硫代硫酸盐氧化菌TX的分离、鉴定及其生物学特性

从聚硫橡胶废水处理系统中分离到一株硫代硫酸盐氧化细菌TX.根据其形态学特征、生理特征和16S rRNA基因序列相似性分析,将该菌株初步鉴定为盐生硫杆菌属(Halothiobacillus sp.)(GenBank登录号为EU871645).该菌株能利用硫代硫酸盐、单质硫、连四硫酸盐、硫化物或亚硫酸盐为唯一能源进行自养生长,不能利用葡萄糖、蔗糖、果糖、乳糖、麦芽糖或酵母粉进行异养生长,为专性化能无机自养型硫杆菌.在以硫代硫酸钠为唯一能源的培养基中其最适生长温度为30℃～35℃,最适起始pH值为3.0～5.0.在矿物盐培养基中,硫代硫酸盐最终被氧化成硫酸,造成培养基pH持续下降.在摇瓶分批培养和硫胶废水处理过程中均检测到连四硫酸盐的积累,表明该菌株主要通过连四硫酸盐途径或"S4I"途径进行硫代硫酸盐的生物氧化.     

一株嗜盐嗜碱硫氧化菌的筛选、鉴定及硫氧化特性

【背景】沼气和天然气等清洁能源中往往会含有一定量的硫化氢,硫化氢的存在不仅污染环境,而且对人类危害很大。【目的】以硫代硫酸钠为唯一硫源从巴丹吉林沙漠盐碱湖岸边沉积物中分离筛选得到一株硫氧化菌BDL05,并研究其硫氧化特性。【方法】通过形态观察、生理生化特征及16S rRNA基因序列分析对硫氧化菌BDL05进行鉴定。【结果】菌株BDL05为革兰氏阴性菌,弧状,其16S rRNA基因序列与Thiomicrospira microaerophila ASL 8-2的相似性达99.8%,将其命名为Thiomicrospira microaerophila BDL05。该菌氧化硫代硫酸盐的最适pH为9.3,最适总钠盐浓度为0.8mol/L,在以硫化钠为硫源的气升式反应器中单质硫的生成率为94.7%,生成速率为3.0 mmol/(L·h)。【结论】菌株Thiomicrospira microaerophila BDL05为嗜盐嗜碱硫氧化菌,其耐盐耐碱性较强,比生长速率快,硫化钠氧化能力较强,是一株在气体生物脱硫方面具有应用价值的菌株。     

一株丁草胺降解菌的分离鉴定及其降解特性的研究

利用富集培养技术从长期施用丁草胺的稻田土壤中分离得到能够降解丁草胺的细菌1株, 标记为LYC-1。经形态特征、生理生化特征和16S rRNA序列分析, 将该菌株鉴定为不动杆菌属(Acinetobacter sp.), 菌株LYC-1的最适生长温度为30°C, 最适pH值为7.5。当接种量为5%时, 该菌株在含100 mg/L 的丁草胺无机盐基础培养液中培养7 d后, 可使丁草胺降解达80%以上。     

一株氨氧化细菌的分离鉴定及其氨氧化特性

以城市污水处理工程的好氧活性污泥为菌源,通过富集、分离纯化,筛选出一株自养型氨氧化细菌CM-NRO14,进一步研究了该菌株的系统发育地位及氨氧化特性。根据菌株的形态学特征、生理生化特性及16S rDNA序列测定分析,确定该菌株属于亚硝酸单胞杆菌。CM-NRO14菌株的最大氨氧化速率24.54 mg/(L·d),利用氨的Km值45.66 mg/L,氨自身的抑制常数2401 mg/L,最大氨氧化速率的氨浓度331.2 mg/L。亚硝酸盐对氨氧化的抑制常数1524 mg/L。氨氧化的最适pH 7.79。氨氧化的最适温度34℃。菌株在初始氨氮浓度为180 mg/L,最佳氨氧化pH、温度条件下,4 d内氨氮降解率达到91.6%,菌株倍增时间2.94 d。     

一株甲烷氧化菌的分离鉴定与特性

甲烷氧化菌是一类能以甲烷作为唯一碳源和能源进行同化和异化代谢的微生物。从若尔盖高原不同地点采集的样品中筛选得到一株名为XN1的甲烷氧化菌,根据此菌株的形态与16SrRNA序列同源性分析,证实该菌株属于Methylomonas属。对该菌株的培养条件进行研究的结果表明,以甲烷与甲醇共同作为碳源,硝酸钾和氯化铵共同为氮源时菌生长最好,最适生长温度为25℃,最适生长pH为6.5,培养基中CuSO4·5H2O和FeSO4·7H2O的浓度以0.03mg/L和0.4mg/L为宜。     

嗜酸硫杆菌属硫氧化系统研究进展

硫化矿的酸溶解和化学氧化过程中(H 和Fe3 作用下,金属硫化矿中分解),伴随着硫元素转变成多聚硫S8或硫代硫酸盐的过程。对嗜酸硫杆菌属硫氧化过程的研究表明,胞外环状多聚硫S8可能通过细胞外膜蛋白巯基活化成线状-SnH后,被转运到细胞周质区域,进而被硫加双氧酶氧化成SO32-,活化过程中同时生成少量H2S;这些酶促反应不需要辅助因子参与,不释放电子。胞外硫代硫酸盐通过未知途径进入细胞周质。细胞周质中的SO32-主要经由亚硫酸-受体氧化还原酶氧化成SO42-,S2O32-可能经由硫代硫酸盐-辅酶Q氧化还原酶、硫代硫酸盐脱氢酶、连四硫酸盐水解酶等氧化为硫酸,少量H2S则经由硫化物-辅酶Q氧化还原酶氧化为多聚硫,后者再经由SO32-和S2O32-氧化生成最后产物SO42-。这些生物氧化过程释放的电子进入呼吸链参与产生细菌生长代谢所需的能量。然而,关于A.ferrooxidans硫氧化系统中各种硫化合物的酶催化氧化机制的研究仍很缺乏,胞内外硫化合物的转运机制、是否存在胞外酶催化氧化等仍然有待解决。另外,硫的型态和价态、酶催化反应的细胞微区域以及硫氧化系统中一些关键酶的分离及其表达基因的鉴定等问题都还有待进一步研究。基于对这些事实的分析,提出了一个嗜酸硫杆菌属硫氧化系统的模型。     

日本三宅岛火山土壤中硫代硫酸盐氧化菌的分离、生物学特性及鉴定

【目的】利用培养法从日本三宅岛火山土壤(堆积年限131年)中分离到一株能氧化分解硫代硫酸盐的细菌MU2A-22T。【方法】用培养法对该菌株MU2A-22T进行了生理生化性质以及分类学位置上的确定。【结果】菌株MU2A-22T为革兰氏阴性,短杆状或球状。理化性质表明该菌株能利用葡萄糖、L-阿拉伯糖、葡萄糖酸盐、己二酸酯、dL-苹果酸钠、硫代硫酸钠(最适浓度为2.5 mmol/L)为唯一碳源进行自养生长。最适生长温度为25°C 30°C,最适pH为6.0 8.0。菌株MU2A-22T的16S rRNA序列与菌株Paracoccus solventivorans 6637T亲缘关系最近,序列相似性为97%,编码核酮糖-1,5-二磷酸羧化酶/加氧酶(Rubisco)的基因也被确定。对Paracoccus属内几种近缘菌的脂肪酸分析,证明菌株MU2A-22T中含有Paracoccus属的特征氨基酸,其中含量大于10%的分别为C18:1(74.7%)和C18:0(12.1%)。DNA-DNA杂交实验表明,菌株MU2A-22T与Paracoccus solventivorans 6637TDNA的相似度为49.3%。MU2A-22T菌株G+C含量为66.5%66.7%。【结论】菌株MU2A-22T为Paracoccus属内的一新种菌(登录号GQ452286),命名为Paracoccus scorialis sp.nov.。     

象山港网箱养殖区沉积物的古菌空间分布

对象山港网箱养殖区及其周边沉积物中古菌群落的空间分布进行研究,应用基于16S rRNA基因的T-RFLP(末端限制性片段多态性分析)技术分析象山港网箱养殖区及其周边不同深度沉积物中古菌的群落结构和多样性,并构建克隆文库进行系统发育学分析。测定沉积物各项理化因子,通过PCA和RDA分析了古菌群落分布及其与环境因子之间的关系。结果表明,泉古菌是港口沉积物中的优势古菌群,占古菌群落的50%以上。网箱养殖区沉积物的古菌群落结构较非养殖区简单,多样性降低。非养殖区古菌群落随深度呈现有规律的变化。营养盐类和pH是造成养殖区域古菌群落结构区别于非养殖区域的主要环境因素。     

一株嗜酸硫氧化菌的分离

从江西德兴分离得到一株嗜酸硫氧化杆菌DX-2,采用双层固体培养基进行分离纯化,对分离菌株进行了形态、生理生化特性研究及16SrRNA序列分析.该菌株为革兰氏阴性细菌,短杆状,菌体大小（0.4～0.5）μm×（1～2）μm,化能自养,可利用硫磺和硫代硫酸盐为能源生长,不能利用亚铁进行生长.以16SrRNA序列同源性为基础构建了相关种属在内的系统发育树,结果表明,DX-2与喜温硫杆菌（Acidithiobacillus caldus）处于同一进化树分支中,相似性达99%以上.考察了不同重金属离子对DX-2的生长的影响.     

锰氧化细菌的分离鉴定及其锰氧化特性的分析

利用选择性培养基对锰矿样品进行分离、筛选, 得到一株高效锰氧化细菌(MN1405)。经形态特征、生理生化特征以及16S rRNA 基因序列分析, 将菌株MN1405 鉴定为Arthrobacter echigonensis。在培养条件下, MN1405 对培养基中的锰离子去除率可达93.38%, 且其培养所获得的培养液也具有良好的除锰效果。     

Citreicella thiooxidans gen. nov., sp. nov., a novel lithoheterotrophic sulfur-oxidizing bacterium from the Black Sea

Four strains of obligately heterotrophic bacteria isolated from the oxygen-sulfide interface of the Black Sea are characterized. The bacteria are aerobic, Gram-negative, with lemon-like, nonmotile cells. Bacteriochlorophyll a is not detected. They are mesophilic and neutrophilic with a temperature range of 8–35 °C (optimum 25) and pH range of 6.5–8.5 (optimum 7.8). Their growth is NaCl dependent within a range of 5 and 60 (optimum 20) g l⁻¹. They are able to oxidize thiosulfate, sulfide and elemental sulfur to sulfate and to use metabolic energy from these reactions (lithoheterotrophy). According to the level of DNA reassociation of more than 40%, all isolates represent a single generic group. The G+C content of the DNA was in the range of 67.5–69.2 mol%. According to phylogenetic analysis, the new isolates form a separate branch in the alpha-3 subdivision of the Proteobacteria together with two undescribed marine bacterial strains. On the basis of phenotypical and genomic properties, the new isolates are described as a new genus and species Citreicella thiooxidans gen. nov., sp. nov. The type strain is CHLG 1^T (=DSM 10146, UNIQEM U 228).     

一株养殖水体中亚硝酸盐去除菌的鉴定及其去除条件

【目的】从养殖污泥中分离筛选优良亚硝酸盐去除菌,并对其去除条件进行研究。【方法】从养殖污泥中分离亚硝酸盐去除菌,进一步通过测定比较分离菌株对亚硝酸盐的去除率,筛选优良的亚硝酸盐去除菌,通过API ID32GN细菌鉴定系统以及16S rDNA序列分析法对其进行鉴定,并采用单因子法研究其去除亚硝酸盐的条件。【结果】从养殖污泥中分离筛选了一株优良的亚硝酸盐去除菌AQ-3,其对50 mg/L亚硝酸盐的去除率高达99.47%。菌株AQ-3被鉴定为鲍曼氏不动杆菌(Acinetobacter baumannii)(GenBank登录号:JF751054.1),其16S rDNA序列与基因库中不动杆菌属菌株的16S rDNA序列有99%?100%的同源性,而且与鲍曼氏不动杆菌KF714株(GenBank登录号:AB109775)的亲缘关系最近。菌株AQ-3去除亚硝酸盐的最适初始pH范围为7?9,最佳碳源为乙酸钠和丁二酸钠,而且随着初始菌浓度的不断增大,菌株AQ-3对亚硝酸盐的去除率显著升高;随着亚硝酸盐浓度的不断增大,菌株AQ-3对亚硝酸盐的去除率逐渐降低。【结论】在丰富亚硝酸盐去除菌种质资源的同时,为该菌在养殖水体中的实际应用提供了理论基础。     

一株产右旋糖苷酶海洋细菌的筛选鉴定

【目的】筛选鉴定产右旋糖苷酶的海洋细菌,并对其所产右旋糖苷酶的酶学性质及在变异链球菌牙菌斑生物膜中的应用进行初步研究。【方法】利用平板透明圈法从海洋环境中筛选产右旋糖苷酶的细菌,根据菌株形态特征、生理特征及16S rDNA序列确定其分类学地位,采用体外生物膜模型研究该酶对变异链球菌牙菌斑生物膜形成的抑制作用。【结果】从海泥中筛选出一株产右旋糖苷酶的细菌KQ11,初步鉴定为节杆菌(Arthrobacter sp.)。该菌株的最适生长温度为30°C,最适生长pH 7.5,最适生长NaCl浓度为0.4%。右旋糖苷酶的最适作用温度为45°C,最适作用pH为5.5。该酶能有效地抑制变异链球菌牙菌斑生物膜的形成。【结论】菌株KQ11右旋糖苷酶能够抑制变异链球菌牙菌斑生物膜的形成,可望用于漱口液等口腔护理产品中。     

Purification and characterization of a novel alginate lyase from the marine bacterium Cobetia sp. NAP1 isolated from brown algae

The application of marine resources, instead of fossil fuels, for biomass production is important for building a sustainable society. Seaweed is valuable as a source of marine biomass for producing biofuels such as ethanol, and can be used in various fields. Alginate is an anionic polysaccharide that forms the main component of brown algae. Various alginate lyases (e.g. exo- and endo-types and oligoalginate lyase) are generally used to degrade alginate. We herein describe a novel alginate lyase, AlgC-PL7, which belongs to the polysaccharide lyase 7 family. AlgC-PL7 was isolated from the halophilic Gram-negative bacterium Cobetia sp. NAP1 collected from the brown algae Padina arborescens Holmes. The optimal temperature and pH for AlgC-PL7 activity were 45 °C and 8, respectively. Additionally, AlgC-PL7 was thermostable and salt-tolerant, exhibited broad substrate specificity, and degraded alginate into monosaccharides. Therefore, AlgC-PL7 is a promising enzyme for the production of biofuels.     

Isolation and characterization of chitinase from a marine bacterium Vibrio sp.

A chitinase was separated from the culture broth of Vibrio sp. 11211 isolated from sediment from the South China Sea. The chitinase was purified 18.3-fold with 33% recovery by ammonium sulphate precipitation and chromatography. The subunit molecular weight of the enzyme was estimated by SDS-PAGE to be about 30kDa. The enzyme showed optimum pH at 6.5 and optimum temperature at 50^°C, and was stable in the pH range of 4 to 9 and at the temperature below 40^°C.     

一株氯霉素降解细菌的分离鉴定与代谢特性研究

微生物是介导环境中氯霉素降解转化的主要驱动者,但高效降解矿化菌株资源匮乏,氧化反应介导的代谢途径不清。为研究微生物介导下氯霉素的环境归趋过程,为氯霉素污染环境强化修复提供菌株资源,文中以受氯霉素污染的活性污泥为接种源,首先富集获得一个由红球菌Rhodococcus主导 (相对丰度>70%) 的氯霉素高效降解菌群,并从中分离获得一株能够高效降解氯霉素的菌株CAP-2,通过16S rRNA基因分析鉴定为红球菌Rhodococcus sp.。菌株CAP-2能在不同营养条件下高效降解氯霉素。基于菌株CAP-2对检测到的代谢产物对硝基苯甲酸和已报道的代谢产物对硝基苯甲醛和原儿茶酸的生物转化特征,提出其降解途径是由氯霉素侧链氧化断裂生成对硝基苯甲醛,进一步氧化为对硝基苯甲酸的新型氧化降解途径。该菌株对于氯霉素分解代谢的分子机制研究以及受氯霉素污染环境的原位生物修复应用具有巨大潜力。     

海洋细菌SSQ500的鉴定及其发酵产物抗肿瘤作用机制研究

[目的]对具有抗肿瘤活性的海洋细菌SSQ500进行分类鉴定,初步探讨其发酵产物的抗肿瘤作用机制.[方法]通过菌株的形态学观察、生理生化特征检测和16S rDNA序列分析及系统发育分析,以确定菌株的分类.不同浓度的活性物质作用于肿瘤细胞不同时间后收集细胞,流式细胞术分析人肝癌细胞(SMMC-7721)凋亡比例,检测Caspase3、Caspase8蛋白的表达.[结果]初步鉴定菌株SSQ500属于粘球菌属(Myxococcaceae sp.).该菌株发酵产物能诱导SMMC-7721凋亡,上调Caspase3、Caspase8蛋白的表达.[结论]SSQ500属于粘球菌属,其发酵产物的抗肿瘤作用机制为诱导肿瘤细胞凋亡,可能与上调Caspase3、Caspase8蛋白的表达有关.     

Effects of inhibitors and NaCl on the oxidation of reduced inorganic sulfur compounds by a marine acidophilic, sulfur-oxidizing bacterium, Acidithiobacillus thiooxidans strain SH

The effect of NaCl and the pathways of the oxidation of reduced inorganic sulfur compounds were studied using resting cells and cell-free extracts of Acidithiobacillus thiooxidans strain SH. This isolate specifically requires NaCl for growth. The oxidation of sulfur and sulfite by resting cells was strongly inhibited by 2-heptyl-4-hydroxyquinoline-N-oxide. Carbonylcyanide m-chlorophenyl-hydrazone and monensin were also relatively strong inhibitors. Thiosulfate-oxidizing activity was not inhibited by these uncouplers. Valinomycin did not inhibit the oxidation of sulfur compounds. NaCl stimulated the sulfur- and sulfite-oxidizing activities in resting cells but not in cell-free extracts. The tetrathionate-oxidizing activity in resting cells was slightly stimulated by NaCl, whereas it did not influence the thiosulfate-oxidizing activity. Sulfide oxidation was biphasic, suggesting the formation of intermediate sulfur. The initial phase of sulfide oxidation was not affected by NaCl, whereas the subsequent oxidation of sulfur in the second phase was Na⁺-dependent. A model is proposed for the role of NaCl in the metabolism of reduced sulfur compounds in A. thiooxidans strain SH.     

一株荧光假单胞杆菌的分离鉴定与反硝化特性

【目的】从污水厂的活性污泥中获得一株高效反硝化细菌。【方法】采用低温驯化,进行初筛、复筛选取一株反硝化活性最高的菌株,命名为L2,通过形态学、生理生化特征及16S r RNA基因序列分析研究其分类地位,系统研究理化因素对该菌株反硝化性能的影响。【结果】菌株在低温条件下能够稳定高效地进行反硝化,鉴定该菌株为荧光假单胞杆菌(Pseudomonas fluorescens),其反硝化最适接种量为10%,温度为20°C,p H为7.0,盐浓度为0.5%,碳源为葡萄糖,C/N为5.0,能够耐受较高初始硝态氮浓度。【结论】菌株L2是一株耐低温、耐高浓度初始硝态氮、耐低C/N、兼性厌氧、高效反硝化的荧光假单胞杆菌。     

Purification and characterization of thermostable pectate-lyases from a newly isolated thermophilic bacterium, Thermoanaerobacter italicus sp. nov.

A novel thermophilic spore-forming anaerobic microorganism (strain Ab9) able to grow on citrus pectin and polygalacturonic acid (pectate) was isolated from a thermal spa in Italy. The newly isolated strain grows optimally at 70°C with a growth rate of 0.23 h⁻¹ with pectin and 0.12 h⁻¹ with pectate as substrates. Xylan, starch, and glycogen are also utilized as carbon sources and thermoactive xylanolytic (highest activity at 70°–75°C), amylolytic as well as pullulolytic enzymes (highest activity at 80°–85°C) are formed. Two thermoactive pectate lyases were isolated from the supernatant of a 300-l culture of isolate Ab9 after growth on citrus pectin. The two enzymes (lyases a and b) were purified to homogeneity by ammonium sulfate treatment, anion exchange chromatography, hydrophobic chromatography and finally by preparative gel electrophoresis. After sodium dodecylsulfate (SDS) gel electrophoresis, lyase a appeared as a single polypeptide with a molecular mass of 135 000 Da whereas lyase b consisted of two subunits with molecular masses of 93 000 Da and 158 000 Da. Both enzymes displayed similar catalytic properties with optimal activity at pH 9.0 and 80°C. The enzymes were very stable at 70°C and at 80°C with a half-life of more than 60 min. The maximal activity of the purified lyases was observed with orange pectate (100%) and pectate-sodium salt (90%), whereas pectin was attacked to a much lesser extent (50%). The K _m values of both lyases for pectate and citrus pectin were 0.5 g·l⁻¹ and 5.0 g·l⁻¹, respectively. After incubation with polygalacturonic acid, mono-, di-, and tri-galacturonate were detected as final products. A 2.5-fold increase of activity was obtained when pectate lyases were incubated in the presence of 1 mM Ca²⁺. The addition of 1 mM ethylenediaminetetraacetic acid (EDTA) resulted in complete inhibition of the enzymes. These heat-stable enzymes represent the first pectate-lyases isolated and characterized from a thermophilic anaerobic bacterium. On the basis of the results of the 16S rRNA sequence comparisons and the observed phenotypic differences, we propose strain Ab9 as a new species of Thermoanaerobacter, namely Thermoanaerobacter italicus sp. nov. Received: May 25, 1997 / Accepted: June 5, 1997