应用DGGE方法构建健康人粪便乳杆菌标准指纹图谱

从健康人的新鲜粪便样品中分离乳杆菌,鉴定属种后构建一种能够快速鉴定未知属种乳杆菌的标准指纹图谱.首先通过形态学观察及K2O2酶等生理生化试验对所分得菌株进行初步鉴定,然后利用16S rDNA序列同源性分析方法,对所分得菌株进行16S rDNA基因扩增与测序,测序结果与GenBank中该属内茵株的16S rDNA基因序列进行同源性分析,从而准确鉴定所分得菌株.最后,利用变性梯度凝胶电泳(DGGE)方法构建标准指纹图谱.结果表明,从健康人新鲜粪便中分离得到8株乳杆茵,经鉴定,其中4株为植物乳杆菌,1株为卷曲乳杆菌,3株为发酵乳杆菌.所得标准指纹图谱中试验菌株大体被分成4类,与16S rDNA基因序列同源性分析结果相吻合.通过传统方法可以从健康成人的新鲜粪便中分离得到乳杆菌,传统方法与分子生物学方法相结合可以将其准确鉴定到种的水平.变性梯度凝胶电泳(DGGE)方法可以在种的水平上作为快速鉴定乳杆菌的一种有效工具.     

科尔沁地区传统发酵奶豆腐中乳酸菌的鉴定

从科尔沁地区采集到5份传统发酵制作的奶豆腐样中分离出12株供试菌株,将分离的菌株的16S rDNA进行PCR扩增、序列分析并构建系统发育树。结果表明,12株菌株都属于乳酸菌(Lactic Acid Bacteria),其中6株乳杆菌(Lactobacillus),分别是植物乳杆菌(Lactobacillus plantarum)4株、干酪乳杆菌(Lactobacillus casei)1株、戊糖乳杆菌(Lactobacillus pentosus)1株;6株乳球菌(Lactococcus)分别是乳糖片球菌(Pediococcus acidilactici)5株和戊糖片球菌(Pediococcus pentosaceus)1株。鉴定结果与生理生化实验结果一致,且以乳酸片球菌和植物乳杆菌为优势菌群。     

新疆特色干酪中乳酸菌的分离鉴定

从新疆不同牧区采集工艺不同的干酪制品,对其中的乳酸菌进行分离纯化、生理生化性质试验和16S rRNA分析.结果表明,分离、纯化出的104株乳酸菌种,有82株为乳杆菌属(Lactobacillus),12株为肠球菌属(Enterococcus),10株为魏斯氏菌属(Weissella).利用16S rRNA序列同源分析和系统发育树分析对具有不同生理生化特性的代表菌株进行了分子鉴定,鉴定结果为TNM-2与干酪乳杆菌(Lactobacillus casei)、Y5-4与食窦魏斯氏菌(Weissella cibaria)、NS2-2与植物乳杆菌(Lactobacillus plantarum)的同源性达到100％,NM-2与瑞士乳杆菌(Lactobacillus helveticus)、Y1-1与马乳酒乳杆菌(Lactobacillus kefiranofaciens)、WG与耐久肠球菌(Enterococcus durans)的同源性达到99％.     

一株产共轭亚油酸乳酸菌的筛选和鉴定

从酸菜汁中分离到一株具有较高产共轭亚油酸(CLA)能力的乳酸菌菌株9#,在小麦胚芽培养基中发酵CLA产量达到64.28μg/mL.通过菌株菌落形态学特征、菌体形态学特征、菌株生化特征和细菌16S rDNA进行的同源性比对,利用MEGA4软件构建系统发育树,表明菌株9#属于干酪乳杆菌(Lactobacillus casei).     

利用16S rDNA序列及tuf-RFLP鉴定蒙古国发酵乳中的乳酸菌

运用16S rDNA序列分析和tuf-RFLP技术对采于蒙古国扎布汗省的25份发酵乳样中分离出的110株乳酸菌进行鉴定。首先将分离的110株乳酸菌的16S rRNA基因进行扩增,测序并构建系统发育树,初步鉴定为41株嗜热链球菌,40株瑞士乳杆菌,11株德氏乳杆菌保加利亚亚种,2株发酵乳杆菌,1株乳明串珠菌,2株肠膜明串肠膜亚种,1株乳酸乳球乳酸亚种和12株属于干酪族的菌株。由于干酪乳杆菌族的16S rDNA序列差异很小,故采用tuf-RFLP技术对这12株进行了进一步的验证,通过分离菌株与模式菌株tuf-RFLP图谱的比较分析,结果表明这12株菌均为干酪乳杆菌。     

不同地区酸马奶中乳杆菌的分离及其生物学特性的研究

从内蒙古锡林浩特市(16份样品)和蒙古国(5份样品)以传统方法制作酸马奶的牧民家庭中共采集到21份酸马奶样品；从蒙古国样品采集的样品中共分离到30株乳杆菌、分别鉴定为Lb．casei(1株)、Lb．plantarum(9株)和Lb．acidophilus Group(20株)的菌株；从内蒙古采集的样品中的共分离到50株乳杆菌，分别为Lb．casei(16株)、Lb．plantarum(6株)和Lb．acidophilus Group(10株)、Lb．paracasei subsp．paracase(3株)、Lb．coryniformis subsp．coryniformis(5株)、Lb．curvatus(1株)、Lb．kefiranofaciens(2株)和异型乳酸发酵的Lb．fermentum(1株)、Weissella kandleri(1株)、Weissella paramesenteroides(1株)以及未明确的Lactobacillus sp．(2株)等。结果表明，蒙古国酸马奶样品中以高温性Lb．ncidophilus Group的菌株分离率较高，而内蒙古酸马奶样品中以中温性Lb．casei菌株最多，其次为Lb.acidophilus Group的菌株。     

产凝集素乳杆菌的筛选及PCR—16S rDNA鉴定

目的 从泡菜汁中分离产凝集素乳杆菌并鉴定.方法 通过凝集实验筛选产凝集素的乳杆菌,用聚合酶链式反应(PCR)扩增16S rDNA可变区基因鉴定产凝集素乳杆菌.结果 此菌株呈革兰阳性,短杆状,产乳酸.通过同源性比较,此菌株与Lactobacillus plantarum strain p215的同源性为99%.结论 分离得到了产凝集素的菌株,鉴定表明此菌株为植物乳杆菌.     

益生菌L.casei Zhang的多项分类鉴定

结合传统生理生化鉴定结果和16S rDNA序列分析方法对L.casei zhang在干酪乳杆菌这个类群中的分类学地位进行了综述.结果表明,该菌应属于干酪乳杆菌干酪亚种.     

rDNA间区序列测定在乳杆菌鉴定中的应用

利用16S-23S rDNA Short ISR区间序列分析方法,对分离自酸马奶中的12株杆菌进行鉴定。通过对Short ISR序列测序并进行BLAST比对,结合系统发育树分析,可将ZL3-1、XM2-1和XF5-2判定为Lacto-bacillus.caseisubsp.casei,MG2-1、A6-2、B10-2和XB2-3归入L.helveticus,C56、D73和E96暂定为L.ferintoshensis,无法准确判定E91和E112的归属。16S-23S rDNA Short ISR区间序列可以识别L.rhamnosus和L.casei,而对于L.ferintoshensis、L.kefir、L.buchneri和L.hilgardii无法加以区分,但是仍有助于将未知杆菌归入这一群。     

益生菌Lactobacillus casei Zhang对酸奶风味、质地及感官特性的影响

将益生菌Lactobacillus casei Zhang(Lb. casei Zhang)以1.0×107g-1的添加量与商业酸奶发酵荆YC-X11共同接种进行发酵乳制备.发酵结束(pH=4.5)于4℃冷藏24 h后,分别测定发酵乳样品的酸度、黏度、脱水收缩性、挥发性风味物质、L.delbrueckii subsp.Bulgaricus、S.thermophilus和Lb.casei Zhang活菌数及对其进行感官鉴评.结果表明.Lb.casei Zhang对发酵乳样品的酸度、黏度、脱水收缩性、S.ther-mophilus活菌数无影响(P>0.05),可促进L.delbrueckii subsp.Bulgaricus生长(P<0.05),并使酸奶样品中挥发性风味物质总的质量分数提高17.1%.从而总体上提高发酵乳的感官品质.同时Lb.casei Zhang在发酵乳中具有良好的稳定性.因而益生菌Lb.casei Zhang与商业酸奶发酵剂YC-X11复配进行发酵乳生产具有极大的可行性.     

0癌症进展Oncology Progress5R-5E0546E

ieved in the study of gynecological cancers,early diagnosis,early treatment and valid recognition of precancerous stage are still the priority of therapy.The standardization of treatment on gynecological cancers remains to be generalized and the surgeries are prone to be humanize and individuate.The technique of clinical treatment should be based on evidence-based medicine and cooperation of multi-ceneri和L.hilgardii无法加以区分,但是仍有助于将未知杆菌归入这一群。     

利用Lactobacillus casei Zhang开发益生菌新鲜干酪

通过添加不同比例Lactobacillus casei Zhang发酵剂制作新鲜干酪,并对其在新鲜干酪中的活力和添加Lactobacillus后新鲜干酪的理化性质进行了研究。结果表明,Lb.casei Zhang在新鲜干酪中具有较高的活力,添加2%、1%、0.5% Lb.casei Zhang发酵剂的干酪中,4℃冷藏开始前,Lb.casei Zhang活菌数分别为2.24×10~8 cfu/g,1.38×10~8 cfu/g,5.55×10~7 cfu/g,4℃冷藏28d后,Lb.casei Zhang存活率分别为99.12%,98.31%,98.61%。在制作过程中和4℃冷藏过程中,与空白组相比,添加Lb.casei Zhang对新鲜干酪的pH值、滴定酸度、蛋白水解活性影响都不显著(P>0.05)。     

Identification and characterization of Lactobacillus florum strains isolated from South African grape and wine samples

A total of 213 strains of lactic acid bacteria were examined in this study. Among these, 30 strains previously isolated from South African grape and wine samples remained unidentified. The identification of these isolates was performed by BLAST and phylogenetic analyses of 16S rDNA gene sequences, which indicated that the isolates belonged to Lactobacillus florum. In this work, we also designed a discriminative species-specific primer FLOR targeting the 16S rDNA gene of Lb. florum. The validity and specificity of this primer was confirmed. Of particular interest in this study was to further evaluate the identified strains for the presence of genes encoding enzymes of oenological relevance. Reference strains included three flower-associated Lb. florum (F9-1(T), F9-2 and F17) and two Lactobacillus lindneri (AWRI B530 and DSM 20691) strains. Lb. lindneri strains were incorporated as being the closest relatives of Lb. florum. PCR detection results revealed that all Lb. florum strains and Lb. lindneri AWRI B530 (grape isolate) possessed the majority of the tested genes relative to DSM 20691 (beer isolate); these enzyme-encoding genes included malolactic enzyme, peptidases (PepC, PepI, PepN), citrate lyase (α- and β-subunits), phenolic acid decarboxylase and arginine deiminase pathway enzymes (arginine deiminase and ornithine transcarbamylase). Sequence verification of PCR-generated fragments was performed by sequencing. The sequence data were used to construct the phylogenetic trees, which indicated that our Lb. florum isolates cluster with other Lb. florum strains of flower origin but rather distinct from other LAB species, with Lb. lindneri being the next closest species.     

Phenotypic and molecular identification and clustering of lactic acid bacteria and yeasts from wheat (species Triticum durum and Triticum aestivum) sourdoughs of Southern Italy

The microflora of 25 wheat sourdoughs from the Apulia region, Southern Italy, was characterized. The sourdoughs were mainly produced from Triticum durum wheat. The number of lactic acid bacteria and yeasts ranged from ca. log 7.5 to log 9.3 colony forming units (cfu)/g and from log 5.5 to log 8.4 cfu/g, respectively. About 38% of the 317 isolates of lactic acid bacteria were identified by conventional physiological and biochemical tests. Phenotypic identification was confirmed by 16S rDNA and 16S/23S rRNA spacer region PCR. Overall, 30% of the isolates were identified as Lactobacillus sanfranciscensis, 20% as Lb. alimentarius, 14% as Lb. brevis, 12% as Leuconostoc citreum, 7% as Lb. plantarum, 6% as Lactococcus lactis subsp. lactis, 4% as Lb. fermentum and Lb. acidophilus, 2% as Weissella confusa and 1% as Lb. delbrueckii subsp. delbrueckii. Some of these species have not been previously isolated from sourdoughs. Since bakers yeast is widely used in sourdough production, Saccharomyces cerevisiae was largely found. The phenotypical relationships within the main lactic acid bacteria identified were established by using cluster analysis. A microbial map of the 25 sourdoughs was plotted showing characteristic associations among lactic acid bacteria and differences in the lactic acid bacteria species which were mainly due to the species of wheat flour, use of bakers yeast and type of bread.     

Phenotypic and genotypic characterisation of Lactobacilli isolated from camel cheese produced in India

Thirty‐two Lactobacilli strains were isolated from four samples of camel cheese collected from Bikaner, India. These isolates were identified based on phenotypic and genotypic characteristics. Sequencing of 16S rDNA was performed for species identification and diversity analysis. Lactobacillus delbrueckii and Lb. fermentum were found to be dominant species followed by Lb. plantarum and Lb. casei. On evaluation of technological properties of these isolates, 20 isolates were observed to be good acid producers, eight were found positive for citrate utilisation and 11 showed presence of Prtp gene. Isolates obtained can be potential for development of defined strain starter for camel cheese.     

哈族传统乳制品中乳酸菌多样性及抑菌性研究

从新疆阿勒泰地区哈萨克族传统发酵乳制品中分离得到11株乳酸菌,采用生理生化特性和16S rDNA序列同源性分析,对其进行鉴定。通过纸片法筛选,大多数乳酸菌对大肠杆菌(Escherichia coli)、金黄色葡萄球菌(Staphylococcus aureus)、铜绿假单胞菌(Pseudomonas aerugonosa)、枯草芽孢杆菌(Bacillus subtilis)和李斯特氏菌(Listeria monocytogenes)这5株指示菌有抑菌效果,其中1株干酪乳杆菌(TNG2)和4株瑞士乳杆菌(NS2-a,TNS1-1,NS1-1和NS2-1)对5株指示菌均有抑菌效果。     

Species-specific identification of commercial probiotic strains

Products containing probiotic bacteria are gaining popularity, increasing the importance of their accurate speciation. Unfortunately, studies have suggested that improper labeling of probiotic species is common in commercial products. Species identification of a bank of commercial probiotic strains was attempted using partial 16S rDNA sequencing, carbohydrate fermentation analysis, and cellular fatty acid methyl ester analysis. Results from partial 16S rDNA sequencing indicated discrepancies between species designations for 26 out of 58 strains tested, including two ATCC Lactobacillus strains. When considering only the commercial strains obtained directly from the manufacturers, 14 of 29 strains carried species designations different from those obtained by partial 16S rDNA sequencing. Strains from six commercial products were species not listed on the label. The discrepancies mainly occurred in Lactobacillus acidophilus and Lactobacillus casei groups. Carbohydrate fermentation analysis was not sensitive enough to identify species within the L. acidophilus group. Fatty acid methyl ester analysis was found to be variable and inaccurate and is not recommended to identify probiotic lactobacilli.     

腊肠中降胆固醇乳酸菌的筛选及鉴定

通过硫磷铁比色法,该试验从腊肠中共分离到具有降胆固醇能力的乳酸菌35株,胆固醇的清除能力为9.65％～48.23％,清除率在40％以上的有11株,而乳酸菌HC12的胆固醇清除率最高(48.23％).通过对乳酸菌HC12的形态观察、生理生化试验、糖发酵试验及16S rDNA序列分析等研究,鉴定HC12为干酪乳杆菌.     

Isolation and identification of lipase producing thermophilic Geobacillus sp. SBS-4S: cloning and characterization of the lipase

A thermophilic microorganism, SBS-4S, was isolated from a hot spring located in Gilgit, Northern Areas of Pakistan. It was found to be an aerobic, gram-positive, rod-shaped, thermophilic bacterium that grew on various sugars, carboxylic acids and hydrocarbons at temperatures between 45°C and 75°C. Complete 16S rRNA gene sequence of the microorganism exhibited homology to various species of genus Geobacillus. A highest homology of 99.8% was found with Geobacillus kaustophilus. A partial (0.7 kbp) chaperonin gene sequence also showed a highest homology of 99.4% to that of G. kaustophilus whereas biochemical characteristics of the microorganism were similar to Geobacillus uzenensis. Based on biochemical characterization, 16S rRNA and chaperonin gene sequences, we identified SBS-4S as a strain of genus Geobacillus. Strain SBS-4S produced several extracellular enzymes including amylase, protease and lipase. The lipase encoding gene was cloned, expressed in Escherichia coli and the gene product was characterized. The recombinant lipase was optimally active at 60°C with stability at wide pH range (6-12). The enzyme activity was enhanced remarkably in the presence of Ca(+2). The K(m) and the V(max) for the hydrolysis of p-nitrophenyl acetate were 3.8mM and 2273 μmol min(-1)mg(-1), respectively. The ability of the recombinant enzyme to be stable at a wide pH range makes it a potential candidate for use in industry.