一株菲降解菌的特性及相关降解基因的克隆

从石油污染土壤中分离到一株菲降解菌2F5-2.根据该菌株生理生化特征和16S rDNA序列相似性分析,将其初步鉴定为鞘氨醇杆菌属(Sphingobium sp.).该菌株在10 h内对100 mg/L的菲的降解率为100%.降解菲的最适温度为30℃,最适pH为7.对降解途径的初步研究显示,该菌株通过水杨酸途径降解菲.克隆了编码芳香烃双加氧酶α亚基的基因phdA,它与菌株Sphingomonas sp.P2、Sphingobium yanoikuyae B1、Sphingomonas sp.ZP1中phdA的同源性分别为97.9%、98%和100%,表明该基因具有保守性.图6参16     

海洋石油降解菌的筛选与降解特性

用采自山东胜利油田的石油污染水体,以原油为唯一碳源,经人工海水培养基富集培养,得到lO株细菌.通过降解率的测定,发现其中一株细菌HB-1具有较强的石油降解能力,200 r/min振荡培养6 d,其原油的降解率达54.74%.根据菌落特征、菌体形态、生理生化特性和16S rDNA序列分析,确定HB-1为不动杆菌属(Acinetobacter sp.).进一步实验表明,该菌株最适宜牛长和降解石油的条件为:温度25～32℃,初始pH值6.5～7.5,盐度3%,此条件符合实际海洋环境的要求.图6表2参22     

孔雀石绿降解菌M3的分离鉴定及降解特性研究

从鱼塘底泥中筛选分离出1株能高效降解低含量孔雀石绿(MG)的细菌M3.经16S rDNA同源性序列分析,鉴定为泛菌属(Pantoea sp.).30 ℃静止培养条件下,该菌株对0.5、1.0、2.0和5.0 mg·L-1孔雀石绿5 d的降解率分别为97.54%、97.1%、100%和77.8%.菌株M3不能以MG为唯一碳源进行生长和代谢.葡萄糖、NH4NO3、KH2PO4/K2HPO4均能影响菌株M3对MG的降解.20～30 ℃温度范围内菌株M3对MG有明显降解效果,且降解速率随温度上升而提高.     

耐热石油烃降解混合菌的筛选及其群落结构分析

对克拉玛依采集的部分石油污染土壤进行了筛选,得到了5组石油烃高效降解混合菌,其中混合菌KL9-1在45℃的条件下,通过7 d的降解,稀油的降解率达到43.27%,稠油的降解率达到20.09%。混合菌KL9-1经过多次分离纯化后,获得3株具有石油烃降解能力的优势单菌,3株单菌对稀油的降解率都在30%以上。结合分离单菌株的形态、生理生化特征和16S rDNA基因序列的分析结果,初步鉴定KL9-1-1为Pseudomonas putida,KL9-1-2和KL9-1-3为Pseudomonas sp.。     

Porphyrobacter sp.D22F的降解特性及其在石油降解菌群中的生态位

为揭示多环芳烃(Polycyclic aromatic hydrocarbons,PAHs)降解微生物资源的多样性和石油降解菌群的优势菌,研究了从南海沉积物中分离得到的一株PAHs降解菌D22F的降解特性及其在石油降解菌群D22-1中的生态位.对菌株D22F进行16S rRNA基因同源性分析及透射电镜观察以初步确定其种属,通过培养法、气相色谱质谱联用(GC-MS)测定其多环芳烃降解范围和降解率,通过简并引物PCR扩增其PAHs双加氧酶大亚基基因片段并进行系统发育分析,采用变性梯度凝胶电泳(DGGE)监测石油降解菌群中的优势菌.结果表明,与菌株D22F的16S rRNA基因相似度最高的模式株为产卟啉杆菌属Porphyrobacter tepidarius DSM 10594T(AF465839;98.55%).该菌株能降解萘、甲基萘、苊、硫芴、菲、蒽等;对初始浓度为0.2 g/L菲10 d后的降解率可达90%以上.从其基因组DNA中克隆到的PAHs起始双加氧酶大亚基基因phnAc与Novosphingobium aromaticivorans DSM 12444中的质粒pNL1(CP000676)上的bphA1f基因相似度最高,达到99.41%.DGGE谱图显示,菌株D22F是石油降解菌群中的3种优势菌之一,在传代菌群中可稳定存在.Porphyrobacter sp.D22F为产卟啉杆菌属(Porphyrobacter)中首株以低分子量PAHs为唯一碳源和能源的菌株,是石油降解菌群的优势菌.     

两株吡啶降解菌的分离与鉴定

为了研究石油污染土壤中含氮杂环化合物的降解情况,该研究选择吡啶作为目标污染物,采用选择性富集培养的方法,从45份石油污染土壤样品中,分离得到260株降解吡啶污染物的高效降解菌株,选择降解效率最高的2株吡啶降解菌命名为菌株4-11和2-13,进行种属鉴定、细菌的生长情况和吡啶降解性能的考察.实验证明,60 h菌株4-11和2-13对质量浓度为1000 mg·L-1吡啶的降解率分别达到65.5%和64.1%.通过形态学、生理生化鉴定和16S rDNA序列比对分析,确定菌株4-11属于产硫酸杆菌(Thiobacillus)与Thiobacillus intermediu 同源性最高,为99.8 %,菌株2-13属屈挠杆菌(Flexibacter)与 Flexibacter giganteus同源性最高,为99.9 %.     

一株菲降解菌的分离鉴定及其降解条件研究

以菲为研究对象,从克拉玛依稠油污染土壤中筛选到1株对菲具有较好降解效果的菌株JZ3-21。通过形态观察、生理生化指标及16S rDNA序列分析对该菌株进行了鉴定。该株菌的16S rDNA序列与Pseudomonas属的相似性达99%,结合分离菌株的形态、生理生化特征和16S rDNA基因序列的分析结果,初步鉴定该菌株为恶臭假单胞菌（Pseudomonas putida.）。对其降解条件进行了研究,结果表明：在40℃,pH 8.0,接种量为1.5%的条件下,菌株对初始质量浓度为100 mg.L-1的菲在64 h内的降解率高达94.2%。该菌对高质量浓度菲有较好的耐受性,其最高耐受质量浓度可达2 000 mg.L-1。     

产生物表面活性剂的石油降解菌Acinetobacter BHSN的研究

从石油污染土壤中分离筛选获得1株产生物表面活性剂的石油降解菌株BHSN.经形态观察、生理生化试验、16S rDNA 序列分析等将其鉴定为不动杆菌属(Acinetobacter sp.).研究表明,BHSN菌株能降解石油中C13～C32正构烷烃.BHSN菌株生长和产生物表面活性剂的最适温度为25～30 ℃,最适pH为5.0～8.5.LB培养基中添加终质量浓度为10 g·L-1葡萄糖能促进菌体的生长,但不利于表面活性剂的合成;添加蔗糖、乳糖、麦芽糖和木糖对菌体的生长和表面活性剂的合成均有促进作用,其中麦芽糖的促进作用最强.初步研究表明,BHSN菌株产的生物表面活性剂为脂肽类;BHSN发酵液的表面张力由初始68.3 mN·m-1降低到28.6 mN·m-1.     

海南红树林沉积物中多环芳烃降解菌群组成及降解率的比较研究

已有研究证明能从有PAHs污染的区域筛选相应的降解菌,海南红树林存在PAHs污染,但少有研究从海南红树林沉积物中筛选PAHs降解菌。文章的研究目的是从海南不同红树林分布区的沉积物中筛选PAHs的降解菌群比较PAHs的降解菌群降解特征,并阐明菌群降解率和菌群结构之间的关系,为环境中PAHs污染区域修复提供理论依据。通过采集海南不同红树林中24个样点的沉积物,以菲、芘、苯并(a)芘3种PAHs为混合碳源,富集培养降解菌群,用HPLC测定菌群降解率,以传统分离培养的方法比较菌群组成差异。所有样点筛得的群对3种底物的平均降解效率为菲(3-环)芘(4-环)苯并(a)芘(5-环)。由贪噬菌属和剑菌属组成的菌群Q15,对菲的降解率达到95.3%,由伯克氏菌属、鞘脂单胞菌属组成的菌群Q12对芘降解率为94%,仅有申氏菌属的菌群Q9,对苯并(a)芘降解率为49.7%。所有样点共筛得23个属60个种的降解菌,除两株为厚壁菌门外,其他均为变形菌门。其中变形菌门中菌DAC9的16S rRNA序列与最相近的菌株序列一致性为97%,可能是潜在新菌。结果表明,红树林沉积物中富集培养的PAHs降解菌群对3种底物的降解效率存在差异,其中中低环PAHs更易被降解。不同的菌属组合对PAHs的降解能力不同。并从沉积物中筛选出了一株潜在新菌。     

一株同时降解3种邻苯二甲酸酯降解菌的筛选鉴定及降解特性

选择邻苯二甲酸二甲酯(DMP)、邻苯二甲酸二丁酯(DnBP)和邻苯二甲酸二(2-乙基己基)酯(DEHP)作为目标污染物,采用富集驯化法从设施菜地土壤中筛选出1株可同时降解DMP、DnBP和DEHP的细菌MB1.经形态、生理生化特征及16S rDNA序列分析,初步鉴定为微杆菌属(Microbacterium sp.).通过正交试验研究了该菌株的最优降解条件以及最优条件下该菌株的生长曲线和降解曲线,最后在培养条件下研究了该菌株对人工污染土壤中邻苯二甲酸酯的降解特性.结果表明,菌株MB1的最优降解条件为:pH值为8,温度为25℃,接菌量为5%,每种邻苯二甲酸酯浓度为300 mg·L~(-1).在此最优条件下该菌株呈S型曲线增长,7 d后无机盐培养液中DMP、DnBP和DEHP的降解率分别为99.62%%、99.65%和55.26%.人工污染土壤中空白试验和投加菌株试验结果为:在不添加菌液的处理中,灭菌土壤21 d时DMP、DnBP和DEHP的降解率分别为3.86%、4.19%和2.01%;未灭菌土壤21 d时对DMP、DnBP和DEHP的降解率分别为4.82%、5.99%和3.44%.在添加菌液的处理中,21 d时土壤灭菌处理中DMP、DnBP和DEHP的降解率分别达94.45%、95.65%和39.21%;而土壤未灭菌处理中DMP、DnBP和DEHP的降解率分别达94.93%、95.99%和41.16%.该结果表明:土壤中土著微生物仅能降解微量PAEs,菌株MB1对土壤中DMP、DnBP和DEHP等3种PAEs污染物具有较为高效的降解能力,未灭菌土壤中邻苯二甲酸酯的降解效果略高于灭菌土壤.     

我国蚕豆根瘤菌的多样性和系统发育研究

采用数值分类、16S rDNA PCR-RFLP、IGS PCR-RFLP等方法对分离自我国11个省的50株蚕豆根瘤菌及11株参比菌株进行了表型测定和遗传型研究,同时对5株蚕豆根瘤菌的代表菌株进行了16S rDNA全序列测定.表型测定的结果表明,在80%的相似水平上供试菌株分为4个群,各群间存在地区交叉;16S rDNA PCR-RFLP的聚类结果与数值分类的聚类结果有很好的一致性;IGSRFLP反映的多样性更明显,形成的遗传群较多,可用于菌株间的鉴别.实验结果表明我国蚕豆根瘤菌具有极大的表型多样性和遗传多样性.系统发育研究结果表明,蚕豆根瘤菌的代表菌株均位于快生根瘤菌属(Rhizobium)系统发育分支,与R.leguminosarum USDA2370的全序列相似性达99.9%,说明蚕豆根瘤菌属于Rhizobium,系豌豆根瘤菌的一个生物型.图4表3参12     

Characterization of two metal resistant Bacillus strains isolated from slag disposal site at Burnpur, India

Two strains of Bacillus sp. resistant to arsenate and lead designated as AsSP9 and PbSP6, respectively were isolated from the slag disposal site. They were identified to be related to Bacillus cereus cluster on the basis of 16S rDNA based sequence analysis and phenotypic characteristics. Both were rod-shaped (AsSP9, 2-5 microm and PbSP6, 2-4 microm), aerobic, salt tolerant (2-8% NaCI), endospore forming bacteria with minor differences like the AsSP9 showed sporangial bulging and PbSP6 had positive lipase activity. The temperature range for their growth was 20-40 degrees C and pH range 6.0-9.0 with an optimum temperature of 37 degrees C and pH of 7 for both strains. The principal nitrogen sources forAsSP9 and PbSP6 were DL-Tryptophan and L-Phenylalanine, respectively. The suitable carbon source forAsSP9 was lactose and for PbSP6 sucrose. The heavy metal accumulation efficiency was found to be 0.0047 mg g(-1) of dry mass forAsSP9 and 0.686 mg g(-1) of dry mass for PbSP6.     

Identification of two alkane hydroxylases involved in long-chain n-alkane degradation by Dietzia sp. DQ12-45-1b北大核心CSCD

The aim of this study was to identify genes involved in long-chain alkane degradation in Dietzia sp. DQ12-45-1b. Functional genes were annotated by genome analysis. Induction of alkane hydroxylase genes by C28 n-alkane was analyzed by using quantitative real-time PCR in wild-type Dietzia sp. DQ12-45-1b and its alkW1 gene knockout mutant strain M 5-5. From the genome of Dietzia sp. DQ12-45-1b, two homologues, G1 and G2 genes were annotated, which showed 50% amino acid sequence similarity with AlmA from Acinetobacter sp. DSM17874, and 48% amino acid sequence similarity with LadA from Geobacillus thermodenitrificans NG80-2, respectively. In addition, G1 showed 71% amino acid sequence similarity with G1a, and G2 showed 34% and 87% amino acid sequence similarities with G2a and G2ß, respectively, which were annotated from Dietzia sp. E1 genome. In addition, the alkW1 gene knockout strain M 5-5 could grow with C28 n-alkane as the sole carbon source, indicating the presence of potential long-chain alkane hydroxylase gene(s) other than alkW1 in Diezia sp. DQ12-45-1b. Accordingly, induction of G1 and G2 genes was observed when Dietzia ap. DQ12-45-1b and alkW1 knockout mutant strain M 5-5 grew with C28 n-alkane as sole carbon source. The results indicated that G1 and G2 genes are mostly responsible for the degradation of long-chain alkanes in Dietzia sp. DQ12-45-1b, which has unique multiple alkane hydroxylase systems.     

硝基芳香烃降解菌株的分离、鉴定和应用

从被二硝基甲苯(DNT)和三硝基甲苯(TNT)污染的土壤中分离到5个高效降解硝基芳香烃类化合物的菌株TD1、TD2、TD3、TD4和TD5。16S rDNA序列分析表明,TD1、TD2、TD3和TD4属于节杆菌属(Arthrobacter),TD5属于不动杆菌属(Acinetobacter)。菌株复配试验表明,TD1、TD2、TD3、TD4和TD5的体积比例为2:2:2:1:1时对含有DNT和TNT的炸药废水的CODC r去除率最好。最佳复配的菌液投加到装有合适填料载体的生物反应器中用于处理炸药废水,运行47d后出水CODC r不超过200 mg/L,硝基苯类化合物的浓度在2 mg/L以下。     

慢生根瘤菌交叉结瘤及其系统发育关系的研究

选用包括6株花往根瘤菌[Bradyrhizobium sp.(Arachis)]在内的13株慢生根瘤菌，进行了6种豆科植物结瘤试验及共系统发育分析。研究结果表明，慢生根瘤菌与试验的豆科宿主植物间普遍存在交叉结瘤，还发现有些菌株在原宿主外的豆科植物根部形成类根瘤的现象。16S rDNA和23S rDNA的PCR－RFLP分析揭示出这些菌株rDNA基因的差异性，不同宿主来源的慢生根瘤菌株可以同属一个rDNA类型，而从同一宿主中分离的根瘤菌菌株可分属多种不同的rDNA类型，研究结果表明，采用rDNA PCR-RFLP等遗传背景分析的手段，可能筛选出广谱的根瘤菌力株，表4参8。     

Removal of nitrogen and phosphorus by eight strains of microalgae and their growth characteristics in Penaeus vannamei sewage北大核心CSCD

Microalgae are the most important primary productive forces in shrimp aquaculture systems. Microalgae not only provide oxygen and natural food for aquaculture objects, but they also absorb nitrogen (N) and phosphorus (P) to reduce water eutrophication. However, there are great differences in N and P absorption among different strains of microalgae. To maintain the sustainable development of shrimp aquaculture, the growth performances of eight microalgal strains in Penaeus vannamei sewage and N and P removal rates were investigated under laboratory conditions. The results indicated that the eight microalgal strains could reduce the N and P content in P. vannamei sewage to some extent. Microcystis aeruginosa, Chlamydomonas sp., and Chlorella pyrenoidosa grew very well, with average growth rates of 0.309 3, 0.246 9, and 0.215 5, respectively. There were significant differences in the removal efficiency among the different strains. The removal rates of total N by M. aeruginosa, Chlamydomonas sp., and C. pyrenoidosa were 74%, 69%, and 60%, respectively, at the end of the experiment, which were higher than the other species. M. aeruginosa and Chlamydomonas sp. had better total P removal efficiency than those of the other microalgal strains and removal rates were greater than 60%, and the second highest total P removal efficiency was by C. pyrenoidosa. Different types of microalgal strains had different absorption rates of different morphological nitrogen. M. aeruginosa and Chlamydomonas sp. had the highest nitrate nitrogen removal rate (approximately 70%). Chlamydomonas sp. had a fast and persistent removal rate of ammonia nitrogen, with the removal rate being as high as 100%. The removal efficiency of M. aeruginosa and C. pyrenoidosa were a little slower, and those of Scenedesmus obliquus, Synedra sp., and Navicula graciloides were the slowest. After 16 d, the removal rate reached more than 90%. Cryptomonas obovate and C. pyrenoidosa displayed the best removal rate of nitrite nitrogen, and the removal rate reached 80% on day 8, and the removal rate of C. obovata was more persistent. These results can provide scientific reference for the orientation and use of microalgae to remove pollutants in tailings water from shrimp aquaculture systems. © 2018 Science Press. All rights reserved.     

不对称还原α-氨基苯乙酮菌种的筛选及转化条件优化

芳香基手性胺醇是许多手性药物合成的重要手性砌块,生物催化不对称还原前手性酮是合成该类醇的重要方法之一.以α-氨基苯乙酮盐酸盐为模型底物从土壤中筛选获得两株能分别高立体选择性催化底物产生R型、S型相应醇的菌株,对映体过量值(e.e.)分别为99%和77%,编号为1403和4802,鉴定菌株所属为镰刀菌属和地霉属.对两株菌培养时期和转化条件的研究表明镰刀菌1403最适生长时间为24 h,最优菌体浓度20 g/L,最优底物浓度5 g/L;地霉4802最适生长时间24 h,最优菌体浓度80 g/L,最优底物浓度3 g/L.底物特异性研究表明,菌株1403和4802均可转化α-氯代苯乙酮、α-溴代苯乙酮、α-羟基苯乙酮和苯乙酮为相应醇,且以α-羟基苯乙酮为底物时,其产物均为S型,e.e.值达99%.     

原废水培养基分离活性污泥中的苯酚降解细菌

用未经处理的含酚焦化工业废水为基础配制培养基，采用平板涂布法，在10^-5稀释度下得到100个单菌落分离物．它们均能在以苯酚(500-800mg L^-1)为唯一碳源的无机盐培养基上生长，其16S rDNA基因的ARDBA多态性分析将这些分离物划分成4个分类操作单元(OTUs)．各单元代表株的16S rRNA基因序列分析和检索结果表明，97个分离物与Alcaligenes faecalis同源性达99％，2个分离物分别与Arthrobacter nicotianae和Klebsiella sp.99％同源．另一个分离物与Ochrobactrum sp．96％同源．苯酚经化酶大亚基基因(LmPHs)PCR扩增表明，除类似Arthrobacter nicotianae的菌株外，其它3个分类操作单元的降酚菌都能利用多组分苯酚经化酶代谢途径进行酚代谢．图2表2参11