regions. The shape of the concentration depth profile indicates methane consumption through sulfate reducing processes. The in situ experiment demonstrates the response of a natural microbial ecosystem to fluctuations in the environmental conditions.

Start‐up of low temperature anaerobic reactors using freshwater methanogenic sediments

Summary The feasibility of the utilisation of freshwater methanogenic sediments as inocula for anaerobic wastewater treatment processes at low temperature (20°C) was studied. This process appeared to be stable for the three substrates studied: pig manure, distillery waste and sewage sludge, with a total solids concentration of 40, 25 and 32 kg.m−3, respectively. The range of organic load applied was 0.1 ‐1.3 kg COD.m−3d−1; biogas production ranged from 0.195 to 0.257 mB 3.mD −3d−1, and the percentage of methane was of 66 ‐ 76%. The results reported permit to conclude that freshwater sediments are a promising alternative to conventional anaerobic sludge for the inoculation of lowtemperature anaerobic digesters.

The kinetics of the degradation of chloroform and benzene in anaerobic sediment from the River Rhine

In anaerobic methanogenic sediment microcosms14C labelled chloroform was degraded mainly to carbon dioxide. At a concentration of 4 μg.l−1 the mineralization followed first order kinetics with a half life of 12 days at 10°C and 2.6 days at 20°C. At a concentration of 400 μg.l−1 the mineralization rate increased with time and followed logarithmic kinetics with a μmax of 0.02.d−1 at 10°C. The logarithmic kinetics can be explained by growth of the bacteria on the higher concentration of chloroform with a generation time of 35 days. Shaking and oxygenation did not inhibit the mineralization of chloroform, probably because of bacterial consumption of the dissolved oxygen. 14C labelled benzene was mineralized only for a small percentage to14C labelled carbon dioxide while other, not acid extractable, degradation products were formed. Under anaerobic conditions after one day when 5% of the benzene was degraded to carbon dioxide, the mineralization ceased, while the disappearance of benzene proceeded. With air in the headspace of the incubation bottles 25% of the benzene was mineralized to carbon dioxide. The anaerobic degradation of benzene at a concentration of 100 μ.l−1 showed similar kinetics as the degradation at 1 μg.l−1. Hence no adaptation of the microflora in the sediment occurred during the 63 days of the experiment at 100 μg.l−1.

Requirement for a Microbial Consortium To Completely Oxidize Glucose in Fe(III)-Reducing Sediments

In various sediments in which Fe(III) reduction was the terminal electron-accepting process, [C]glucose was fermented to C-fatty acids in a manner similar to that observed in methanogenic sediments. These results are consistent with the hypothesis that, in Fe(III)-reducing sediments, fermentable substrates are oxidized to carbon dioxide by the combined activity of fermentative bacteria and fatty acid-oxidizing, Fe(III)-reducing bacteria.

Populations of Methanogenic Bacteria in a Georgia Salt Marsh

Methanogens represented about 0.5% of the total bacteria in sediments from a Georgia salt marsh in which Spartina alterniflora is the predominant vegetation. The population of methanogens was composed of at least two groups of nearly equal size. One group was represented by cocci which were able to utilize trimethylamine and were unable to use H(2) or acetate. The second group was composed of two subgroups which were able to utilize H(2) but were unable to use trimethylamine or acetate. The more common subgroup included rod- or plate-shaped methanogens which could utilize isopropanol in addition to H(2) and formate. The second subgroup included Methanococcus maripaludis, which utilized only H(2) and formate. Other groups of methanogens were also present, including Methanosarcina sp. which utilized acetate, H(2), and methylamines. In addition to the overall variability in the types of methanogens, the numbers of methanogens in sediments also exhibited significant spatial variability both within and between tall- and short-Spartina zones.