硫酸盐还原菌的分离纯化方法

硫酸盐还原菌在硫元素的地球生物循环及环境保护中都发挥着重要作用。介绍了硫酸盐还原菌的分离与纯化的方法,比较了厌氧袋、厌氧罐、厌氧工作站、厌氧管等方法的应用,以及在使用中需要注意的问题;同时介绍了分子生物学技术在硫酸盐还原菌分离纯化方面的应用,旨在为硫酸盐还原菌的分离纯化提供一定的参考依据。     

锅炉管道回水、厌氧污泥中硫酸盐还原菌的筛选

目的研究硫酸盐还原菌及其筛选、分离鉴定体系.方法在锅炉管道回水和厌氧污泥中驯化、筛选硫酸盐还原能力强的菌株,以铬酸钡光度方法检测液体培养基中的硫酸根含量,测定了菌株的硫酸盐降解能力.结果在水样A和泥样B中,A1和B2菌株能够较好地降解培养基中的硫酸盐,通过计数得出A1和B2的最大可能菌数分别为1.65×10(6)个/ml、2.25×10(6)个/ml.并利用改进的稀释涂布-叠皿夹层培养方法分离鉴定A1和B2菌株,观察硫酸盐还原菌的形态,为进一步了解SRB的生长特性,进而从根本上解决硫酸盐还原菌腐蚀问题提供理论基础.结论经过实验,所筛选分离鉴定体系可行.     

不同pH值条件下硫酸盐还原菌组成及硫酸盐还原机制分析

【目的】从海洋沉积物中富集获得硫酸盐还原菌群,改变pH值进行培养,分析pH值对硫酸盐还原性质的影响,明确菌群组成和进行硫酸盐还原功能基因预测,探究硫酸盐还原机制。【方法】分析硫酸盐还原菌群在不同pH值条件下的硫酸盐还原率,在此基础上,利用高通量测序技术和PICRUSt软件分析硫酸盐还原菌群优势菌组成及硫酸盐还原相关基因相对丰度。【结果】硫酸盐还原菌群在不同pH值培养条件下的生长和硫酸盐还原率出现显著变化(P<0.01),在pH 5.0时达到峰值,分别为0.34±0.01和96.52%±0.44%。高通量测序数据显示,pH 5.0时菌群丰富度和多样性最高,优势菌属为假单胞菌(Pseudomonas)和芽孢杆菌(Bacillus),相对丰度较高的基因为同化性硫酸盐还原相关基因。【结论】硫酸盐还原菌富集生长的最适pH 5.0,在此条件下的高硫酸盐还原率由同化性硫酸盐还原途径主导,为揭示硫酸盐还原机制提供了实验支持,并拓宽了硫酸盐还原菌实践应用方面的种质资源。     

铁还原菌降解石油烃的研究进展

铁还原菌是指能够利用细胞外Fe(III)作为末端电子受体,通过氧化有机物将Fe(III)还原为Fe(II)微生物的总称。铁还原作用广泛存在于土壤、河流、海洋、地表含水层以及高温高压的地下深部油藏。在厌氧或兼性厌氧条件下,Fe(III)还原耦合有机物的降解,对铁、碳元素的生物地球化学循环具有重要意义。本文介绍了铁还原菌的多样性和铁还原作用机理,综述了铁还原菌在石油烃降解方面的研究进展。此外,还总结了铁还原菌在生物修复中的潜在作用,并对未来的研究方向进行了展望。     

用硫酸盐还原菌处理重金属废水的研究

介绍了用硫酸盐还原菌处理重金属废水的几种主要方法和原理。硫酸盐还原菌处理含重金属废水主要是通过将可溶性的重金属离子转化成不溶性的金属硫化物、氢氧化物、碳酸盐的方式 ,或直接通过以菌体对重金属离子的吸附完成的。目前研究用硫酸盐还原菌处理重金属废水的主要方法有分批沉淀工艺、吸附处理工艺、化学法和硫酸盐还原菌的混合工艺、全混合处理工艺及硫酸盐还原菌的厌氧上流式污泥床和流化床工艺 ,并对其主要的工艺指标进行了比较。     

一种肠道硫酸盐还原菌的增菌培养基

目的针对已经分离、纯化的肠道硫酸盐还原菌,建立一种能快速、高效地培养菌体的培养基。方法比较营养丰富的GAM肉汤与常用于培养硫酸盐还原菌的选择性培养基Postgate的培养效果,摸索在GAM肉汤中添加不同浓度的硫酸盐对两种肠道硫酸盐还原菌-Desulfovibrio desulfuricans和Desulfovibrio intestina—zis的培养效果。确定效果最佳的改良GAM培养基配方,并测定在该培养基中D．desulful＇icans的生长曲线。结果与Postgate培养基相比,GAM肉汤能在2d内快速培养D．desulfugicans,但培养至6d时细菌数量大幅降低。在GAM肉汤中添加Na2SO4与FeSO4,在实验浓度范围内,均显著地促进硫酸盐还原菌的生长。在此基础上改良GAM肉汤培养基,培养得到的细菌数量较GAM肉汤显著提高。D．desulfuricans的生长曲线显示,2d时细菌生长达到最高峰,数量可达3．5×10^7 CFU／mL;培养6d,细菌数量为7．3×10^6 CFU／mL。结论基于GAM肉汤改良而得到的增菌培养基,能快速、高效地培养肠道硫酸盐还原菌,为后续进一步研究肠道硫酸盐还原菌的生理功能提供了支持。     

硫酸盐还原菌及其还原解毒Cr(Ⅵ)的研究进展

硫酸盐还原菌是一类分布广泛, 能进行硫酸盐异化还原反应的严格厌氧菌。利用硫酸盐还原菌可去除环境中的许多污染物, 因而该类细菌在环境污染治理中具有广阔的应用前景。本文介绍了硫酸盐还原菌的生物学特性和代谢特征及其在环境污染治理中的应用, 并对硫酸盐还原菌还原解毒Cr(Ⅵ)及应用于含Cr(Ⅵ)废水处理的研究进展作了综述, 分析了其未来的研究方向。     

硫酸盐还原菌净化工业废水的研究

本文报道了硫酸盐还原菌的菌学特征,以其具有吸附和絮凝作用,使工业废水得到净化。用硫酸盐还原菌处理印染废水,废水的脱色率为92．5％,COD(cr)和BOD5亦达到排放标准。对城市生活废水和含铬的电镀废水亦有很好的处理效果,各项指标分别达到了排放标准,其中净化后的电镀废水还可作为循环水使用。     

硫酸盐还原菌检测与应用研究进展

硫酸盐还原菌是自然界存在最为广泛的细菌之一,近年来该类微生物研究日益受到国内外学者的关注。本文对硫酸盐还原菌的鉴定、量化和应用等方面的研究进展进行综述。     

土壤腐蚀网站硫酸盐还原菌的研究

从我国东北、西北、西南和华北的的10多种土壤的苗蚀试验站的钢件周围及腐蚀产物中,分离、纯化了13株硫酸盐还原菌(SRB),测定了它们的形态、生理生化特性及氢化酶活性,据以确定我国广大地区土壤中分布的钢铁晦蚀厌氧腐蚀萄主要为普通脱硫弧菌(Desulfovibriavulgaris)和脱硫脱硫弧菌(D.desulfuricans)。他们对钢的腐蚀速率和其氧化酶活性存在着很好的相关性。     

Kinetics of SO4−2 reduction under different growth media by sulfate reducing bacteria

Sulfate Reducing Bacteria (SRB) were used to reduce the SO₄ ^–2 concentration in waste water. The growth pattern of SRB was found by varying the concentration of nutrients and the biomass. The specific reaction constant was evaluated in each case.     

Hydrolysis of newspaper polysaccharides under sulfate reducing and methane producing conditions

The initial decomposition rates of cellulose and hemicellulose were measured using toluene to specifically inhibit the microbial uptake of hydrolysis products during the degradation of newspaper under sulfate reducing and methane producing conditions. The amount of glucose and xylose accumulation in the first 2 weeks of incubation period was higher in the sulfate reducing condition compared to the methane producing condition. It was estimated that 28 and 6% of initially loaded cellulose in the sulfate reducing condition and the methane producing condition was hydrolyzed, respectively. Accordingly, the newspaper-cellulose hydrolysis rate constant was estimated to be 6.7 times higher in sulfate reducing condition than in methane producing condition. Based on the glucose accumulation patterns, when sulfate reducing bacteria (SRB) were inhibited by anthraquinone and molybdate (Na₂MoO₄), it may be suggested that SRB might have contributed to the hydrolysis of cellulose, while their effect on the hydrolysis of hemicellulose could not be elucidated.     

Anaerobic degradation and dehalogenation of chlorosalicylates and salicylate under four reducing conditions

The anaerobic biodegradability and transformationof the mono-and dichlorinated salicylates(2-hydroxybenzoates) was examined under denitrifying,Fe (III) reducing, sulfate reducing andmethanogenic conditions. 3,6-Dichlorosalicylateand 6-chlorosalicylate are anaerobic microbialmetabolites of dicamba, a widely used herbicide.Anaerobic microcosms were established withdicamba treated soil from Wyoming, and golfcourse drainage stream sediments from NewJersey, which were each spiked with salicylate,3,6-dichlorosalicylate or one of the fourmonochlorosalicylate isomers. Salicylatewas degraded under denitrifying, sulfidogenic andmethanogenic conditions. In methanogenicenrichments 5-chlorosalicylate and 3-chlorosalicylatewere reductively dehalogenated to salicylatewhich was then utilized. Dehalogenation ofmonochlorinated salicylates to salicylate wasalso observed in denitrifying chlorosalicylatedegrading cultures. The study revealed thatthe position of the chlorine substituent as well as thepredominant electron accepting process affectthe rate and extent of chlorosalicylate degradationin anoxic environments.     

Effect of hydrogen sulfide on growth of sulfate reducing bacteria

A culture of sulfate reducing bacteria (SRB) growing on lactate and sulfate was incubated at different pH values in the range of 5.8-7.0. The effect of pH on growth rate was determined in this pH range; the highest growth rate was observed at pH 6.7. Hydrogen sulfide produced from sulfate reduction was found to have a direct and reversible toxicity effect on the SRB. A hydrogen sulfide Concentration of 547 mg/L (16.1 mM) completely inhibited the culture growth. Comparison between acetic acid and hydrogen sulfide inhibition is presented and the concomitant inhibition kinetics are mathematically described. (c) 1992 John Wiley & Sons, Inc.     

Anaerobic degradation of 3-halobenzoates by a denitrifying bacterium

A denitrifying bacterium was isolated from a river sediment after enrichment on 3-chlorobenzoate under anoxic, denitrifying conditions. The bacterium, designated strain 3CB-1, degraded 3-chlorobenzoate, 3-bromobenzoate, and 3-iodobenzoate with stoichiometric release of halide under conditions supporting anaerobic growth by denitrification. The 3-halobenzoates and 3-hydroxybenzoate were used as growth substrates with nitrate as the terminal electron acceptor. The doubling time when growing on 3-halobenzoates ranged from 18 to 25 h. On agar plates with 1 mM 3-chlorobenzoate as the sole carbon source and 30 mM nitrate as the electron acceptor, strain 3CB-1 formed small colonies (1–2 mm in diameter) in 2 to 3 weeks. Anaerobic degradation of both 3-chlorobenzoate and 3-hydroxybenzoate was dependent on nitrate as an electron acceptor and resulted in nitrate reduction corresponding to the stoichiometric values for complete oxidation of the substrate to CO₂. 3-Chlorobenzoate was not degraded in the presence of oxygen. 3-Bromobenzoate and 3-iodobenzoate were also degraded under denitrifying conditions with stoichiometric release of halide, but 3-fluorobenzoate was not utilized by the bacterium. Utilization of 3-chlorobenzoate was inducible, while synthesis of enzymes for 3-hydroxybenzoate degradation was constitutively low, but inducible. Degradation was specific to the position of the halogen substituent, and strain 3CB-1 did not utilize 2- or 4-chlorobenzoate. Received: 6 November 1998 / Accepted: 19 January 1999     

硫酸盐还原菌固态发酵产品酶活测定方法的建立

为建立生物质固载硫酸盐还原菌（SRB）产品酶活的检测方法,首先确定亚硫酸盐还原酶（SiR）作为SRB胞外酶表征其生物活性的可行性;其次对固态发酵产品制备粗酶液的各种条件进行探讨,研究浸泡介质、浸泡时间、超声功率、超声时间、预处理方式等单因素对酶活的影响;由正交实验确定粗酶液制备的最佳工况。实验结果表明,先浸提后超声处理的酶活值显著高于仅用超声、浸提的结果。为确保酶活值测定不受干扰,排除超声温度对结果的影响,确定实验最佳工况：2g固态发酵产品经磷酸盐缓冲液25mL、浸提60min,酶活值为1．0799U／g。生物质固载SRB酶活检测方法的建立为固态发酵产品活性评价、固态发酵条件优化、可渗透反应墙生化性能评估提供一定的理论依据。     

Anaerobic degradation of methoxylated aromatic compounds by Clostridium methoxybenzovorans and a nitrate-reducing bacterium Thauera sp. strain Cin3,4

The coupling of growth of the o-demethylating bacterium, Clostridium methoxybenzovorans SR3, with a nitrate-reducing bacterium able to degrade aromatic compounds, Thauera sp. Cin3,4, allowed complete mineralization of poorly oxidizable methoxylated aromatic compounds such as vanillate, isovanillate, vanilline, anisate, ferulate and veratrate. C. methoxybenzovorans o-demethylated these aromatic compounds to their corresponding hydroxylated derivatives and fermented the side chains to acetate and butyrate. The hydroxylated compounds and the fermentation end-products in the C. methoxybenzovorans spent growth medium were then completely metabolized to CO₂ on inoculation with the Thauera strain. Kinetic studies with veratrate indicated that C. methoxybenzovorans initially o-demethylated the substrate to vanillate and then further to protocatechuate together with the production of acetate and butyrate from the demethylated side chains. Protocatechuate, acetate and butyrate were then utilized as a carbon source by the Thauera strain aerobically or anaerobically in the presence of nitrate. The results therefore suggest that mono- or dimethoxylated aromatic compounds can be completely mineralized by coupling the growth of a fermentative bacterium with a nitrate-reducing bacterium, and a metabolic pathway for this is proposed.     

Anaerobic degradation of 3-hydroxybenzoate by a newly isolated nitrate-reducing bacterium

A Gram-negative nitrate-reducing bacterium, strain Asl-3, was isolated from activated sludge with nitrate and 3-hydroxybenzoate as sole source of carbon and energy. The new isolate was facultatively anaerobic, catalase- and oxidase-positive and polarly monotrichously flagellated. In addition to nitrate, nitrite, N2O, and O2 served as electron acceptors. Growth with 3-hydroxybenzoate and nitrate was biphasic: nitrate was completely reduced to nitrite before nitrite reduction to N2 started. Benzoate, 3-hydroxybenzoate, 4-hydroxybenzoate, protocatechuate or phenyl-acetate served as electron and carbon source under aerobic and anaerobic conditions. During growth with excess carbon source, poly-beta-hydroxybutyrate was formed. These characteristics allow the affiliation of strain Asl-3 with the family Pseudomonadaceae. Analogous to the pathway of 4-hydroxybenzoate degradation in other bacteria, the initial step in anaerobic 3-hydroxybenzoate degradation by this organism was activation to 3-hydroxy-benzoyl-CoA in an ATP-consuming reaction. Cell extracts of 3-hydroxybenzoate-grown cells exhibited 3-hydroxybenzoyl-CoA synthetase activity of 190 nmol min-1 mg protein-1 as well as benzoyl-CoA synthetase activity of 86 nmol min-1 mg protein-1. A reductive dehydroxylation of 3-hydroxybenzoyl-CoA could not be demonstrated due to rapid hydrolysis of chemically synthesized 3-hydroxybenzoyl-CoA by cell extracts.     

In vitro Activity of Gatifloxacin Against 238 Strains of Anaerobic Bacteria

The activity of gatifloxacin, a new 8-methoxy-fluoroquinolone, was tested against 208 pulmonary pathogens and against an additional 30 isolates of the Bacteroides fragilis group. Pulmonary isolates were from patients with documented anaerobic pleuropulmonary infections and were obtained by appropriate sampling methods. MICs were determined using the NCCLS-approved Wadsworth brucella laked blood agar method and compared to those of clindamycin, imipenem, metronidazole and trovafloxacin. Breakpoints used to define susceptible and [resistant] categories were (in μg/ml): Clindamycin-2, imipenem-4, metronidazole 8 and trovafloxacin. No breakpoint has been defined for gatifloxacin. Gatifloxacin inhibited 99% of all anaerobes tested at 4 μg/ml and 97% of all strains at 2 μg/ml. One strain of B. fragilis was resistant to gatifloxacin at 4 μg/ml; all strains of other B. fragilis group species were susceptible. One strain of Peptostreptococcus sp. was resistant to both gatifloxacin and trovafloxacin (MIC >4 μg/ml). All other strains were susceptible to all agents at ≤μg/ml. All of the non-sporeforming Gram-positive rods were susceptible to gatifloxacin at ≤μg/ml (three strains had an MIC of 4 μg/ml). Trovafloxacin had MICs of 4 μg/ml for two strains, and an MIC of 8 μg/ml for one strain. Five percent of B. fragilis, 21% of other B. fragilis group species and 20% of Clostridium species (other than C. difficile, C. perfringens or C. ramosum) were resistant to clindamycin. No imipenem resistant isolates were found in this study. Gatifloxacin appears to have excellentin vitro activity against pulmonary isolates of anaerobes and very good activity against strains of the B. fragilis group.