Physiological and immunological evidence for two distinct C1-utilizing pathways in Bathymodiolus puteoserpentis (Bivalvia: Mytilidae), a dual endosymbiotic mussel from the Mid-Atlantic Ridge

The existence of endosymbiotic sulfur-oxidizing chemoautotrophic and methanotrophic bacteria associating with marine mytilid mussels has previously been inferred by 16S rDNA analysis in Bathymodiolus puteoserpentis Von Cosel et al. 1994, a hydrothermal vent mussel from a site on the Mid-Atlantic Ridge. In mussels collected in June 1993, we found evidence of enzymes diagnostic of two distinct C₁ assimilation pathways in this symbiosis. Assays for the utilization of radiolabelled methane and for immunodetection of methanol dehydrogenase were positive, indicating that oxidation and incorporation of this substrate are occurring in this symbiosis. Sulfide or thiosulfate had no detectable stimulatory effect on CO₂ incorporation, and assays for the enzyme ribulose-1,5-bisphosphate carboxylase/oxygenase (RubisCO), an enzyme diagnostic for the Calvin–Benson cycle, were negative. RubisCO was detected in all samples examined by immunoblot analysis, indicating this enzyme is expressed in the B. puteoserpentis symbiosis. Stable isotope data showed that carbon isotope values were in agreement with previously reported values, and nitrogen isotope values were among the most depleted ever reported for bivalve symbioses. The carbon isotope values do not preclude the utilization of vent-derived methane. These data could be explained by the presence of two metabolically distinct bacterial symbionts or a Type X methanotrophic symbiont. Received: 3 October 1997 / Accepted: 23 July 1998     

S–O–C isotopic picture of sulphate–methane–carbonate system in freshwater lakes from Poland. A review

Microbial oxidation of organic compounds (including methane), in freshwater sediments, may result in precipitation of carbonates, which may become an important geochemical archive of paleoenvironmental variations. Most probably low δ¹³C value in calcite in eutrophic systems results from an advanced oxidation of organic compounds in turbulent or/and sulphate-rich conditions. Likewise, high δ¹³C value in calcite from organic-rich sediments may evidence low redox potential of the freshwater system. Oxidation of methane and organic matter results in significant isotope effects in sulphates dissolved in water. Therefore, to better understand the origin of carbon isotope signal in carbonates, concentration and stable isotope measurements in dissolved sulphate (water column), bubble methane and calcite (freshwater sediments) have been carried out in 24 lakes, 2 ponds and 4 rivers in Poland. The highest concentration of sulphate has been detected in rivers (85.47 SO₄ ²⁻ mg/l) and an artificial lake (70.30 SO₄ ²⁻ mg/l) located in the extremely SO₄ ²⁻-polluted region called the “Black Triangle”. The lowest concentration of sulphate is found in dystrophic and mountain lakes (from 0.5 SO₄ ²⁻ to about 3 mg/l). The lowest δ³⁴S(SO₄ ²⁻) and δ¹⁸O(SO₄ ²⁻) values occur in unpolluted lakes in eastern Poland (−0.94 and 1.38‰, respectively). The highest S and O isotopic ratios are found in a polluted lake in western Poland (δ¹⁴S(SO₄ ²)=12.95‰) and in a dystrophic lake in eastern Poland (δ¹⁸O(SO₄ ²) = 16.15‰) respectively. It is proposed that δ³⁴SO₄ ²⁻ and (¹⁸O(SO₄ ²⁻) values in lakes represent a good tool to assess and quantify anthropogenic impact by acid precipitation and to monitor variations in the trophic state and redox processes controlled by biodegradation of organic compounds in sediments and water column. In general, increasing depth (up to 12 m) of the water column is associated with decreasing trend the δ¹³C(CH₄) value from about –35 to about –78‰. However, δ¹³C value in sedimentary calcite (δ¹³C vary from –10 to 0.5‰) shows opposite trends as compared to the corresponding methane. This is probably due to redox processes and distribution of heavy isotopes between methane and calcite. Likewise, turbulent water (river) show high δ¹³C value in methane and low δ¹³C value in calcite—this is probably due to an enhanced oxidation of methane producing ¹³C-depleted CO₂. In contrast to clean lakes, it is observed that an increase of the δ¹³C(CH₄) value occurs with increasing depth of the water column in a strongly SO₄ ²⁻-contaminated lake. This is probably due to a loss of biological buffering potential of the lake accompanied by an active oxidation of methane precursors.     

Interrelationships between sulfate reducing and methane producing bacteria in coastal sediments with intense sulfide production

Coastal sediments receiving different amounts of organic carbon through sedimentation were investigated with respect to sulfate reduction and methanogenic activity. Sampling was carried out at sediment temperatures of 7° and 15°C. Sulfate-reducing and methanogenic bacteria were found at all depths. Sulfate reduction decreased with depth and the highest sulfide concentrations were found a few centimeters below the sediment surface, up to 15 mM at 15°C and pH 7.1. In the same segments a maximum in the methane concentration was also found, 0.91 mM. The high sulfide concentration inhibited the methane formation from acetate but not from carbon dioxide. In the organic rich sediment sulfate reduction was limited by the diffusion of SO ₄ ⁺ into the sediment and methane production from acetate by sulfide diffusion out of the sediment. When electron acceptor concentration limits sulfate reduction, thermodynamic calculations show that the utilization of electron donors more reduced than acetate is favored. In the sediment with the high carbon-input, acetate predominated at 15°C whereas in the low carbon-input sediment hardly any short chain organic acids were detected. The possibility of a shift in sulfate reduction from acetate oxidation to acetate production is discussed.     

Stable C and N isotope analysis elucidated the importance of zooplankton in a tropical seagrass bed of Santiago Island,Northwestern Philippines

The role of zooplankton in a tropical seagrass ecosystem was investigated in milkfish farms pollution-impacted and -unimpacted seagrass beds in Santiago Island coral reefs, Northwestern Philippines. The aim was to compare between the two sites: (1) abiotic factors and zooplankton community parameters, and (2) the trophic structure using C and N stable isotopes. Low water (98–119?mV) and sediment (–121 to ?138?mV) Oxidation Reduction Potential values indicated a reducing environment in the impacted site. Zooplankton in the impacted site showed the typical community response to eutrophication (low diversity, but high total abundance due to the dominance of the cyclopoid copepod Oithona oculata), generally few elevated δ¹⁵N values, but a significant shift towards depleted ¹³C due to the organic enrichment of fish-farm feeds. Apart from suggesting a highly complex food web with POM and zooplankton as main food sources in the unimpacted site, the Bayesian mixing model simulation generated reduced complexity in feeding interactions between basal sources, zooplankton, and fish including adults of a key fish species, Siganus fuscescens, in the impacted sites. In this study, C and N stable isotope analysis has clarified the importance of zooplankton as fish prey in a seagrass bed food web.     

A study on cometabolic bioventing for the in situ remediation of trichloroethylene

Cometabolic bioventing for removal of TCE in the unsaturated zone was studied in a soil column study using methane as growth substrate. A numerical model was developed for simulating the behavior of TCE during cometabolic bioventing. The model parameters were estimated independently through laboratory batch experiments or from the literature. Simulations were found to provide reasonable agreement with the experimental data. The experimental data show that a total TCE remediation efficiency of over 95% was obtained. The volatilization-to-biodegradation ratio of TCE was about 7:1 and T _c values ranging from 0.0078 to 0.07 were obtained in this methane-driven system. Due to the toxicity of the high TCE concentrations to the microbial biomass in the initial stages of the experiment, cometabolic biodegradation was enhanced and was more efficient in the later stages of cometabolic bioventing.     

甲醇对土壤甲烷氧化的影响及其微生物学机理

对设施栽培土壤甲烷氧化及其微生物学机理进行了研究，结果表明，不同土壤对甲烷的氧化能力各异，这可能与土壤的理化性质有关。土壤微生物是甲烷氧化的主要生物类群，不同碳源对甲烷氧化的影响各异，纤维素对甲烷氧化的抑制作用最小，高浓度的甲醇则对甲烷氧化具有强烈的抑制作用，而适当浓度的甲醇可极大地促进土壤甲烷的氧化。在甲烷释放极少的设施栽培土壤中，兼性营养甲烷氧化菌可能在甲烷氧化中占据主导地位。     

Photocatalytic degradation of chlorophenols using Ru(bpy) 3 2+ /S2O 8 2−

Advanced oxidation processes, such as photocatalysed oxidation, provide an important route for degradation of wastes. In this study, the lowest excited state (³MLCT) of Ru(bpy)₃²⁺ is used to break down chlorophenol pollutant molecules to harmless products. This has the advantage of using visible light and a short-lived catalytically active species. Photolysis of deaerated aqueous solutions of a variety of mono- and poly-substituted chlorophenols has been followed in the presence of Ru(bpy)₃²⁺/S₂O₈²⁻ with near visible light (λ > 350 nm) by UV/visible absorption spectroscopy, luminescence, potentiometry, NMR and HPLC techniques. Upon irradiation, a decrease is observed in the chlorophenol concentration, accompanied by the formation of Cl⁻, H⁺ and SO₄²⁻ ions as the main inorganic products. Benzoquinone, phenol, dihydroxybenzenes and chlorinated compounds were the dominant organic products. As the ruthenium(II) complex is regenerated in the reaction, the scheme corresponds to an overall catalytic process. The kinetics of the rapid chlorophenol photodechlorination has been studied, and are described quite well by pseudo-first order behaviour. Further studies on this were made by following Cl⁻ release with respect to the initial Ru(bpy)₃²⁺ and S₂O₈²⁻ concentrations. A comparison is presented of the photodechlorination reactivity of the mono and polychlorophenols studied at acidic and alkaline pH.     

Photochemical reactions of musk ketone

The photolysis of musk ketone in cyclohexane and methanol afforded several cyclization products while the product spectrum in cyclohexene and squalene was characterized by an amino derivative and a ring expansion product, respectively. On cellulose stearate layers the photochemical reactions of musk ketone proceeded readily both under sun simulator and in natural sunlight and several photoproducts identified in solvent experiments were detected. Photoproducts were isolated by preparative HPLC and TLC and analyzed by means of HPLC, GC/MS, MS, LC/MS, ¹H‐ and ¹³C‐NMR.     

Enhanced methane recovery and exoelectrogen-methanogen evolution from low-strength wastewater in an up-flow biofilm reactor with conductive granular graphite fillers

Methane production from low-strength wastewater (LSWW) is generally difficult because of the low metabolism rate of methanogens. Here, an up-flow biofilm reactor equipped with conductive granular graphite (GG) as fillers was developed to enhance direct interspecies electron transfer (DIET) between syntrophic electroactive bacteria and methanogens to stimulate methanogenesis process. Compared to quartz sand fillers, using conductive fillers significantly enhanced methane production and accelerated the start-up stage of biofilm reactor. At HRT of 6 h, the average methane production rate and methane yield of reactor with GG were 0.106 m³/(m³·d) and 74.5 L/kg COD, which increased by 34.3 times and 22.4 times respectively compared with the reactor with common quartz sand fillers. The microbial community analysis revealed that methanogens structure was significantly altered and the archaea that are involved in DIET (such as Methanobacterium) were enriched in GG filler. The beneficial effects of conductive fillers on methane production implied a practical strategy for efficient methane recovery from LSWW.

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Mechanism on minimization of excess sludge in oxic-settling-anaerobic (OSA) process

The oxic-settling-anaerobic (OSA) process is a promising wastewater treatment technique for efficiently reducing sludge production and improving the stability of process operation. In this paper, the possible factors of sludge reduction such as sludge decay, uncoupled metabolism, and anaerobic oxidation with low sludge production were discussed in the OSA process. It has been confirmed that sludge decay is the decisive cause in the OSA process, accounting for 66.7% of sludge production reduction. Sludge decay includes hydrolysis and acidogenesis of dead microorganisms and particle organic carbon adsorbed in sludge floc and endogenous metabolism. By batch experiments, it has been proven that there is energetic uncoupling in the OSA system since microorganisms were exposed to alternative anaerobic and aerobic environment. It accounts for about 7.5% of sludge production reduction. Soluble chemical oxygen demand (SCOD) released from the anaerobic sludge tank in the OSA process was used as the substrate for cryptic growth. The substrate was used for anoxic denitrifying, anaerobic phosphorus release, sulfate reduction, and methane production. These anaerobic reactions in the sludge anaerobic tank have lower sludge production than in the aerobic oxidation when equivalent SCOD is consumed, which may lead to approximately 23% of sludge reduction in the OSA process. It has been concluded that multiple causes resulted in the minimization of excess sludge in the OSA system. The microbial community structure and diversity of sludge samples from the CAS (conventional activated sludge) and OSA systems were investigated by 16 SrDNA PCR-DG-DGGE (polymerase chain reaction-double gradient-denaturing gradient gel electrophoresis). DGGE profile and cluster analysis showed more abundant species in the OSA system contrasting to microbial communities in the CAS system.     

Binding of trinitrotoluene (TNT) to water extractable humus

Solid-state ¹⁵N NMR was applied to the aqueous extracts of a ¹³C-enriched plant slurry (Lolium perenne), anaerobically incubated with ¹⁵N₃-trinitrotoluene (TNT). Almost all ¹⁵N₃-TNT transformation products became covalently bound to the plant-derived organic material extractable with water. DCPMAS ¹⁵N ¹³C NMR revealed a three-step reaction scheme. After reduction of TNT, the aryl amines are acetylated. Subsequent alkylation of the resulting amides strengthens the incorporation of TNT-transformation products into humic material. Comparable results have been recently obtained under aerobic conditions, which indicates that this pathway is a common process during biological TNT transformation.     

Microbial activity, oxygen status and fermentation in the gut of the irregular sea urchin Echinocardium cordatum (Spatangoida: Echinodermata)

Microbial activity in the gut of the detrivore irregular sea urchin Echinocardium cordatum (Pennant) was studied by measurements of oxygen profiles, fermentation end products, C/N ratios, pH and concentrations of sulphide and sulphate in the gut segments, including the nodules of the intestinal caecum. The highest oxygen flux and consumption, and highest concentrations of short-chained fatty acids, and the lowest pH values occurred in the anterior stomach segments, including the anterior caecum. The C/N ratios indicate synthesis of microbial biomass in the caeca. The concentration of sulphate was high in the anterior stomach segments, whereas sulphide was only detectable in the nodules of the intestinal caecum. The anterior and intestinal caeca were the major sites for microbial activity, and oxidation of acetate and propionate produced in the two caeca corresponded to at least 9% of the total respiration of the sea urchin. Microbial fermentation, especially in the anterior caecum, seems to be important for the metabolism of E. cordatum, allowing the sea urchin to utilise refractile carbohydrates. The functions of the intestinal caecum are probably both residual fermentation and oxidation of accumulated sulphide. Received: 25 February 1998 / Accepted: 30 June 1998     

Microbial community structure and dynamics of starch-fed and glucose-fed chemostats during two years of continuous operation

The microbial community structures of two mesophilic anaerobic chemostats, one fed with glucose, the other with starch as sole carbon sources, were studied at various dilution rates (0.05–0.25 d^–1 for glucose and 0.025–0.1 d^–1 for starch) during two years continuous operation. In the glucose-fed chemostat, the aceticlastic methanogen Methanosaeta spp. and hydrogenotrophic methanogen Methanoculleus spp. predominated at low dilution rates, whereas Methanosaeta spp. and the hydrogenotrophic Methanobacterium spp. predominated together when dilution rates were greater than 0.1 d^–1. Bacteria affiliated with the phyla Bacteroidetes, Spirochaetes, and Actinobacteria predominated at dilution rates of 0.05, 0.1, and 0.15 d^–1, respectively, while Firmicutes predominated at higher dilution rates (0.2 and 0.25 d^–1). In the starch-fed chemostat, the aceticlastic and hydrogenotrophic methanogens coexisted at all dilution rates. Although bacteria belonging to only two phyla were mainly responsible for starch degradation (Spirochaetes at the dilution rate of 0.08 d^–1 and Firmicutes at other dilution rates), different bacterial genera were identified at different dilution rates. With the exception of Archaea in the glucose-fed chemostat, the band patterns revealed by denaturing gradient gel electrophoresis (DGGE) of the microbial communities in the two chemostats displayed marked changes during long-term operation at a constant dilution rate. The bacterial community changed with changes in the dilution rate, and was erratic during longterm operation in both glucose-fed and starch-fed chemostats.     

Reduction of hexavalent chromium using fungi and bacteria isolated from contaminated soil and water samples

The reductive adsorption of hexavalent chromium (Cr⁶⁺) using six indigenous microorganisms isolated from contaminated soil and water samples was investigated. Quantification of Cr⁶⁺ reduction was determined using the 1,5-diphenylcarbazide method followed measuring the absorbance at OD₅₄₀. Bacterial isolates identified as Klebsiella pneumoniae, Bacillus firmus and Mycobacterium sp. were capable of absorbing Cr⁶⁺ efficiently into their biomass, whereas the fungal isolates, Aspergillus flavus, Aspergillus sp. and A. niger were capable of transforming Cr⁶⁺ to Cr³⁺ relative to cell-wall-binding properties. Infrared spectral analysis of functional groups showed that ?OH, ?NH₂ and C?O with conjugated ?NH were the binding groups responsible for adsorption of Cr⁶⁺ within the biomass of isolates. The data highlight the promising biotechnological application of these isolates in removing carcinogenic and mutagenic Cr⁶⁺ from contaminated ecosystems.     

Seasonal variation of microbial populations and biomass in Tatachia grassland soils of Taiwan

To investigate the seasonal variations of microbial ecology in grassland of Tatachia forest, soil properties, microbial populations, microbial biomass, and 16S rDNA clone library analysis were determined. The soil had temperatures 6.6–18.4°C, pH 3.6–5.1, total organic carbon 1.11–10.68%, total nitrogen 0.18–0.78%, and C/N ratios 3.46–20.55. Each gram of dry soil contained bacteria, actinomycetes, fungi, cellulolytic, phosphate-solubilizing microbes, and nitrogen-fixing microbes 4.54 × 10⁴ to 3.79 × 10⁷, 3.43 × 10² to 2.17 × 10⁵, 5.74 × 10³ to 3.76 × 10⁶, 1.97 × 10³ to 1.34 × 10⁶, 8.49 × 10² to 5.59 × 10⁵, and 3.86 × 10² to 3.75 × 10⁵ CFU, respectively. Each gram of soil contained 117–2,482 μg of microbial biomass carbon, 23–216 μg of microbial biomass nitrogen and 9–29 μg of DNA. The microbial populations, microbial biomass, and DNA decreased stepwise with the depth of soil, and they had low values in winter seasons. The microbial populations, microbial biomass carbon, microbial biomass nitrogen, and DNA at the BW2 horizon were 8.42–17.84, 19.26–64.40, 16.84–61.11, and 31.03–46.26% of those at the O horizon, respectively. When analyzing 16S rDNA library, members of Proteobacteria, Acidobacteria, Actinobacteria, Bacteroidetes, Chloroflexi, Firmicutes, candidate division TM1, candidate division TM7, Gammatimonadetes, and Verrucomicrobia were identified. Members of Proteobacteria (44.4%) and Acidobacteria (33.3%) dominated the clone libraries. Within the phylum Proteobacteria, α-, β-, and γ-Proteobacteria were most numerous, followed by δ-Proteobacteria.     

Enhancement of radon exposure in smoking areas

Radium-226 is a significant source of radon-222 which enters buildings through soil, construction materials or water supply. When cigarette smoke is present, the radon daughters attach to smoke particles. Thus, the alpha radiation to a smoker’s lungs from the natural radon daughters is increased because of smoking. To investigate whether the cigarette tobacco itself is a potential source of indoor radon, the α potential energy exposure level contents of radon (²²²Rn, 3.82d) and Thoron (²²⁰Rn, 55.60s) were measured in 10 different cigarette tobacco samples using CR-39 solid-state nuclear track detectors (SSNTDs). The results showed that the ^{222, 220}Rn concentrations in these samples ranged from 128 to 266 and 49 to 148 Bqm⁻³, respectively. The radon concentrations emerged from all investigated samples were significantly higher than the background level. Also, the annual equivalent doses from the samples were determined. The mean values of the equivalent dose were 3.51 (0.89) and 1.44 (0.08) mSvy⁻¹, respectively. Measurement of the average indoor radon concentrations in 20 café rooms was, significantly, higher than 20 smoking-free residential houses. The result refers to the dual (chemical and radioactive) effect of smoking as a risk factor for lung cancer.     

Vacuum promotes metabolic shifts and increases biogenic hydrogen production in dark fermentation systems

The successful operation of any type of hydrogen-producing bioreactor depends on the performance of the microorganisms present in the system. Both substrate and partial gas pressures are crucial factors affecting dark fermentation metabolic pathways. The main objective of this study was to evaluate the impact of both factors on hydrogen production using anaerobic granular sludge as inoculum and, secondly, to study the metabolic shifts of an anaerobic community subjected to low partial gas pressures. With this goal in mind, seven different wastewater (four synthetic media, two industrial wastewater, and one domestic effluent) and the effect of applying vacuum on the systems were analyzed. The application of vacuum promoted an increase in the diversity of hydrogenproducing bacteria, such as Clostridium, and promoted the dominance of acetoclastic- over hydrogenotrophic methanogens. The application of different media promoted a wide variety of metabolic pathways. Nevertheless, reduction of the hydrogen partial pressure by application of vacuum lead to further oxidation of reaction intermediates irrespective of the medium used, which resulted in higher hydrogen and methane production, and improved the COD removal. Interestingly, vacuum greatly promoted biogenic hydrogen production from a real wastewater, which opens possibilities for future application of dark fermentation systems to enhance biohydrogen yields.     

Interaction and independence on methane oxidation of landfill cover soil among three impact factors: water, oxygen and ammonium

To understand the influence patterns and interactions of three important environmental factors, i.e. soil water content, oxygen concentration, and ammonium addition, on methane oxidation, the soils from landfill cover layers were incubated under full factorial parameter settings. In addition to the methane oxidation rate, the quantities and community structures of methanotrophs were analyzed to determine the methane oxidation capacity of the soils. Canonical correspondence analysis was utilized to distinguish the important impact factors. Water content was found to be the most important factor influencing the methane oxidation rate and Type II methanotrophs, and the optimum value was 15% (w/w), which induced methane oxidation rates 10- and 6- times greater than those observed at 5% (w/w) and 20% (w/w), respectively. Ambient oxygen conditions were more suitable for methane oxidation than 3% oxygen. The addition of 100 mg-N·kg _drysoil ^?1 of ammonium induced different effects on methane oxidation capacity when conducted at low or high water content. With regard to the methanotrophs, Type II was sensitive to the changes of water content, while Type I was influenced by oxygen content. Furthermore, the methanotrophic acidophile, Verrucomicrobia, was detected in soils with a pH of 4.9, which extended their known living environments.     

Soil organic matter chemistry. Part 1. Characterization of several humic preparations by proton and carbon‐13 nuclear magnetic resonance spectroscopy

Both laboratory and commercial preparations of humic substances (HSs) such as fulvic acids and humic acids along with HC1‐HF preparation of Manitoba peat soil organic matter were characterized using Fourier Transformation (FT) proton (¹H) and carbon‐13 (¹³C) nuclear magnetic resonance (NMR) spectroscopy. All the samples were dissolved in a solution of 0.4 N NaOD in D₂O. In the case of ‘H‐NMR spectroscopy, all the investigated humic samples displayed resonance absorption peaks in the region of 1–4 ppm indicating the likely presence of aliphatic protons in the preparations. However, with the exception of one fulvic acid preparation (extracted from Manitoba Carrol clay‐loam soil with 0.5 N NaOH), ¹H‐NMR spectra of all other samples provided evidence for strong aromatic character. The aliphatic and aromatic characteristics of such samples of HSs were further confirmed with the aid of ¹³C‐NMR spectra.