小溪自然保护区嗜盐及耐盐菌抗菌活性筛选及生物学特征

以8个敏感菌株(3株革兰阳性菌、3株革兰阴性菌和2株真菌)作为指示菌,采用管碟法对分离自湖南小溪自然保护区普通非盐环境(ordinary non-saline environment)土样中的114株嗜盐及耐盐细菌(含放线菌)进行抗菌活性筛选,并对抗菌活性较强的菌株进行了基于16S rRNA基因序列的系统发育分析和生物学特性研究。结果显示,74株受试菌株的发酵产物具有抗菌活性,阳性率为64.9%。综合分析形态特征、生理生化特征和基于16S rRNA基因序列的系统发育分析数据,表明9株具有较强抗菌活性的菌株分别属于Bacillus属(JSM 081049、JSM 082021-1、JSM 082056、JSM 082080、JSM 082081-1、JSM 082097)、Arthrobacter属(JSM082018)、Brachybacterium属(JSM082044)和Streptomyces属(JSM082030)。     

栉江珧相关细菌抗菌活性筛选及系统发育分析

采用8种敏感指示菌(革兰阳性和阴性菌各3株,真菌2株),利用琼脂扩散法,对分离自南海硇洲岛潮汐带栉江珧(Atrina pectinata)样品的细菌进行抗菌活性筛选,并对具有较强生物活性的菌株进行系统发育分析和生物学特征观察.结果显示,125个分离菌株中,有18株的发酵液具有抗菌活性,占受试菌种总数的14.4％,其中6株(JSM 112006、JSM 114095、JSM 114060、JSM 112019、JSM 114054、JSM 114093)具有较强的抗茵活性.基于16S rRNA基因序列的系统发育分析表明,这些活性菌株类群多样性较高,分属于4个属:菌株JSM 112006属于Marinobacter属;JSM 114095属于Halomonas属;JSM 114060和JSM 112019属于Salinivibrio属;JSM 114054和JSM 114093属于Staphylococcus属.研究结果表明,南海硇洲岛潮汐带栉江珧分离菌株中存在一定比例的抗菌活性菌株,且这些细茵具有较为丰富的类群多样性.     

泥河湾大长梁遗址和马圈沟Ⅰ文化层放线菌的分离及系统发育分析

为分析泥河湾盆地大长梁遗址和马圈沟Ⅰ文化层中可培养放线菌的物种多样性,本实验通过平板稀释涂布法对该区域土样进行放线菌的物种分离,得到197株放线菌纯培养物。采用链霉菌属特异引物对放线菌纯培养物进行初步鉴定,确定143株放线菌归属于链霉菌属。对未被鉴定的54株放线菌进行16S rDNA全序列测定并分析其系统发育关系。最终,将197株放线菌归属于放线菌纲下的5个亚目,6个科,10个属。其中,马圈沟Ⅰ文化层的分离菌株12-9与Nocardioides albus KCTC 9186T之间的16S rDNA序列相似性是93.73%,可能是一个潜在新属;大长梁遗址的分离菌株18-64与Haloglycomyces albus YIM 92370T的16S rDNA序列相似性是93.64%,可能是一个潜在新属。     

分离自甘肃天祝五台岭土壤的放线菌拮抗性的初步研究

从甘肃天祝县土壤中分离到的27株放线菌,作为测试放线菌拮抗性的供试茵株.抗菌试验表明,14株放线菌对6株革兰氏阳性菌和阴性菌以及白色念殊菌等指示菌有抗性.菌株WTL-4、WTL-20和WTL-26抗菌谱最广,WTL-4能抗3种革兰氏阳性菌,WTL-20能抗革兰氏阳性的金黄色葡萄球菌和单增李斯特菌、革兰氏阴性的沙门氏菌及真菌白色念殊菌,WTL-26能抗革兰氏阳性的金黄色葡萄球菌和单增李斯特菌及革兰氏阴性的沙门氏菌.抗肿瘤试验表明,有8株菌株对肿瘤细胞有抑制作用,其中抑制率在60%以上的菌株占18.5%.对抗肿瘤活性较高的3株菌WTL-4、WTL-14和WTL-15以及没有抗菌和抗肿瘤活性的菌株WTL-27进行了系统发育分析.从系统发育树可以看出,4株菌均属于链霉菌属(Streptomyces).菌株WTL-4与Streptomyces viridochromoqenes的相似性为98%;菌株WTL-14和WTL-15的相似性达到99.2%,它们与S.flavolimosus 174457的相似性达到99.4%;WTL-27与S.mycroflavus的相似性为99%.     

硇洲岛潮汐带牡蛎相关可培养细菌多样性

[目的]了解南海硇洲岛潮汐带香港巨牡蛎(Crassostrea hongkongensis)相关可培养细菌的多样性.[方法]应用纯培养法分离样品中的细菌(含放线菌),采用基于16S rRNA基因序列的系统发育分析方法研究这些分离菌株的生物多样性.[结果]用补充0-25％(W/V) NaC1的MA、MH和NA培养基从样品中分离到102株细菌,在形态观察和部分生理生化实验结果的基础上去冗余,选取74个代表性菌株进行基于16S rRNA基因序列的系统发育分析.结果表明,这些菌株归为38个物种,属于4个大的系统发育类群(Gamma-Proteobacteria,Alpha-Proteobacteria,Bacteroidetes,Firmicutes)的16个科、18个属.大多数菌株属于Gamma-Proteobacteria亚门(45株,60.8％),其余依次是Firmicutes门(12株,16.2％)、Bacteroidetes门(11株,14.9％)和Alpha-Proteobacteria亚门(6株,8.1％).大多数菌株与其系统发育关系最密切的有效发表种典型菌株之间(16S rRNA基因序列相似性为94.3％-99.8％)存在一定的遗传差异,其中JSM 111069可能代表Carnobacteriaceae科的潜在新属;菌株JSM 111039和JSM 111085可能分别代表Bacillus属的两个新物种,菌株JSM 111020、JSM 111072和JSM 111090可能分别代表Pseudoalteromonas、Proteus和Idiomarina属的新物种.[结论]南海硇洲岛潮汐带牡蛎中存在较为丰富的原核生物多样性,并潜藏着一定数量的微生物新类群(物种).     

硇洲岛牡蛎相关细菌抗菌筛选及系统发育分析

以8个敏感菌株作为指示菌,采用管碟法对分离自湛江硇洲岛（20°52′N～20°56′N,110°33′E～110°38′E）潮汐带香港巨牡蛎(Crassostrea hongkongensis)中的72株细菌的发酵液进行抗菌筛选,并对阳性菌株进行基因组DNA提取、16S rRNA基因PCR扩增和序列测定,继而进行系统发育分析。抗菌实验结果表明,受试菌株中有23株菌的发酵产物具有抗菌活性（阳性率31.9%）,其中有5个菌株（JSM 111024、JSM 111027、JSM 111029、JSM 111076、JSM 111083）具有较强的抗菌活性。基于16S rRNA基因序列的系统发育分析表明,这23株菌具有较高的类群多样性和物种多样性,属于3个系统发育群/门(Alpha Proteobacteria、Gamma Proteobacteria、Bacteroidetes)中的8个科(Aeromonadaceae、Flavobacteriaceae、Halomonadaceae、Idiomarinaceae、Phyllobacteriaceae、Pseudoalteromonadaceae、Shewanellaceae、Xanthomonadaceae)的8个属(Idiomarina、Halomonas、Myroides、Nitratireductor、Oceanimonas、Pseudoalteromonas、Shewanella、Wohlfahrtiimonas),可分为11个物种。优势类群为Gamma Proteobacteria亚门（14株）,其中优势属为Oceanimonas属（6株）;第二大类群为Bacteroidetes门（7株）,都属于Flavobacteriaceae科的Myroides属。具有较强抗菌活性的5个菌株中,有4个菌株（JSM 111024、JSM 111027、JSM 111029、JSM 111083）属于Alpha Proteobacteria 亚门Phyllobacteriaceae科〖WTBX〗Oceanimonas属,而菌株JSM 111076属于Gamma Proteobacteria 亚门Aeromonadaceae科Nitratireductor属。     

小溪自然保护区非盐环境土壤中嗜盐和耐盐菌多样性

【目的】研究湖南小溪国家级自然保护区普通非盐环境(ordinary non-saline environment)土壤样品中可培养嗜盐及耐盐细菌(含放线菌)多样性。【方法】采用纯培养法和基于16S rRNA基因序列的系统发育分析对样品中嗜盐及耐盐细菌多样性进行研究。【结果】用补充5%-20%(w/v)NaCl的MA、ISP2、ISP5、NA和HAA培养基从土壤样品中分离到114株细菌,其中8株为中度嗜盐菌,19株为轻度嗜盐菌,87株为耐盐菌。根据形态观察和部分生理生化实验结果去冗余,选取61个代表性菌株进行基于16S rRNA基因序列的系统发育多样性分析。结果表明,这些菌株属于细菌域(Bacteria)的3个大的系统发育类群(门;phylum)(Actinobacteria,Firmicutes,Proteobacteria)的16个科、18个属,代表了41个物种。多数菌株属于Firmicutes门(38株,62.3%)和Actinobacteria门(18株,29.5%)。大多数菌株与其系统发育关系最密切的已知物种的典型菌株之间存在一定的遗传差异(16S rRNA基因序列相似性为96.9%-99.8%),其中有7个菌株(JSM070026,JSM081004,JSM081006,JSM081008,JSM083058,JSM083085,JSM084035)代表7个潜在新种(potential novel species)。【结论】研究结果表明,湖南小溪国家级自然保护区普通非盐环境土壤中存在较为丰富的可培养嗜盐及耐盐细菌多样性,并且潜藏着较多新的微生物类群(物种)。     

拮抗放线菌B-20的种类鉴定及其16S rDNA序列分析

对分离自东北蔬菜保护地土壤的1株拮抗放线菌菌株B-20进行了形态特征、培养特征、生理生化、细胞壁组分分析及16S rDNA序列分析。基内菌丝无横隔、不断裂,气生菌丝多分枝;孢子丝波曲至螺旋形,孢子椭圆形,表面光滑。细胞壁化学组分Ⅰ型。以16s rDNA序列为基础构建了包括13株相关种属细菌在内的系统发育树。根据多相分类鉴定结果表明,放线菌B-20的上述特征与淡紫灰链霉菌(Streptomyces lavendulae)的高度一致。二者的16S rDNA序列的相似性达到了99%。因此,可将链霉菌B-20定名为淡紫灰链霉菌B-20 (S.lavendulae B-20)。     

药用植物内生放线菌的分离、筛选及活性菌株YIM 61470鉴定

从云南西双版纳热带雨林多种药用植物中分离到272株内生放线菌,活性筛选表明 146株菌的发酵产物具有抗菌活性,其中94株菌具有拮抗病原细菌活性,127株菌具有抑制病原真菌的功能.分离菌株YIM 61470具有广谱抗菌活性,通过形态特征、培养特征、生理生化特征、细胞化学分类特征和基于16S rRNA基因序列的相似性分析等研究,菌株YIM 61470被鉴定为链霉菌属(Streptomyces)氢化链霉菌(S.llydrogenans)的一个菌株.     

一株白芍内生放线菌的分离、活性及系统发育分析

目的:分离白芍中拮抗农作物致病菌和人类常见病原菌的内生放线菌,并进行系统发育分析。方法:采用3种分离培养基,从白芍根部分离内生放线菌;通过滤纸片法筛选具有拮抗活性的菌株,观察菌丝形态,并进行16S rDNA序列系统发育分析。结果:从白芍中分离得到16株内生放线菌,其中从FYSCA培养基中分离到9株;16株内生放线菌中有6株具有拮抗作用,菌株S-BS033004对5种病原菌有拮抗活性,尤其是对棉花黄萎病菌和小孢拟盘多毛孢菌和耐青霉素类金黄色葡萄球菌的拮抗作用显著,抑菌圈≥20mm。经16S rDNA系统发育分析表明该菌株与Streptomyces anulatus NBRC13369T等6株链霉菌模式菌株亲缘关系较近,相似性均为99.7%。结论:白芍内生放线菌S-BS033004是一株杀菌谱较广的链霉菌,具有很好的开发潜力。     

发酵车间空气源细菌淀粉酶产生菌筛选及多样性分析

采用平板水解圈法(plate hydrolysis spot method)对分离自酒鬼酒发酵车间空气样品的细菌进行淀粉酶产生菌筛选,运用基于16S rRNA基因序列的分析方法对高酶活菌株进行系统发育多样性分析。结果表明,73个受试菌株中,有23株为淀粉酶产生菌,占受试菌株的31.5%,其中有9株为高酶活菌。23个淀粉酶产生菌类群多样性和物种多样性较高,属于4个大的系统发育类群(Actinobacteria、Deinococcus-Thermus、Firmicutes、Proteobacteria)中的10个科(Bacillaceae、Deinococcaceae、Intrasporangiaceae、Microbacteriaceae、Micrococcaceae、Nocardiaceae、Propionibacteriaceae、Pseudomonadaceae、Rhodobacteraceae、Xanthomonadaceae)的13个属,可分为21个物种。进一步分析表明,9株高酶活菌属于细菌域(Eubacteria)的4个大的系统发育类群(Actinobacteria、Deinococcus Thermus、Firmicutes、Proteobacteria)的7个科(Bacillaceae、Deinococcaceae、Micrococcaceae、Microbacteriaceae、Nocardiaceae、Rhodobacteraceae、Xanthomonadaceae),归属于8个属。研究结果表明,酒鬼酒发酵车间空气源细菌存在较高比例的淀粉酶产生菌,且这些菌株具有较高的类群多样性和物种多样性。     

<Emphasis Type="Italic">Halobacillus hunanensis</Emphasis> sp. nov., a moderately halophilic bacterium isolated from a subterranean brine

A moderately halophilic, Gram-positive, catalase- and oxidase-positive, rod-shaped, aerobic bacterium, designated strain JSM 071077^T, was isolated from a subterranean brine sample collected from a salt mine in Hunan Province, China. Cells were motile by means of peritrichous flagella and formed ellipsoidal endospores lying in subterminal swollen sporangia. Strain JSM 071077^T was able to grow with 2–25% (w/v) total salts (optimum, 5–10%), at pH 6.0–10.0 (optimum, pH 7.5) and 10–40°C (optimum, 25–30°C). meso-Diaminopimelic acid was present in the cell-wall peptidoglycan. The predominant menaquinone was MK-7, and the major cellular fatty acids were anteiso-C_15:0, anteiso-C_17:0 and iso-C_15:0. The genomic DNA G+C content was 41.8 mol%. Phylogenetic analyses based on 16S rRNA gene sequence comparisons revealed that strain JSM 071077^T should be assigned to the genus Halobacillus, being related most closely to the type strain of Halobacillus naozhouensis (98.8% sequence similarity), and the two strains formed a distinct subline in the neighbor-joining, minimum-evolution and maximum-parsimony phylogenetic trees. The sequence similarities between the novel isolate and the type strains of other recognized Halobacillus species ranged from 97.6% (with Halobacillus alkaliphilus) to 95.2% (with Halobacillus kuroshimensis). The results of the phylogenetic analyses, combined with DNA–DNA relatedness data, phenotypic characteristics and chemotaxonomic information, support that strain JSM 071077^T represents a new species of the genus Halobacillus, for which the name Halobacillus hunanensis sp. nov. is proposed. The type strain is JSM 071077^T (=DSM 21184^T = KCTC 13235^T).     

<Emphasis Type="Italic">Virgibacillus litoralis</Emphasis> sp. nov., a moderately halophilic bacterium isolated from saline soil

A Gram-positive, moderately halophilic, endospore-forming, catalase- and oxidase-positive, motile, rod-shaped, aerobic bacterium, designated strain JSM 089168^T, was isolated from saline soil collected from Naozhou Island, Leizhou Bay, South China Sea. The organism was able to grow with 2–25% (w/v) total salts (optimum, 5–10%), at pH 6.0–10.0 (optimum, pH 8.0) and 10–45°C (optimum, 30°C). meso-Diaminopimelic acid was present in the cell-wall peptidoglycan. The strain contained MK-7 as the predominant menaquinone, and diphosphatidylglycerol and phosphatidylglycerol as the major polar lipids. The major cellular fatty acids were anteiso-C_15:0, iso-C_15:0 and anteiso-C_17:0, and the DNA G + C content was 40.2 mol%. Phylogenetic analysis based on 16S rRNA gene sequence comparisons revealed that strain JSM 089168^T should be assigned to the genus Virgibacillus, being related most closely to the type strains of Virgibacillus carmonensis (sequence similarity 97.6%), Virgibacillus necropolis (97.3%) and Virgibacillus halodenitrificans (97.1%). Levels of DNA–DNA relatedness between strain JSM 089168^T and the type strains of V. carmonensis, V. necropolis and V. halodenitrificans were 20.4, 14.3 and 12.0%, respectively. The combination of phylogenetic analysis, DNA–DNA relatedness, phenotypic characteristics and chemotaxonomic data supported the view that strain JSM 089168^T represents a novel species of the genus Virgibacillus, for which the name Virgibacillus litoralis sp. nov. is proposed. The type strain is JSM 089168^T (=DSM 21085^T =KCTC 13228^T). The GenBank/EMBL/DDBJ accession number for the 16S rRNA gene sequence of strain JSM 089168^T is FJ425909.     

Virgibacillus zhanjiangensis sp. nov., a marine bacterium isolated from sea water

A Gram-positive, endospore-forming, catalase- and oxidase-positive, motile, rod-shaped, aerobic bacterium, designated strain JSM 079157^T, was isolated from surface seawater off the coastline of Naozhou Island in South China Sea. The organism was able to grow with 1–15% (w/v) total salts (optimum, 4–7%), and at pH 6.0–10.0 (optimum, pH 7.5) and 10–45°C (optimum, 30°C). meso-Diaminopimelic acid was present in the cell-wall peptidoglycan. The predominant menaquinone was MK-7, and the polar lipids were diphosphatidylglycerol and phosphatidylglycerol. The major cellular fatty acids were anteiso-C_15:0 (45.1%) and anteiso-C_17:0 (16.2%), and the DNA G + C content was 39.5 mol%. A phylogenetic analysis based on 16S rRNA gene sequence comparisons revealed that strain JSM 079157^T should be assigned to the genus Virgibacillus, being related most closely to the type strains of Virgibacillus litoralis (97.4% sequence similarity), Virgibacillus necropolis (97.3%) and Virgibacillus carmonensis (97.1%). These four strains formed a distinct subcluster in the phylogenetic tree. The levels of DNA–DNA relatedness between the new isolate and the type strains of V. litoralis, V. necropolis and V. carmonensis were 30.4, 19.3 and 12.6%, respectively. The results of the phylogenetic analysis, combined with DNA–DNA relatedness data, phenotypic characteristics and chemotaxonomic information, support the suggestion that strain JSM 079157^T represents a new species of the genus Virgibacillus, for which the name Virgibacillus zhanjiangensis sp. nov. is proposed. The type strain is JSM 079157^T (=DSM 21084^T = KCTC 13227^T).     

Bacillus hunanensis sp. nov., a slightly halophilic bacterium isolated from non-saline forest soil

A novel Gram-stain-positive, slightly halophilic, catalase- and oxidase-positive, endospore-forming, motile, aerobic, rod-shaped bacterium, designated strain JSM 081003^T, was isolated from non-saline forest soil in Hunan Province, China. Growth occurred with 0.5?C15% (w/v) NaCl (optimum 2?C4%) at pH 6.5?C10.5 (optimum pH 7.5?C8.5) and at 5?C40°C (optimum 30°C). meso-Diaminopimelic acid was present in the cell-wall peptidoglycan. The major cellular fatty acids were iso-C15:0, anteiso-C15:0 and iso-C14:0. Strain JSM 081003^T contained MK-7 as the predominant respiratory quinone, and diphosphatidylglycerol, phosphatidylethanolamine and phosphatidylglycerol as the major polar lipids. The genomic DNA G + C content of strain JSM 081003^T was 40.9 mol%. A phylogenetic analysis based on 16S rRNA gene sequence comparisons revealed that strain JSM 081003^T should be assigned to the genus Bacillus, and was related most closely to the type strains of Bacillus lehensis (sequence similarity 99.6%), Bacillus oshimensis (99.4%) and Bacillus patagoniensis (96.6%); lower than 96.0% sequence similarity was observed with other Bacillus species. The combination of phylogenetic analysis, DNA?CDNA relatedness values, phenotypic characteristics and chemotaxonomic data supports the view that strain JSM 081003^T represents a new species of the genus Bacillus, for which the name Bacillus hunanensis sp. nov. is proposed. The type strain is JSM 081003^T (= DSM 23008^T = KCTC 13711^T).     

Identification of Streptomyces sp. nov. WH26 producing cytotoxic compounds isolated from marine solar saltern in China

A moderately halophilic actinomycetes strain, designated as WH26, was isolated from Weihai Solar Saltern in China. The identification of the strain WH26 was performed by its morphological characteristics, physiological and biochemical tests as well as phylogenetic analysis based on 16S rRNA sequence comparison. The results showed that the nucleotide sequence of the 16S rRNA gene (1,677 bp) of the strain WH26 exhibited close similarity (97–99 %) with other Streptomyces 16S rRNA genes and the strain WH26 was identified to belong to the genus Streptomyces. An ethyl acetate extraction of Streptomyces sp. nov. WH26 demonstrated significant cellular toxicity. Two compounds, 8-O-methyltetrangulol and naphthomycin A were isolated from the extract via silica gel column chromatography and HPLC. These two compounds showed potent cytotoxic activity against several human tumor cell lines including A549, HeLa, BEL-7402 and HT-29. The present studies suggest that moderately halophilic actinomycetes may be a novel biological source for the discovery of anticancer agents.     

Streptomyces polyrhachii sp. nov., a novel actinomycete isolated from an edible Chinese black ant (Polyrhachis vicina Roger)

A novel actinomycete, designated strain NEAU-ycm1^T, was isolated from an edible Chinese black ant (Polyrhachis vicina Roger) and characterized with a polyphasic approach. The organism was found to have morphological and chemotaxonomic characteristics typical of streptomycetes. Phylogenetic analysis based on the almost complete 16S rRNA gene sequence show that the novel isolate belongs to the genus Streptomyces and forms a separate subclade. The closest phylogenetic relatives were identified as the type strains of Streptomyces intermedius NBRC 13049^T (97.74 %), Streptomyces aureoverticillatus NRRL B-3326^T (97.69 %), Streptomyces rutgersensis NBRC 12819^T (97.68 %), Streptomyces gougerotii NBRC 3198^T (97.68 %) and Streptomyces diastaticus subsp. diastaticus NBRC 3714^T (97.68 %). Similarities to other type strains of the genus Streptomyces were lower than 97.55 %. A comparison between strain NEAU-ycm1^T and the closest related Streptomyces type strains revealed that it is different from them in morphological, physiological and biochemical characteristics. Therefore, it is proposed that NEAU-ycm1^T (=CGMCC 4.7094^T = DSM 42102^T) represents a novel species of the genus of Streptomyces, for which the name Streptomyces polyrhachii sp. nov. is proposed.     

<Emphasis Type="Italic">Arthrobacter halodurans</Emphasis> sp. nov., a new halotolerant bacterium isolated from sea water

A novel Gram-positive, halotolerant, non-sporulating, non-motile, catalase-positive, oxidase-negative and aerobic bacterium, designated strain JSM 078085^T, was isolated from sea water collected from the South China Sea. Strain JSM 078085^T exhibited a rod-coccus growth cycle and produced a yellow pigment. The strain was able to grow in the presence of 0–12% (w/v) NaCl and at pH 6.0–9.5 and 4–35°C; optimum growth was observed at pH 7.0 and 25–30°C in the absence of NaCl. The peptidoglycan type was A4α (l-Lys–l-Ala–l-Glu). Cell-wall sugars contained galactose and glucose. Strain JSM 078085^T contained menaquinone MK-9(H₂) as the major respiratory quinone and diphosphatidylglycerol, phosphatidylglycerol and phosphatidylinositol as the major polar lipids. The major cellular fatty acids were anteiso-C_15:0, iso-C_15:0 and anteiso-C_17:0 and the DNA G + C content was 63.3 mol%. Phylogenetic analysis based on 16S rRNA gene sequence comparisons revealed that strain JSM 078085^T should be assigned to the genus Arthrobacter, being most closely related to the type strain of Arthrobacter rhombi (sequence similarity 97.1%), and the two strains formed a distinct lineage in the phylogenetic tree. The level of DNA–DNA relatedness between strain JSM 078085^T and the type strain of Arthrobacter rhombi was 10.6%. The combination of phylogenetic analysis, DNA–DNA relatedness, phenotypic characteristics and chemotaxonomic data supported the view that strain JSM 078085^T represents a novel species of the genus Arthrobacter, for which the name Arthrobacter halodurans sp. nov. is proposed. The type strain is JSM 078085^T (=DSM 21081^T=KCTC 19430^T). The GenBank/EMBL/DDBJ accession number for the 16S rRNA gene sequence of strain JSM 078085^T is EU583729.     

Isolation of Three Xylanase-Producing Strains of Actinomycetes and Their Identification Using Molecular Methods

Eighty-eight actinomycetes were isolated from 20 samples collected from different locations in and around Delhi, India. Among these, 69 isolates were found positive for xylanase production. Of 69 isolates, 3 (SN32, SN77, and SN83) produced >125 IU/ml xylanase. Modern genetic tools were used for revealing the identities of these potent xylanase producers. The selected isolates were categorized under the genus Streptomyces based on their cultural and morphologic characteristics. Genetic diversity among these species of Streptomyces was established based on restriction length fragment polymorphism and random amplified polymorphic DNA analysis. The closest phylogenetic neighbours according to the 16S rRNA gene-sequence data for the three isolates SN32, SN77, and SN83 were Streptomyces cyaneus, S. tendae, and S. caelestis, respectively.     

Application of rpoB sequence similarity analysis, REP-PCR and BOX-PCR for the differentiation of species within the genus Geobacillus

Aim:  To investigate the applicability of rpoB gene, which encodes the β subunit of RNA polymerase, to be used as an alternative to 16S rRNA for sequence similarity analysis in the thermophilic genus Geobacillus. Rapid and reproducible repetitive extragenic palindromic fingerprinting techniques (REP‐ and BOX‐polymerase chain reaction) were also used. Methods and Results:  rpoB DNA (458 bp) were amplified from 21 Geobacillus‐ and Bacillus type strains, producing different BOX‐ and REP‐PCR profiles, in addition to 11 thermophilic isolates of Geobacillus and Bacillus species from a Santorini volcano habitat. The sequences and the phylogenetic tree of rpoB were compared with those obtained from 16S rRNA gene analysis. The results demonstrated between 90–100% (16S rRNA) and 74–100% (rpoB) similarity among examined bacteria. Conclusion:  BOX‐ and REP‐PCR can be applied for molecular typing within Geobacillus genus. rpoB sequence similarity analysis permits a more accurate discrimination of the species within the Geobacillus genus than the more commonly used 16S rRNA. Significance and Impact of the Study:  The obtained results suggested that rpoB sequence similarity analysis is a powerful tool for discrimination between species within the ecologically and industrially important strains of Geobacillus genus.