Microbial Formation of Dimethyl Sulfide in Anoxic Sphagnum Peat

Peat bogs dominated by Sphagnum spp. have relatively high areal rates of dimethyl sulfide (DMS) emission to the atmosphere. DMS was produced in anoxic slurries of Sphagnum peat with a linear time course and with an average rate of 40.4 (range, 22.0 to 68.6) nmol per liter of slurry (middot) day(sup-1) observed in nine batches of slurry. Methanethiol (MeSH) was produced at roughly similar rates over the typical 4- to 8-day incubations. DMS and MeSH production in these acidic (pH 4.2 to 4.6) peats were biological, as they were stopped completely by autoclaving and inhibited strongly by addition of antibiotics and 500 (mu)M chloroform. Endogenous DMS production may be due to the degradation of S-methyl-methionine, dimethyl sulfoxide, or methoxyaromatic compounds (e.g., syringic acid), each of which stimulated DMS formation when added at 5 to 10 (mu)M concentrations. However, on the basis of the high rates of thiol (MeSH and ethanethiol) methylation activity that we observed and the availability of endogenous MeSH, we suggest that methylation of MeSH is the major pathway leading to DMS formation in anaerobic peat. Solid-phase adsorption of MeSH plays a key role in its availability for biomethylation reactions. Additions of acetate (1.5 mM) or compounds which could cause acetate to accumulate (e.g., glucose, alanine, and 2-bromoethanesulfonate) suppressed DMS formation. It is likely that acetogenic bacteria are involved in DMS formation, but our data are insufficient to allow firm conclusions about the metabolic pathways or organisms involved. Our observations are the first which point to the methylation of MeSH as the major mechanism for endogenous DMS production in any environment. The rates of net DMS production observed are sufficient to explain the relatively high fluxes of DMS emitted to the atmosphere from Sphagnum sp.-dominated wetlands.     

Dynamics of Dimethyl Sulfide in a Marine Microbial Mat

Abstract The microbial mat was chosen as a model ecosystem to study dynamics of dimethyl sulfide (DMS) in marine sediments in order to gain insight into key processes and factors which determine emission rates. A practical advantage, compared to open ocean ecosystems, is that microbial mats contain high biomasses of different functional groups of bacteria involved in DMS dynamics, and that DMS concentrations are generally high enough to allow direct measurement of emission rates. Field data showed that, during the seasonal development of microbial mats, concentrations of chlorophyll a corresponded to dimethylsulfoniopropionate (DMSP). DMSP is an important precursor of DMS. It was demonstrated, with laboratory cultures, that various species of benthic diatoms produce substantial amounts of DMSP. The abundances of aerobic and anaerobic DMS- or DMSO-utilizing bacteria were estimated using the most-probable-number technique. Laboratory experiments with relatively undisturbed sediment cores showed that microbial mats act as a sink for DMS under oxic/light (day) conditions, and as a source of DMS under anoxic/dark (night) conditions. Axenic culture studies with Chromatium vinosum M2 and Thiocapsa pfennigii M8 (isolated from a microbial mat) showed that, under anoxic/light conditions, DMS was quantitatively converted to dimethylsulfoxide (DMSO). T. roseopersicina M11 converted DMSP to DMS and acrylate, apparently without use of either substrate. Received: 5 May 1997; Accepted: 21 August 1997     

Microbial Populations Involved in Cycling of Dimethyl Sulfide and Methanethiol in Freshwater Sediments

Although several microorganisms that produce and degrade methanethiol (MT) and dimethyl sulfide (DMS) have been isolated from various habitats, little is known about the numbers of these microorganisms in situ. This study reports on the identification and quantification of microorganisms involved in the cycling of MT and DMS in freshwater sediments. Sediment incubation studies revealed that the formation of MT and DMS is well balanced with their degradation. MT formation depends on the concentrations of both sulfide and methyl group-donating compounds. A most-probable number (MPN) dilution series with syringate as the growth substrate showed that methylation of sulfide with methyl groups derived from syringate is a commonly occurring process in situ. MT appeared to be primarily degraded by obligately methylotrophic methanogens, which were found in the highest positive dilutions on DMS and mixed substrates (methanol, trimethylamine [TMA], and DMS). Amplified ribosomal DNA restriction analysis (ARDRA) and 16S rRNA gene sequence analysis of the total DNA isolated from the sediments and of the DNA isolated from the highest positive dilutions of the MPN series (mixed substrates) revealed that the methanogens that are responsible for the degradation of MT, DMS, methanol, and TMA in situ are all phylogenetically closely related to Methanomethylovorans hollandica. This was confirmed by sequence analysis of the product obtained from a nested PCR developed for the selective amplification of the 16S rRNA gene from M. hollandica. The data from sediment incubation experiments, MPN series, and molecular-genetics detection correlated well and provide convincing evidence for the suggested mechanisms for MT and DMS cycling and the common presence of the DMS-degrading methanogen M. hollandica in freshwater sediments.     

施用微生物制剂对育蚌水体微生物区系的影响

研究了育蚌水体分别施用微生物制剂利水素与鱼虾菌乐后水样中微生物区系的变化。试验结果表明:施用利水素的水样革兰氏阴性杆菌和嗜水气单胞菌含量分别下降26.67%、20%,芽胞杆菌和表皮葡萄球菌含量分别上升13.33%、26.67%;施用鱼虾菌乐的水样革兰阴性杆菌和嗜水气单胞菌含量分别下降20%、13.33%,芽胞杆菌和表皮葡萄球菌含量分别上升6.67%、13.33%;未施用微生物制剂的水样革兰阴性杆菌和嗜水气单胞菌含量均上升13.33%,芽胞杆菌消失,新增肠杆菌和链球菌。施用微生物制剂利水素和鱼虾菌乐,可有效优化养殖水体微生物种群结构,改良养殖水质环境。     

Second-Order Model to Predict Microbial Degradation of Organic Compounds in Natural Waters

The reliability of second-order rate constants for assessing microbial degradation kinetics in natural waters was examined by using three compounds that undergo hydrolytic degradation. The butoxyethyl ester of 2,4-dichlorophenoxyacetic acid was studied in water samples from 31 sites, malathion was examined in water from 14 sites, and chlorpropham was studied in samples from 11 sites. The coefficient of variation for rate constants for each compound was less than 65% over all sites. Additional studies indicated that the rate conformed to second-order kinetics; that is, the rate was proportional to both bacterial and xenobiotic concentrations.     

Kinetics of Mixed Microbial Assemblages Enhance Removal of Highly Dilute Organic Substrates

Our experiments with selected organic substrates reveal that the rate-limiting process governing microbial degradation rates changes with substrate concentration, S, in such a manner that substrate removal is enhanced at lower values of S. This enhancement is the result of the dominance of very efficient systems for substrate removal at low substrate concentrations. The variability of dominant kinetic parameters over a range of S causes the kinetics of complex assemblages to be profoundly dissimilar to those of systems possessing a single set of kinetic parameters; these findings necessitate taking a new approach to predicting substrate removal rates over wide ranges of S.     

Effects of Organic Substrates on Dechlorination of Aroclor 1242 in Anaerobic Sediments

The effects of different organic substrates on the abilities of anaerobic sediment enrichments to reductively dechlorinate polychlorinated biphenyls (PCBs) were studied. Sediments collected from a site previously contaminated with PCBs were dosed with additional PCBs (Aroclor 1242; approximately 300 ppm [300 μg/g], sediment dry weight) and incubated anaerobically with acetate, acetone, methanol, or glucose. The pattern of dechlorination was similar for each substrate-fed batch; however, the extents and rates of dechlorination were different. Significant dechlorination over time was observed, with the relative rates and extents of dechlorination being greatest for methanol-, glucose-, and acetone-fed batches and least for acetate-fed batches. Dechlorination occurred primarily on the meta- and para- positions of the highly chlorinated congeners, resulting in the accumulation of less-chlorinated, primarily ortho-substituted products. No significant dechlorination was observed in incubation batches receiving no additional organic substrate, even though identical inorganic nutrients were added to all incubation batches. In addition, dechlorination was not observed in autoclaved controls that received substrate and nutrients.     

Denitrification, Nitrate Reduction, and Oxygen Consumption in Coastal and Estuarine Sediments

Denitrification and consumption of oxygen and nitrate in sediments from Tama Estuary, Odawa Bay, and Tokyo Bay were measured in an experimental sediment-water system. Filtered seawater containing [¹⁵N]nitrate flowed continuously over undisturbed sediments, and the concentrations of O₂, nitrate, and nitrite in the influent and effluent and of ¹⁵N₂ in the effluent were monitored. Under steady-state conditions, the rate of nitrate consumption was the same order of magnitude as the rate of oxygen consumption in Tama Estuary sediments, whereas the former rate was one order of magnitude lower than the latter rate in Odawa Bay and Tokyo Bay sediments. Denitrification accounted for 27 to 57% of the nitrate consumption.     

Water Organic Pollution and Eutrophication Influence Soil Microbial Processes,Increasing Soil Respiration of Estuarine Wetlands: Site Study in Jiuduansha Wetland

Undisturbed natural wetlands are important carbon sinks due to their low soil respiration. When compared with inland alpine wetlands, estuarine wetlands in densely populated areas are subjected to great pressure associated with environmental pollution. However, the effects of water pollution and eutrophication on soil respiration of estuarine and their mechanism have still not been thoroughly investigated. In this study, two representative zones of a tidal wetland located in the upstream and downstream were investigated to determine the effects of water organic pollution and eutrophication on soil respiration of estuarine wetlands and its mechanism. The results showed that eutrophication, which is a result of there being an excess of nutrients including nitrogen and phosphorus, and organic pollutants in the water near Shang shoal located upstream were higher than in downstream Xia shoal. Due to the absorption and interception function of shoals, there to be more nitrogen, phosphorus and organic matter in Shang shoal soil than in Xia shoal. Abundant nitrogen, phosphorus and organic carbon input to soil of Shang shoal promoted reproduction and growth of some highly heterotrophic metabolic microorganisms such as β-Proteobacteria, γ-Proteobacteria and Acidobacteria which is not conducive to carbon sequestration. These results imply that the performance of pollutant interception and purification function of estuarine wetlands may weaken their carbon sequestration function to some extent.     

Formation of Dimethyl Sulfide and Methanethiol in Anoxic Freshwater Sediments

Concentrations of volatile organic sulfur compounds (VOSC) were measured in water and sediment columns of ditches in a minerotrophic peatland in The Netherlands. VOSC, with methanethiol (4 to 40 nM) as the major compound, appeared to be mainly of sediment origin. Both VOSC and hydrogen sulfide concentrations decreased dramatically towards the water surface. High methanethiol and high dimethyl sulfide concentrations in the sediment and just above the sediment surface coincided with high concentrations of hydrogen sulfide (correlation factors, r = 0.91 and r = 0.81, respectively). Production and degradation of VOSC were studied in 32 sediment slurries collected from various freshwater systems in The Netherlands. Maximal endogenous methanethiol production rates of the sediments tested (up to 1.44 (mu)mol per liter of sediment slurry (middot) day(sup-1)) were determined after inhibition of methanogenic and sulfate-reducing populations in order to stop VOSC degradation. These experiments showed that the production and degradation of VOSC in sediments are well balanced. Statistical analysis revealed multiple relationships of methanethiol production rates with the combination of methane production rates (indicative of total anaerobic mineralization) and hydrogen sulfide concentrations (r = 0.90) or with the combination of methane production rates and the sulfate/iron ratios in the sediment (r = 0.82). These findings and the observed stimulation of methanethiol formation in sediment slurry incubations in which the hydrogen sulfide concentrations were artificially increased provided strong evidence that the anaerobic methylation of hydrogen sulfide is the main mechanism for VOSC formation in most freshwater systems. Methoxylated aromatic compounds are likely a major source of methyl groups for this methylation of hydrogen sulfide, since they are important degradation products of the abundant biopolymer lignin. Increased sulfate concentrations in several freshwater ecosystems caused by the inflow of water from the river Rhine into these systems result in higher hydrogen sulfide concentrations. As a consequence, higher fluxes of VOSC towards the atmosphere are conceivable.     

Occurrence of Vibrio parahaemolyticus in Estuarine Waters and Oysters of New Hampshire

Vibrio parahaemolyticus was isolated from water and oysters collected from seven different sampling stations in the Great Bay and Little Bay estuarine areas of New Hampshire. The morphological and biochemical characteristics of 50 isolates conformed in general to those described for this organism in the literature. All isolates produced hemolysis on blood-agar. To date, there have been no reports of V. parahaemolyticus food poisoning outbreaks due to the consumption of fish or shellfish harvested from this estuarine region.     

Microbial Formation of Ethane in Anoxic Estuarine Sediments

Estuarine sediment slurries produced methane and traces of ethane when incubated under hydrogen. Formation of methane occurred over a broad temperature range with an optimum above 65°C. Ethane formation had a temperature optimum at 40°C. Formation of these two gases was inhibited by air, autoclaving, incubation at 4 and 80°C, and by the methanogenic inhibitor, 2-bromoethanesulfonic acid. Ethane production was stimulated by addition of ethylthioethanesulfonic acid, and production from ethylthioethanesulfonic acid was blocked by 2-bromoethanesulfonic acid. A highly purified enrichment culture of a methanogenic bacterium obtained from sediments produced traces of ethane from ethylthioethanesulfonic acid. These results indicate that the small quantities of ethane found in anaerobic sediments can be formed by certain methanogenic bacteria.     

Preliminary Evidence for the Amplification of Global Warming in Shallow,Intertidal Estuarine Waters

Over the past 50 years, mean annual water temperature in northeastern U.S. estuaries has increased by approximately 1.2°C, with most of the warming recorded in the winter and early spring. A recent survey and synthesis of data from four locations in Southern Rhode Island has led us to hypothesize that this warming may be amplified in the shallow (<1 m), nearshore portions of these estuaries. While intertidal areas are not typically selected as locations for long-term monitoring, we compiled data from published literature, theses, and reports that suggest that enhanced warming may be occurring, perhaps at rates three times higher than deeper estuarine waters. Warmer spring waters may be one of the factors influencing biota residing in intertidal regions both in general as well as at our specific sites. We observed greater abundance of fish, and size of Menidia sp., in recent (2010–2012) seine surveys compared to similar collections in 1962. While any linkages are speculative and data are preliminary, taken together they suggest that shallow intertidal portions of estuaries may be important places to look for the effects of climate change.     

Respiratory Succession and Community Succession of Bacterioplankton in Seasonally Anoxic Estuarine Waters

Anoxia occurs in bottom waters of stratified estuaries when respiratory consumption of oxygen, primarily by bacteria, outpaces atmospheric and photosynthetic reoxygenation. Once water becomes anoxic, bacterioplankton must change their metabolism to some form of anaerobic respiration. Analysis of redox chemistry in water samples spanning the oxycline of Chesapeake Bay during the summer of 2004 suggested that there was a succession of respiratory metabolism following the loss of oxygen. Bacterial community doubling time, calculated from bacterial abundance (direct counts) and production (anaerobic leucine incorporation), ranged from 0.36 to 0.75 day and was always much shorter than estimates of the time that the bottom water was anoxic (18 to 44 days), indicating that there was adequate time for bacterial community composition to shift in response to changing redox conditions. However, community composition (as determined by PCR-denaturing gradient gel electrophoresis analysis of 16S rRNA genes) in anoxic waters was very similar to that in surface waters in June when nitrate respiration was apparent in the water column and only partially shifted away from the composition of the surface community after nitrate was depleted. Anoxic water communities did not change dramatically until August, when sulfate respiration appeared to dominate. Surface water populations that remained dominant in anoxic waters were Synechococcus sp., Gammaproteobacteria in the SAR86 clade, and Alphaproteobacteria relatives of Pelagibacter ubique, including a putative estuarine-specific Pelagibacter cluster. Populations that developed in anoxic water were most similar (<92% similarity) to uncultivated Firmicutes, uncultivated Bacteroidetes, Gammaproteobacteria in the genus Thioalcalovibrio, and the uncultivated SAR406 cluster. These results indicate that typical estuarine bacterioplankton switch to anaerobic metabolism under anoxic conditions but are ultimately replaced by different organisms under sulfidic conditions.     

Comparison of the Seasonal Variations of Synechococcus Assemblage Structures in Estuarine Waters and Coastal Waters of Hong Kong

Seasonal variation in the phylogenetic composition of Synechococcus assemblages in estuarine and coastal waters of Hong Kong was examined through pyrosequencing of the rpoC1 gene. Sixteen samples were collected in 2009 from two stations representing estuarine and ocean-influenced coastal waters, respectively. Synechococcus abundance in coastal waters gradually increased from 3.6 × 10³ cells ml⁻¹ in March, reaching a peak value of 5.7 × 10⁵ cells ml⁻¹ in July, and then gradually decreased to 9.3 × 10³ cells ml⁻¹ in December. The changes in Synechococcus abundance in estuarine waters followed a pattern similar to that in coastal waters, whereas its composition shifted from being dominated by phycoerythrin-rich (PE-type) strains in winter to phycocyanin-only (PC-type) strains in summer owing to the increase in freshwater discharge from the Pearl River and higher water temperature. The high abundance of PC-type Synechococcus was composed of subcluster 5.2 marine Synechococcus, freshwater Synechococcus (F-PC), and Cyanobium. The Synechococcus assemblage in the coastal waters, on the other hand, was dominated by marine PE-type Synechococcus, with subcluster 5.1 clades II and VI as the major lineages from April to September, when the summer monsoon prevailed. Besides these two clades, clade III cooccurred with clade V at relatively high abundance in summer. During winter, the Synechococcus assemblage compositions at the two sites were similar and were dominated by subcluster 5.1 clades II and IX and an undescribed clade (represented by Synechococcus sp. strain miyav). Clade IX Synechococcus was a relatively ubiquitous PE-type Synechococcus found at both sites, and our study demonstrates that some strains of the clade have the ability to deal with large variation of salinity in subtropical estuarine environments. Our study suggests that changes in seawater temperature and salinity caused by the seasonal variation of monsoonal forcing are two major determinants of the community composition and abundance of Synechococcus assemblages in Hong Kong waters.     

Effects of Endogenous Substrates on Adaptation of Anaerobic Microbial Communities to 3-Chlorobenzoate

Lengthy adaptation periods in laboratory studies evaluating the potential for contaminant biodegradation in natural or engineered environments may indicate that the native microbial communities are not metabolizing the contaminants in situ. In this study, we characterized the adaptation period preceding the biodegradation of 3-chlorobenzoate in anaerobic communities derived from lake sediment and wastewater sludge digesters. The importance of alternative mechanisms of adaptation of the anaerobic communities to 3-chlorobenzoate was evaluated by monitoring the concentrations of metabolic substrates and products as well as the levels of total small subunit (SSU) rRNA and SSU rRNA from populations thought to be important in 3-chlorobenzoate mineralization. The anaerobic environments from which the 3-chlorobenzoate-degrading communities were derived contained different levels of endogenous substrates. Increasing methane levels in the digester and sediment communities and decreasing chemical oxygen demand concentrations in the sediment community during the adaptation periods revealed that endogenous substrates were preferentially utilized relative to 3-chlorobenzoate. Methane and chemical oxygen demand concentrations leveled off concomitantly with the onset of 3-chlorobenzoate biodegradation, suggesting that depletion of the preferentially degraded endogenous substrates stimulated 3-chlorobenzoate metabolism. Consistent with these observations, adaptation to 3-chlorobenzoate occurred more rapidly in digester samples that were depleted of endogenous substrates compared to samples that contained high levels of these biodegradable compounds. Other potential adaptation mechanisms, e.g., genetic change or selective population enrichment, appeared to be less important based on the reproducibility and relative lengths of the adaptation events, trends in the SSU rRNA levels, and/or amplification of SSU rRNA genes from key populations.     

Methanogenesis and Sulfate Reduction: Competitive and Noncompetitive Substrates in Estuarine Sediments

Sulfate ions did not inhibit methanogenesis in estuarine sediments supplemented with methanol, trimethylamine, or methionine. However, sulfate greatly retarded methanogenesis when hydrogen or acetate was the substrate. Sulfate reduction was stimulated by acetate, hydrogen, and acetate plus hydrogen, but not by methanol or trimethylamine. These results indicate that sulfate-reducing bacteria will outcompete methanogens for hydrogen, acetate, or both, but will not compete with methanogens for compounds like methanol, trimethylamine, or methionine, thereby allowing methanogenesis and sulfate reduction to operate simultaneously within anoxic, sulfate-containing sediments.     

Ecology of Vibrio vulnificus in Estuarine Waters of Eastern North Carolina

While several studies on the ecology of Vibrio vulnificus in Gulf Coast environments have been reported, there is little information on the distribution of this pathogen in East Coast waters. Thus, we conducted a multiyear study on the ecology of V. vulnificus in estuarine waters of the eastern United States, employing extensive multiple regression analyses to reveal the major environmental factors controlling the presence of this pathogen, and of Vibrio spp., in these environments. Monthly field samplings were conducted between July 2000 and April 2002 at six different estuarine sites along the eastern coast of North Carolina. At each site, water samples were taken and nine physicochemical parameters were measured. V. vulnificus isolates, along with estuarine bacteria, Vibrio spp., Escherichia coli organisms, and total coliforms, were enumerated in samples from each site by using selective media. During the last 6 months of the study, sediment samples were also analyzed for the presence of vibrios, including V. vulnificus. Isolates were confirmed as V. vulnificus by using hemolysin gene PCR or colony hybridization. V. vulnificus was isolated only when water temperatures were between 15 and 27°C, and its presence correlated with water temperature and dissolved oxygen and vibrio levels. Levels of V. vulnificus in sediments were low, and no evidence for an overwintering in this environment was found. Multiple regression analysis indicated that vibrio levels were controlled primarily by temperature, turbidity, and levels of dissolved oxygen, estuarine bacteria, and coliforms. Water temperature accounted for most of the variability in the concentrations of both V. vulnificus (47%) and Vibrio spp. (48%).     

不同底物对海洋混合菌群产氢的影响

将采自天津潮间带的污泥进行热休克处理,厌氧条件下富集混合菌群进行产氢试验。混合菌群分别接种于含葡萄糖、蔗糖、乳糖、淀粉和蛋白胨的培养液中,测定不同底物培养条件下混合菌群产氢量及菌群组成。结果表明,以蔗糖为底物时,混合菌群产氢量最高,为（787±24）mL／L;混合菌群以葡糖糖、乳糖和淀粉为底物时,产氢量依次为（530±20）、（46±5）和（455±35）mL／L;混合菌群不能利用蛋白胨为底物产氢。变性梯度凝胶电泳（DGGE）分析不同底物培养条件下的产氢混合菌群组成。混合菌群16S rRNA基因的DGGE分离结果表明,在蛋白胨培养条件下,混合菌群没有能够形成优势菌,其他底物培养时,混合菌群的优势菌是Clostridium sp．。