In vitro demonstration of anaerobic oxidation of methane coupled to sulphate reduction in sediment from a marine gas hydrate area

Anaerobic oxidation of methane (AOM) and sulphate reduction were examined in sediment samples from a marine gas hydrate area (Hydrate Ridge, NE Pacific). The sediment contained high numbers of microbial consortia consisting of organisms that affiliate with methanogenic archaea and with sulphate-reducing bacteria. Sediment samples incubated under strictly anoxic conditions in defined mineral medium (salinity as in seawater) produced sulphide from sulphate if methane was added as the sole organic substrate. No sulphide production occurred in control experiments without methane. Methane-dependent sulphide production was fastest between 4 degree C and 16 degree C, the average rate with 0.1 MPa (approximately 1 atm) methane being 2.5 micro mol sulphide day(-1) and (g dry mass sediment)(-1). An increase of the methane pressure to 1.1 MPa (approximately 11 atm) resulted in a four to fivefold increase of the sulphide production rate. Quantitative measurements using a special anoxic incubation device without gas phase revealed continuous consumption of dissolved methane (from initially 3.2 to 0.7 mM) with simultaneous production of sulphide at a molar ratio of nearly 1:1. To test the response of the indigenous community to possible intermediates of AOM, molecular hydrogen, formate, acetate or methanol were added in the absence of methane; however, sulphide production from sulphate with any of these compounds was much slower than with methane. In the presence of methane, such additions neither stimulated nor inhibited sulphate reduction. Hence, the experiments did not provide evidence for one of these compounds acting as a free extracellular intermediate (intercellular shuttle) during AOM by the presently investigated consortia.     

好氧和缺氧条件下游离亚硝酸对氨氧化菌和亚硝酸盐氧化菌的选择性抑制

【背景】稳定短程硝化是实现城市污水厌氧氨氧化技术的瓶颈,目前国内外关于游离亚硝酸(Free nitrous acid,FNA)对硝化菌活性的影响大多是在曝气条件下进行研究,鲜有关于缺氧条件下FNA对硝化菌活性影响的报道。【目的】探究好氧和缺氧下FNA对氨氧化菌(Ammonia oxidizing bacteria,AOB)和亚硝酸盐氧化菌(Nitrite oxidizing bacteria,NOB:Nitrospira和Nitrobacter)活性的抑制影响。【方法】采用序批式反应器(Sequencing batch reactor,SBR),基于混合液悬浮固体浓度(Mixed liquid suspended solids,MLSS)为8 300 mg/L的全程硝化污泥条件,通过批次试验分别考察好氧和缺氧下FNA(初始浓度为1.16 mg/L)处理48 h后,AOB和NOB活性的变化情况。【结果】好氧FNA处理活性污泥48 h后,FNA浓度维持在1.16-1.17 mg/L,游离氨(Free ammonia,FA)浓度小于0.017 mg/L,AOB、Nitrospira、Nitrobacter丰度均未发生明显变化;过曝气至99 h时,与空白组相比,比氨氮氧化速率(r~+_(NH4-N))、比亚硝酸盐氮氧化速率(r_(NO2-N))均出现小幅下降,分别由3.5、4.828 mg N/(g VSS·h)降至3.3、4.668 mg N/(g VSS·h),且亚硝酸盐氮累积率(Nitrite accumulation rate,NAR)始终低于33.2%。缺氧FNA处理活性污泥48 h后,FNA浓度维持在0.64-1.16 mg/L,FA浓度低于0.039 mg/L,AOB丰度变化较小,而Nitrospira、Nitrobacter丰度均明显下降,分别由3.002 9×10~9、4.245×10~8 copies/g VSS降至1.666 5×10~8、5.163 8×10~7 copies/g VSS;过曝气至99 h时,与空白组相比,r~+_(NH4-N)值下降幅度较小,而r_(NO2-N)值明显降低,由4.828 mg N/(g VSS·h)降至0.007 mg N/(g VSS·h),且在过曝气0-292 h内,NAR均大于94%。【结论】好氧FNA处理活性污泥48 h后对AOB和NOB无明显抑制作用,但缺氧FNA处理活性污泥48 h后对AOB具有轻微抑制作用,而对NOB具有强烈的抑制作用,可以实现稳定的短程硝化。     

Sulphur metabolism in Thiorhodaceae. II. stoichiometric relationship of CO2 fixation to oxidation of hydrogen sulphide and intracellular sulphur inChromatium okenii

Stoichiometry of sulphide and intracellular sulphur oxidation in connection with CO₂ fixation was studied inChromatium okenii. The equipment used was a special stirred cuvette with a rapid-sampling arrangement, which allowed short-time experiments with illuminated bacterial suspensions under anaerobic conditions. Turnover of the sulphur compounds is controlled by a linear CO₂ fixation rate which amounts to 0.069µmoles of CO₂/min mg of cell protein at light saturation. Van Niel's equations for bacterial photosynthesis could be confirmed for short periods under the condition that sulphate is produced during increase of intracellular sulphur; i.e., oxidation of sulphide and of intracellular sulphur do not occur consecutively but simultaneously. The full oxidation rate of intracellular sulphur starts after complete consumption of sulphide. The time during which sulphide is oxidized to intracellular sulphur amounts to 1/3–1/4 of the time necessary for the complete quantitative oxidation of the sulphide to sulphate.     

Biotechnological process for sulphide removal with sulphur reclamation

A new biotechnological process for sulphide removal is proposed. The process is based on the oxidation of sulphide into elemental sulphur, which can be removed by sedimentation. In this study it was found that elemental sulphur and sulphate are the main oxidation products of the biological sulphide oxidation. The settling characteristics become worse as the sulphide concentration increases, due to polysulphide formation. The start-up phase of this biological system is very short; Only four days are needed to reduce the sulphide concentration of 100 to 2 mg/l at a HRT (Residence time) of 22 minutes. Also some environmental factors were evaluated. The optimal pH is situated in the pH-range 8.0–8.5. Significantly lower conversion rates are found at pH = 6.5 to 7.5 and pH = 9.0, while at pH = 9.5 the sulphide oxidation capacity of the system detoriates. The process temperature was 20°C, although the optimal temperature is situated in the range 25–35°C. No substrate inhibition of sulphide was found at sulphide concentrations up to 100 mg/l.     

Biological sulphide oxidation in a fed-batch reactor

This study shows that, in a sulphide-oxidizing bioreactor with a mixed culture of Thiobacilli, the formation of sulphur and sulphate as end-products from the oxidation of sulphide can be controiledinstantaneously and reversibiy by the amount of oxygen supplied. It was found that at sulphide loading rates of up to 2.33 mmol7/L . h, both products can be formed already at oxygen concentrations below 0.1 mg/L. Because the microorganisms tend to form sulphate rather than forming sulphur, the oxygen concentration is not appropriate to optimize the sulphur production. Within less than 2 h, the system can be switched reversibly from sulphur to sulphate formation by adjusting the oxygen flow. This is below the minimum doubling time (2.85 h) of, e.g., Thiobacillus neapolitanus and Thiobacillus 0,(18) which indicates that one metabolic type of organism can probably perform both reactions. Under highly oxygen-limited circumstances, that is, at an oxygen/sulphide consumption ratio below 0.7 mol . h(-1) mol . h(-1) thiosulphate is abundantly formed. Because the chemical sulphide oxidation results mainly in the formation of thiosulphate, it is concluded that, under these circumstances, the biological oxidation capacity of the system is lower than the chemical oxidation capacity. The oxidation rate of the chemical sulphide oxidation can be described by a first-order process (k =-0.87 h(-1)).(c) 1995 John Wiley & Sons, Inc.     

Optimal operational factors for nitrite accumulation in batch reactors

The environmental factors that affected the accumulation of nitrite in nitrifying reactors were investigated using a mixed culture. A batch reactor with 50 mg-N/l of ammonia was used. The pH, temperature and dissolved oxygen concentration were varied. The concentration of unionized free ammonia also changed with the oxidation of ammonia and the variation of pH and temperature. The accumulation of nitrite was affected sensitively by pH and temperature. A higher nitrite concentration was observed at pH 8-9 or temperature around 30 °C. The dissolved oxygen also affected, giving the highest nitrite accumulation at around 1.5 mg/l. These were the favoredconditions for nitrite production. The free ammonia concentration influenced thenitrite accumulation also, by inhibiting nitrite oxidation. The inhibition becameapparent at a concentration of approximately 4 mg/l or above, but insignificant atbelow 1 mg/l. Thus, simultaneously high free ammonia concentration and maximumspecific ammonia-oxidation rate (above 15 × 10^-3 mg-N/mg-VSSh)were needed for a significant nitrite accumulation. When the two conditions were met, thenthe highest accumulation was observed when the ratio of the maximum specific oxidationrate of ammonia to the maximum specific oxidation rate of nitrite (k_a/k_n) was highest.Under the optimal operating conditions of pH 8, 30 °C and 1.5 mg/l of dissolvedoxygen, as much as 77% of the removed ammonia accumulated in nitrite.     

Sulphide formation and chemical composition of bottom sediments of some Italian lakes

Studies of sulphate reduction and rates of sulphide formation were made in the bottom sediments of the alpine lakes Lago Maggiore and Lago Lugano. The stock of sulphide sulphur was found to be 500–1500 mg/l. The rate of sulphate reduction was 1–10 mg S/l/day. Total numbers of bacteria in sediments varied from 0,5 to 5.10⁹ cells/cm³ of wet mud. Chemical analyses of the carbon, nitrogen and phosphorus were also made. The possible influence of pollution on the sulphur cycle in these lakes is discussed.     

The Effect of Phenols and Heterocyclic Bases on Nitrification in Activated Sludges

S ummary . Activated sludge receiving ammonium thiocyanate (500 mg/l) was able to nitrify. The rate of ammonia oxidation was decreased when >3 mg/l of phenol (or cresols) was added to the sludge, and at 10 mg/l was inhibited completely. Concentrations of up to 100 mg/l of phenols did not affect nitrite oxidation. The 2- and 4-methyl pyridine derivatives inhibited both ammonia and nitrite oxidation.     

Propionate-driven sulphate-reduction by oil-field bacteria in a pressurised porous rock bioreactor

The occurrence of high concentrations (up to 4900 mg/l) of anions of acetic, propionic and butyric acids in co-produced water from oil reservoirs represents a large pool of potential electron donors for bacterial sulphate reduction. Enrichment cultures in defined media, isolated from a variety of oil-filed environments demonstrated the wide distribution of acetate-and propionate-utilising sulphate-reducing bacteria (SRB). A propionate-utilising enrichment culture consisting predominantly of SRB, tentatively identified as species of Desulfobulbus, was used to inoculate a pressurised porous rack bioreator operating under simulated reservoir conditions. Using a flood velocity of 6.3 cm/h with an inlet propionate concentration of 168 mg/l, a sulphide generation rate of 2.5 g/ml of rock per hour was achieved at 30° C and 20 MPa. This rate indicates that a sphere of reservoir rock 9.3 m in radius, colonised with propionate-utilising SRB, could produce 50 kg sulphide day. The rates of propionate-driven bacterial sulphate reduction observed in the porous rock bioreactor could sustain the H₂S production rates observed from wells in souring reservoirs. Correspondence to: I. Vance     

Free nitrous acid inhibition on anoxic phosphorus uptake and denitrification by poly-phosphate accumulating organisms

Nitrite has been found in previous research an inhibitor on anoxic phosphorus uptake in enhanced biological phosphorus removal systems (EBPR). However, the inhibiting nitrite concentration reported varied in a large range. This study investigates the nitrite inhibition on anoxic phosphorus uptake by using four different mixed cultures performing EBPR with pH considered an important factor. The results showed that the protonated species of nitrite, HNO(2) (or free nitrous acid, FNA), rather than nitrite, is likely the actual inhibitor on the anoxic phosphorus uptake, as revealed by the much stronger correlation of the phosphorus uptake rate with the FNA than with the nitrite concentration. All the four EBPR sludges showed decreased anoxic phosphorus uptake rates with increased FNA concentrations in the studied range of 0.002-0.02 mg HNO(2)-N/L. The phosphorus uptake by all four cultures was completely inhibited at 0.02 mg HNO(2)-N/L. Granular sludge appeared to be more tolerant to HNO(2) than flocular sludge likely due to its stronger resistance to the transfer of nitrite into the bacterial aggregates. Furthermore, denitrification by the phosphorus-accumulating organisms (PAOs) was also found to be inhibited by HNO(2). The denitrification rate decreased by approximately 40% when the FNA concentration was increased from 0.002 to 0.02 mg HNO(2)-N/L.     

Dynamics of corrosion rates associated with nitrite or nitrate mediated control of souring under biological conditions simulating an oil reservoir

Representative microbial cultures from an oil reservoir and electrochemical techniques including potentiodynamic scan and linear polarization were used to investigate the time dependent corrosion rate associated with control of biogenic sulphide production through addition of nitrite, nitrate and a combination of nitrate-reducing, sulphide-oxidizing bacteria (NR-SOB) and nitrate. The addition of nitrate alone did not prevent the biogenic production of sulphide but the produced sulphide was eventually oxidized and removed from the system. The addition of nitrate and NR-SOB had a similar effect on oxidation and removal of sulphide present in the system. However, as the addition of nitrate and NR-SOB was performed towards the end of sulphide production phase, the assessment of immediate impact was not possible. The addition of nitrite inhibited the biogenic production of sulphide immediately and led to removal of sulphide through nitrite mediated chemical oxidation of sulphide. The real time corrosion rate measurement revealed that in all three cases an acceleration in the corrosion rate occurred during the oxidation and removal of sulphide. Amendments of nitrate and NR-SOB or nitrate alone both gave rise to localized corrosion in the form of pits, with the maximum observed corrosion rates of 0.72 and 1.4 mm year⁻¹, respectively. The addition of nitrite also accelerated the corrosion rate but the maximum corrosion rate observed following nitrite addition was 0.3 mm year⁻¹. Furthermore, in the presence of nitrite the extent of pitting was not as high as those observed with other control methods.     

Optimization of sulphide production in an anaerobic continuous biofilm process with sulphate reducing bacteria

Different biofilm reactors for sulphide production by sulphate reducing bacteria were compared in a packed bed reactor and in two suspended carrier biofilm reactors. Lactate was used as carbon source in the experiments. The process was reversibly inhibited by free sulphide at 14 mM. The packed bed reactor was more efficient and less sensitive to changes. The maximum load in the system was 5.3 g sulphate/l.d.     

Multi-stage chemostat investigation of interspecies interactions in a hexanoate-catabolizing microbial association isolated from anoxic landfill

Spatial separations of physico-chemical environments (habitat domains) with maintenance of overlapping zones of influence (activity domains) with multi-stage chemostat models were used to facilitate examination of interspecies interactions within a hexanoate-catabolizing microbial association isolated from anoxic landfill. Under a constant dilution rate regime, in the presence of 1.4 and 5 mmol/l influent sulphate, consolidation of the major metabolic events of hexanoate catabolism, sulphate reduction, acetogenesis and methanogenesis took place in the first vessel of a five-vessel array suggesting considerable overlap of activity domains or localization around the habitat domains. Evidence of partial competitive displacement was not apparent until 10 mmol/l of sulphate was used. Introduction of a non-constant dilution rate regime to a three-vessel model, subjected to an influent sulphate concentration of 1.4 mmol/l, effected the displacement of the methanogenic species and, as a consequence, in the presence of sulphate limitation, CO₂ reduction became the major sink for excess electrons generated from hexanoate oxidation. A hypothetical scheme of anaerobic hexanoate catabolism by the interacting microbial association was developed.     

Sulphate reduction and sulphur cycling in lake sediments: a review

1. The concentration of sulphate is low in lakes and sulphur cycling has often been neglected in studies of organic matter diagenesis in lake sediments. The cycling of sulphur is, however, both spatially and temporally dynamic and strongly influences many biogeochemical reactions in sediments, such as the binding of phosphorus. This review examines the control of sulphate reduction and sulphur cycling in sediments of lakes with different trophic status. 2. The factors that control the rate of sulphate reduction have not been identified with certainty in the various environments because many factors are involved, e.g. oxygen and sulphate concentrations, temperature and organic matter availability. 3. Sulphate reduction is less significant under oligotrophic conditions, where mineralization is dominated by oxic decomposition. The supply of organic matter may not be sufficient to support sulphate reduction in the anoxic parts of sediments and, also, sulphate availability may control the rate as the concentration is generally low in oligotrophic lakes. 4. There is a potential for significant sulphate reduction in eutrophic lakes, as both the availability of organic matter and sulphate concentration are often higher than in oligotrophic lakes. Sulphate is rapidly depleted with sediment depth, however, and methanogenesis is generally the most important process in overall carbon mineralization. Sulphate reduction is generally low in acidic lakes because of low sulphate availability and reduced microbial activity. 5. It is still unclear which of the forms of sulphur deposits are the most important and under which conditions burial occurs. Sulphur deposition is controlled by the rate of sulphate reduction and reoxidation. Reoxidation of sulphides occurs rapidly through several pathways, both under oxic and anoxic conditions. Only a few studies have been able to examine the importance of reoxidation, but it is hypothesized that most of the reoxidation takes place under anoxic conditions and that disproportionation is often involved. The presence of sulphide oxidizing bacteria, benthic fauna and rooted macrophytes may substantially enhance oxic reoxidation. Deposition of sulphur is generally higher in eutrophic than in oligotrophic lakes because of a number of factors: a higher rate of sulphate reduction, enhanced sedimentation of organic sulphur and less reoxidation as a result of reduced penetration of oxygen into the sediments, a lack of faunal activity and rooted macrophytes.     

Effect of pH on anoxic sulfide oxidizing reactor performance

The effects of pH on the performance of anoxic sulfide oxidizing (ASO) reactor were evaluated. Performance was investigated under various operational conditions at influent pH range of 4-11. At the influent pH of 7-7.5 during loading tests and HRT tests, the sulfide oxidation was partial. In general, the amount of sulfate formed decreased with the increasing sulfide and nitrite loadings. The bacterial communities in ASO reactors were more sensitive to acidic pH compared with alkaline pH, as nitrite and sulfide removal rates dropped significantly when exposed to acidic pH 3. High dissolved bisulfide ions, nitrite and excess of sulfate (>300 mg/L) might have inhibited the sulfide oxidation under highly acidic and alkaline conditions in the ASO reactor. Based on sulfide and nitrite removal efficiencies, the ASO reactor can be operated in a wide range of pH, i.e. 5-11.     

Degradation of phenol and phenolic compounds by a defined denitrifying bacterial culture

Phenol, a major pollutant in several industrial waste waters is often used as a model compound for studies on biodegradation. This study investigated the anoxic degradation of phenol and other phenolic compounds by a defined mixed culture of Alcaligenes faecalis and Enterobacter species. The culture was capable of degrading high concentrations of phenol (up to 600 mg/l) under anoxic conditions in a simple minimal mineral medium at an initial cell mass of 8 mg/l. However, the lag phase in growth and phenol removal increased with increase in phenol concentration. Dissolved CO₂ was an absolute requirement for phenol degradation. In addition to nitrate, nitrite and oxygen could be used as electron acceptors. The kinetic constants, maximum specific growth rate _max; inhibition constant, K _i and saturation constant, K _s were determined to be 0.206 h^–1, 113 and 15 mg phenol/l respectively. p-Hydroxybenzoic acid was identified as an intermediate during phenol degradation. Apart from phenol, the culture utilized few other monocyclic aromatic compounds as growth substrates. The defined culture has remained stable with consistent phenol-degrading ability for more than 3 years and thus shows promise for its application in anoxic treatment of industrial waste waters containing phenolic compounds.     

Kinetics of denitrification using sulphur compounds: effects of S/N ratio, endogenous and exogenous compounds

The influence of different sulphur to nitrogen (S/N) ratios on the specific autotrophic denitrification activity was studied in batch experiments using thiosulphate and nitrate as substrates. Transitory accumulations of nitrite were observed for assays with S/N ratios of 3.70 and 6.67 g/g, probably due to the higher specific reduction rate of nitrate compared to that of nitrite. Nitrite was the main end product when S/N ratios of 1.16 and 2.44 g/g were tested. The effects of endogenous (NO(3)(-),NO(2)(-),S(2)O(3)(2-)and SO(4)(2-)) and exogenous compounds (acetate and NaCl) on the specific denitrifying activity of the sludge were tested. Nitrite and sulphate did exert clear inhibitory effects over the process while thiosulphate, acetate and NaCl did not have strong effects at the concentrations tested. Similar experiments also showed that sulphur was not a suitable electron donor for these microorganisms, but sulphide was used successfully.     

Bio catalytic oxidation of sulphide laden wastewater from leather industry using sulfide: Quinone oxidoreductase immobilized bio reactor

Abstract

Anaerobic treatment of sulphate rich wastewater results in high amount of sulphide in liquid phase and gaseous phase. Sulphide is malodorous in gaseous phase and toxic even at very low concentrations in liquid phase and causes objectionable environmental issues. In the present investigation the sulphide present in the UASB treated post tanning wastewater was oxidised into elemental sulphur using Sulfide: Quinone oxidoreductase (SQR) immobilized Functionalized Carbon-silica matrix (FCSM) packed bed reactor. The variables employed for the production of Bacillus clausii biomass for the extraction of SQR were optimized using RSM. The purified SQR showed the maximum activity and stability at pH 6.0 to 8.0 and the percentage oxidation of sulphide at HRT of 24?h were 99.2%?±?0.2 at pH 6.0; 99.6%?±?0.2 at pH 7.0 and 99.6%?±?2.12 at pH 8.0. The effect of temperature on SQR activity was optimized and it established the maximum activity and stability at 40?°C with sulphide oxidation of 99.6%?±?0.8%. The optimum conditions for immobilization of SQR onto FCSM were time, 240?min; pH, 7.0; temperature, 40?°C and SQR concentration, 10?mg/g. The immobilization of SQR onto FCSM obeyed the Langmuir isotherm model. The immobilization of SQR onto FCSM was confirmed by SEM, FT-IR, XRD, TGA and DSC analyses. The SQR-FCSM packed bed reactor was operated separately for the oxidation of sulfide in UASB treated post-tanning wastewater under continuous mode at different HRTs and it recorded the maximum sulphide oxidation by 99?±?0.1% at HRT of 15?h with residual sulphide of 2.4?±?1.1?mg/L. The formation of elemental Sulphur was confirmed by XRD studies. The present investigation provides the scope for the removal of sulphide and thereby substantial reduction in Total Dissolved Solids from post tanning wastewater without addition of chemicals.