苯酚降解菌phen8的分离筛选及其16SrDNA序列分析

为筛选高效苯酚降解菌株 ,从炼油厂排污废水中分离筛选到 1株苯酚降解菌 phen8。利用PCR方法和琼脂糖凝胶电泳技术检测到 phen8菌中苯酚羟化酶基因片段的特异性条带 ,从基因水平上证实了 phen8菌的苯酚降解功能的遗传基础。应用PCR技术克隆到 16SrDNA片段 ,其核苷酸序列分析结果表明 ,该菌株的 16SrDNA全序列与斯氏假单胞菌DSM 5 0 2 2 7和DSM 5 0 2 38的同源性为 98% (在GenBank中的登记号为AF 2 8476 4)。初步确立了该菌在微生物系统发育学上的地位 ,暂定为假单胞菌 (Pseudomonassp .) phen8。     

大西洋洋中脊深海多环芳烃降解菌群的优势菌分析

摘要：【目的】为了分析大西洋洋中脊深海海水及表层沉积物中多环芳烃(PAHs)降解菌群中的优势菌。【方法】采用富集培养法和平板涂布法从深海样品中分离可培养细菌及PAHs降解菌。通过16S rRNA基因测序完成系统发育分析。采用变性梯度凝胶电泳（DGGE）及DNA测序分析降解菌群中的优势菌。【结果】总共分离到16株细菌,包括一株PAHs降解菌Novosphingobium sp. 4D。系统发育分析发现,可培养细菌中两个最大的类群分别与Alcanivorax dieselolei NO1A（5/16）和Tistrella mobilis TISTR 1108T（5/16）亲缘关系最近。DGGE结果表明,在菌群MC2D中菌株4L（以及4M、4N, Alcanivorax dieselolei NO1A, 99.21％）、4D（Novosphingobium pentaromativorans US6-1T,97.07％）和4B（以及4E、4H、4K,Tistrella mobilis TISTR 1108T,＞99％）是降解菌群中的优势菌。而降解菌群MC3CO中的优势菌是菌株5C（以及5H,Alcanivorax dieselolei NO1A,＞99％）、条带5-8代表的未培养菌株（Novosphingobium aromaticivorans DSM 12444T,99.41%）、5J（Tistrella mobilis TISTR 1108T,99.52％）和5F（以及5G,Thalassospira lucentensis DSM 14000T,＜97％）。【结论】本研究发现在大西洋洋中脊深海海水及表层沉积物中Alcanivorax、Novosphingobium、Thalassospira、Tistrella属的细菌是PAHs降解菌群中的优势菌,其中的主要降解菌是Novosphingobium属的细菌。     

一株菊酯类农药降解菌的分离鉴定及其降解酶基因的克隆

摘要：【目的】筛选分离高效降解菊酯类农药的光合细菌,研究其降解特性,并对该菌株中降解酶基因进行克隆与初步分析。【方法】根据分离菌株的细胞形态结构、活细胞光吸收特征、生理生化特征及其16S rDNA序列系统发育分析鉴定降解菌,气相色谱法测定该菌株降解菊酯类农药的能力,PCR方法克隆降解酶基因。【结果】菌株PSB07-21属红假单胞菌属（Rhodopseudomonas sp.）,其降解最佳条件为3000 lx、35℃、pH 7,在此条件下培养15 d对600 mg/L甲氰菊酯、氯氰菊酯、联苯菊酯降解率分别为     

一株玉米小斑病拮抗菌的分离、筛选及鉴定

目的：分离对玉米小斑病菌（Bipolaris maydis）有拮抗作用的内生菌株并进行菌种鉴定。方法：经表面消毒用碾碎法从水浮莲（Pistia strotiotes）中分离内生菌,用平板扩散法筛选出对玉米小斑病菌（Bipolaris maydis）有明显拮抗作用的内生菌株XJ-PL-YB-50,利用16SrDNA通用引物对其基因组DNA进行扩增,并测序得到XJPL-YB-50的部分16SrDNA序列,GenBank接收号为EU262981。经Blastn调出与菌株16SrDNA同源的序列,用软件MEGA3．1按照Neighbor-Joining方法构建16SrDNA系统发育树。结果：XJPL-YB-50的主要生理生化指标与Bacillus subtilis相符,与Bacillus subtilis AB271744处于同一分支,相似性为100％,G＋Cm01％为47．7％。结论：水浮莲内生菌XJPL-YB-50对玉米小斑病菌有明显拮抗作用,并最终鉴定为枯草芽孢杆菌Bacillus subtilis。     

拟诺卡氏菌16S rRNA,gyrB,sod和rpoB基因的系统发育分析

为了更好地了解拟诺卡氏菌属(Nocardiopsis)各物种间的系统发育关系,该属现有有效描述种的gyrB,sod和rpoB基因的部分序列被测定,结合16S rRNA基因,对拟诺卡氏菌属进行了系统发育重建。研究发现拟诺卡氏菌属gyrB,sod和rpoB基因的平均相似性分别为87.7%、87.3%和94.1%,而16S rRNA基因的平均相似性则达到96.65%,3个看家基因均比16S rRNA具有更高的分歧度。比较基于不同基因的系统树发现,由gyrB基因得到的系统树拓扑结构与16S rRNA得到的结构在亚群上基本一致。因此,gyrB基因在拟诺卡氏菌属的系统分类上比16S rRNA基因更具优越性。     

一株海藻糖合成酶产生菌的鉴定及产酶条件优化

目的对一株海洋来源的产海藻糖合成酶菌株进行鉴定及产酶条件的初步优化。方法通过16SrDNA基因序列的同源性分析,对一株来源于东海海水的海藻糖合成酶产生菌进行鉴定,并通过单因素分析初步研究其培养特性和最佳的发酵条件。结果该菌16SrDNA序列与GenBank中已知序列相比,最高相似度为100％,鉴定为假单胞菌属（Pseudomonas）,命名为Pseudomonassp．A50。其最佳碳源和氮源分别为2％麦芽糖和0．5％酵母膏,最佳NaCl浓度为2．5％,在初始pH7．8,接种量1％,装液量125mL／250mL,28℃,130r／min发酵48h,海藻糖合成酶活力达到最高。结论此产海藻糖合成酶菌株为假单胞菌属,优化后,海藻糖合成酶活力达到14．16U／mL。     

污染土壤中有机磷农药降解菌的分离及其多样性

采用添加有机磷农药的选择性培养基,在长期受有机磷农药污染的土壤中分离到7株有机磷农药降解菌,经生理生化鉴定和系统发育分析,菌株mp-1鉴定为Pseudaminobactersp.,菌株mp-2为Alcaligenessp.,菌株mp-7为Brucellasp.,其他菌株为Ochrobactrumsp.。16SrDNA序列同源性比较、系统发育分析和染色体ERIC-PCR指纹图谱扩增表明有机磷农药长期污染的土壤中有机磷农药降解菌具有丰富的多样性。     

1株根结线虫拮抗放线菌新种的分离与鉴定

从海南东寨港红树林底泥中分离到206株放线菌。利用24孔板法进行抗线虫活性筛选,获得根结线虫拮抗菌HAlll66。该菌株16SrDNA序列与Nocardiopsis trehalosi VKM Ac-942^T相似性最高（98．3％）,并在系统发育树上聚为同一分支,二者DNA—DNA杂交率38．8％。形态和生理生化特征也表现出一定差异。基于表型与分子特征,鉴定菌株HA11166为Nocardiopsis属新种,命名为东寨港拟诺卡氏菌（Nocardiopsis dongzhaigangensis sp. nov. ）。     

黑水虻幼虫肠道和体表产酶细菌的分离和鉴定

从野外收集和室内饲养的黑水虻Hermetia illucens L.幼虫体表和肠道分离出同时具蛋白质和有机磷分解能力的细菌10株,其中编号为T2、T4、T6、T7和T8的菌株分离自体表,编号为S14、S15、S16、S19和S20的菌株分离自肠道。克隆细菌的16S rDNA和DNA促旋酶B亚基编码基因gyrB对10株细菌进行了鉴定。通过16S rDNA序列确定10株菌为芽孢杆菌属,根据gyrB基因以及BLAST结果构建系统发育树,将10株细菌鉴定为枯草芽孢杆菌。然后将10株菌的gyrB基因以B.subtilis subsp.subtilisstr.168和模式菌株B.subtilis subsp.subtilis BCRC10255相应基因序列为参照构建系统发育树,分析10株菌在种的水平上的亲缘关系,发现10株菌存在亲缘关系差异,没有重复菌株。     

耐高温石油烃降解菌的筛选及性能研究

目的筛选耐高温石油烃降解菌并对降解条件进行优化。方法以大庆地区石油污染土壤的堆肥样品为研究材料,通过富集培养后分离得到耐高温石油降解菌株,选取降解率最高的菌株,对其降解条件进行了探讨。结果得到6株耐高温石油烃降解菌,其中WY 2最适温度52~58℃,pH值7,石油浓度0.5%,接种量2mL,最佳氮源为硫酸铵,通过16SrDNA序列分析,确定该菌株为地衣芽孢杆菌。结论确定了耐高温石油烃降解菌的最佳降解条件。     

The phylogenetic relationships of cyanobacteria inferred from 16S rRNA,gyrB, rpoC1 and rpoD1 gene sequences

Phylogenetic analysis of cyanobacteria was carried out using the small subunit rRNA (16S rRNA), DNA gyrase subunit B (gyrB), DNA-dependent RNA polymerase gamma subunit (rpoC1) and a principal sigma factor of E. coli sigma(70) type for DNA-dependent RNA polymerase (rpoD1) gene sequences of 24 strains which contained 5 subgroups of cyanobacteria-3 strains of the Chroococcales, 5 strains of the Pluerocapsales, 7 strains of the Oscillatoriales, 7 strains of the Nostocales and 2 strains of the Stigonematales. Degenerated PCR primers of gyrB, rpoC1 and rpoD1 genes were designed using consensus amino acid sequences registered in GenBank. The phylogenetic positions of cyanobacteria were resolved through phylogenetic analysis based on 16S rDNA, gyrB, rpoC1 and rpoD1 gene sequences. Phylogenies of gyrB, rpoC1 and rpoD1 support 16S rRNA-based classification of cyanobacteria. Interestingly, phylogenies from amino acid sequences deduced from gyrB and combined amino acid sequences deduced from rpoC1 and rpoD1 genes strongly support that of 16S rRNA, but the branching pattens of the trees based on 16S rDNA, GyrB, rpoC1, rpoD1 and combined amino acid sequences deduced from rpoC1 and rpoD1 were not congruent. In this study, we showed the correlation among phylogenetic relationships of 16S rDNA, gyrB, rpoC1 and rpoD1 genes. The phylogenetic trees based on the sequences of 16S rDNA, GyrB, rpoC1, rpoD1 and the combined amino acid sequences deduced from rpoC1 and rpoD1 showed that the lateral gene transfer of rRNA might be suspected for Synechocystis sp. PCC 6803.     

拟诺卡氏菌16S rRNA,gyrB,sod和rpoB基因的系统发育分析

为了更好地了解拟诺卡氏菌属(Nocardiopsis)各物种间的系统发育关系,该属现有有效描述种的gyrB,sod和rpoB基因的部分序列被测定,结合16S rRNA基因,对拟诺卡氏菌属进行了系统发育重建。研究发现拟诺卡氏菌属gyrB,sod和rpoB基因的平均相似性分别为87.7%、87.3%和94.1%,而16S rRNA基因的平均相似性则达到96.65%,3个看家基因均比16S rRNA具有更高的分歧度。比较基于不同基因的系统树发现,由gyrB基因得到的系统树拓扑结构与16S rRNA得到的结构在亚群上基本一致。因此,gyrB基因在拟诺卡氏菌属的系统分类上比16S rRNA基因更具优越性。     

Bacillus tianmuensis sp. nov., isolated from soil in Tianmu Mountain national natural reserve, Hangzhou, China

In a project aiming to isolate strains with the ability to produce nonribosomal peptides, a Gram-negative, endospore-forming, rod-shaped strain, designated B5^T, was isolated from a soil sample collected from Tianmu Mountain national natural reserve in Hangzhou, China. Strain B5^T contained meso -diaminopimelic acid in the cell wall peptidoglycan. The major cellular fatty acids were anteiso-C_15:0 and iso-C_15:0. The DNA G+C content was 42.5 mol%. The phylogenetic analysis based on 16S rRNA gene sequence indicated that strain B5^T fell within the genus Bacillus , with highest sequence similarity values to Bacillus barbaricus DSM 14730^T (96.4%) and Bacillus macauensis JCM 13285^T (95.5%). The isolate, however, could be distinguished from Bacillus strains with validly published names by low 16S rRNA gene sequence similarity values, distinct phenotypic and chemotaxonomic characteristics. On the basis of these polyphasic evidences, it is demonstrated that the isolate B5^T represents a novel species of the genus Bacillus , for which the name Bacillus tianmuensis sp. nov. is proposed. The type strain is B5^T (=DSM 22111^T=CGMCC 1.8879^T).     

<Emphasis Type="Italic">Acinetobacter soli</Emphasis> sp. nov., isolated from forest soil

A non-motile and rod shaped bacterium, designated strain B1(T), was isolated from forest soil at Mt. Baekwoon, Republic of Korea. Cells were Gram-negative, catalase-positive, and oxidase-negative. The major fatty acids were 9-octadecenoic acid (C(18:1) omega9c; 42%) and hexadecanoic acid (C(16:0); 25.9%) and summed feature 3 (comprising iso-C(15:0) 2-OH and/or C(16:1) omega7c; 10.0%). The DNA G+C content was 44.1 mol%. A phylogenetic tree based on 16S rRNA gene sequences showed that strain B1(T) formed a lineage within the genus Acinetobacter and was closely related to A. baylyi DSM 14961(T) (98.6% sequence similarity), followed by A. baumannii DSM 30007(T) (97.4%), A. calcoaceticus DSM 30006(T) (97.0%) and 3 genomic species (96.8 approximately 7.6%). Phenotypic characteristics, gyrB gene sequence analysis and DNA-DNA relatedness data distinguished strain B1(T) from type strains of A. baylyi, A. baumannii, and A. calcoaceticus. On the basis of the evidence presented in this study, strain B1(T) represents a novel species of the genus Acinetobacter, for which the name Acinetobacter soli sp. nov. is proposed. The type strain is B1(T) (= KCTC 22184(T)= JCM 15062(T)).     

<Emphasis Type="Italic">Arthrobacter soli</Emphasis> sp. nov., a novel bacterium isolated from wastewater reservoir sediment

A novel Gram-positive bacterium, designated SYB2T, was isolated from wastewater reservoir sediment, and a polyphasic taxonomic study was conducted based on its morphological, physiological, and biochemical features, as well as the analysis of its 16S rRNA gene sequence. During the phylogenetic analysis of the strain SYB2T, results of a 16S rRNA gene sequence analysis placed this bacterium in the genus Arthrobacter within the family Micrococcaceae. SYB2T and Arthrobacter protophormiae ATCC 19271T, the most closely related species, both exhibited a 16S rRNA gene sequence similarity of 98.99%. The genomic DNA G+C content of the novel strain was found to be 62.0 mol%. The predominant fatty acid composition was anteiso-C15:0, anteiso-C17:0, iso-C16:0, and iso-C15:0. Analysis of 16S rRNA gene sequences and DNA-DNA relatedness, as well as physiological and biochemical tests, showed genotypic and phenotypic differences between strain SYB2T and other Arthrobacter species. The type strain of the novel species was identified as SYB2T (= KCTC 19291T= DSM 19449T).     

The phylogenetic significance of peptidoglycan types: Molecular analysis of the genera Microbacterium and Aureobacterium based upon sequence comparison of gyrB, rpoB, recA and ppk and 16SrRNA genes

The type strains of 27 species of the genus Microbacterium, family Microbacteriaceae, were analyzed with respect to the phylogeny of the housekeeping genes coding for DNA gyrase subunit B (gyrB), RNA-polymerase subunit B (rpoB), recombinase A (recA) and polyphosphate kinase (ppk). The resulting gene trees were compared to the 16S rRNA gene phylogeny of the same species. The topology of neighbour-joining and maximum parsimony phylogenetic trees based upon nucleic acid sequences and protein sequences of housekeeping genes differed among each other and no gene tree was identical to that of the 16S rRNA gene tree. Only some species showed consistent clustering by all genes analyzed, but the majority of species branched with different neighbours in most gene trees. The failure to phylogenetically cluster type strains into two groups based upon differences in the amino acid composition of peptidoglycan on the basis of 16S rRNA gene sequence similarity, once leading to the union of the genera Microbacterium and Aureobacterium, was also seen in the analysis of recA, rpoB and gyrB gene and protein phylogenies. Analysis of the pkk gene and protein as well as of a concatenate tree, combining sequences of all five genes (total of 3.700 nucleotides), sees members of the former genus Aureobacterium and other type strains with lysine as diagnostic diamino acid to form a coherent cluster that branches within the radiation of Microbacterium species with ornithine in the peptidoglycan.     

Acinetobacter marinus sp. nov. and Acinetobacter seohaensis sp. nov., isolated from sea water of the Yellow Sea in Korea

Two Gram-negative, nonmotile, coccobacilli, SW-3T and SW-100T, were isolated from sea water of the Yellow Sea in Korea. Strains SW-3T and SW-100T contained ubiquinone-9 (Q-9) as the predominant respiratory lipoquinone and C18:1 omega9c and C16:0 as the major fatty acids. The DNA G+C contents of strains SW-3T and SW-100T were 44.1 mol% and 41.9 mol%, respectively. A neighbor-joining tree based on 16S rRNA gene sequences showed that the two isolates fell within the evolutionary radiation enclosed by the genus Acinetobacter. Strains SW-3T and SW-100T exhibited a 16S rRNA gene similarity value of 95.7% and a mean DNA-DNA relatedness level of 9.2%. Strain SW-3T exhibited 16S rRNA gene sequence similarity levels of 93.5-96.9% to the validly described Acinetobacter species and fifteen Acinetobacter genomic species. Strain SW-100T exhibited 16S rRNA gene sequence similarity levels of less than 97.0% to the other Acinetobacter species except Acinetobacter towneri DSM 14962T (98.0% similarity). Strains SW-3T and SW-100T exhibited mean levels of DNA-DNA relatedness of 7.3-16.7% to the type strains of some phylogenetically related Acinetobacter species. On the basis of phenotypic, phylogenetic, and genetic data, strains SW-3T and SW-100T were classified in the genus Acinetobacter as two distinct novel species, for which the names Acinetobacter marinus sp. nov. (type strain SW-3T=KCTC 12259T=DSM 16312T) and Acinetobacter seohaensis sp. nov. (type strain SW-100T=KCTC 12260T=DSM 16313T) are proposed, respectively.     

Cohnella panacarvi sp. nov., a xylanolytic bacterium isolated from ginseng cultivating soil

A Gram-positive, aerobic, rod-shaped, nonmotile, endospore-forming bacterium, designated Gsoil 349T, was isolated from soil of a ginseng field and characterized using a polyphasic approach. Comparative analysis of 16S rRNA gene sequences revealed that the strain Gsoil 349T belongs to the family Paenibacillaceae, and the sequence showed closest similarity with Cohnella thermotolerans DSM 17683T (94.1%) and Cohnella hongkongensis DSM 17642T (93.6%). The strain showed less than 91.3% 16S rRNA gene sequence similarity with Paenibacillus species. In addition, the presence of MK-7 as the major menaquinone and anteiso-C(15:0), iso-C(16:0), and C(16:0) as major fatty acids suggested its affiliation to the genus Cohnella. The G+C content of the genomic DNA was 53.4 mol%. On the basis of its phenotypic characteristics and phylogenetic distinctiveness, strain Gsoil 349T should be treated as a novel species within the genus Cohnella for which the name Cohnella panacarvi sp. nov. is proposed. The type strain is Gsoil 349T (=KCTC 13060T = DSM 18696T).     

Clostridium asparagiforme sp. nov., isolated from a human faecal sample

An obligatory anaerobic, Gram-positive, rod-shaped organism was isolated from faeces of a healthy human donor. It was characterized using biochemical, phenotypic and molecular taxonomic methods. The organism produced acetate, lactate, and ethanol as the major products of glucose fermentation. The G + C content was 53 mol%. Based on comparative 16S rRNA gene sequencing, the unidentified bacterium is a member of the Clostridium subphylum of the Gram-positive bacteria, and most closely related to species of the Clostridium coccoides cluster (rRNA cluster XIVa) [M.D. Collins et al., The phylogeny of the genus Clostridium: proposal of five new genera and eleven new species combinations, Int. J. Syst. Bacteriol. 44 (1994) 812-826]. Clostridium bolteae and Clostridium clostridioforme were identified as the most closely related described species. A 16S rRNA sequence divergence value of > 3% suggested that the isolate represents a new species. This was also supported by the gyrase-encoding gyrB gene sequences. Based on these findings, we propose the novel bacterium from human faeces to be classified as a new species, Clostridium asparagiforme. The type strain of C. asparagiforme is N6 (DSM 15981 and CCUG 48471).     

一株海洋产电菌Shewanella sp. S2的筛选和产电分析

以厦门白城海域的潮间带表面沉积物为菌种来源筛选得到一株具有电催化活性的菌株S2,该菌株的16S rRNA和gyrB基因发育树与Shewanella oneidensis MR-1同支,相似性分别为98.5%和87%,葡萄糖、木糖、半乳糖等碳源利用及最佳生长的NaCl浓度与S.oneidensis MR-1有显著差别,因此初步鉴定为Shewanella属菌株,命名为Shewanella sp.S2。初步研究了菌株S2产电活性,在以乳酸作为碳源产电时,电压最高为150mV,相应的电流密度为66.1mA/m2。