白念珠菌多位点序列分型研究

目的研究白念珠菌的分子特征,建立分子流行病学研究方法—多位点序列分型技术(MLST),并对临床分离白念珠菌进行分型研究。方法对33株白念珠菌提取DNA,选择7对管家基因进行PCR扩增并将目的基因产物测序,然后将测序结果与标准序列比对后上传至数据库,获得相应的由7对管家基因组成的等位基因谱,最后将等位基因谱再次提交网站确认序列分型(sequence types,STs)。同时与ABC(ATP-binding cassette)分型结果进行比较。结果 33株临床白念珠菌通过MLST产生了33个STs,通过ABC分类为3型。结论 MLST具有较高的分辨力,是一种方便、快速的分子生物学方法,可用于流行病学和菌群多态性的研究。     

Rapid identification of uropathogenic Escherichia coli of the O25:H4-ST131 clonal lineage using the Diversi-Lab repetitive sequence-based PCR system

Several recent studies have highlighted the emergence of a globally disseminated clone of uropathogenic and invasive Escherichia coli isolates of serotype O25:H4 and sequence type 131. The ability to characterize rapidly E. coli isolates of this lineage would facilitate enhanced surveillance for this pathogen. We have used the semi-automated Diversi-Lab repetitive PCR-based system to analyse a collection of 35 clinical isolates of uropathogenic E. coli from across the UK, with particular focus on the O25:H4-ST131 lineage. All isolates had been characterized using multilocus sequence typing (MLST), and 14 had previously been typed using pulsed-field gel electrophoresis (PFGE). The Diversi-Lab system allowed discrimination of O25:H4-ST131 isolates from those of other E. coli lineages. It was slightly more discriminatory than MLST, but was less discriminatory than PFGE. With an analysis time of <4 h between receipt of a cultured organism and provision of a typing result, the system offers information on a real-time basis, a major advantage over current practice. We suggest that introduction of the Diversi-Lab system would be useful for rapid exclusion of E. coli isolates during outbreak investigations, and that the approach could be employed for surveillance for pathogenic or antibiotic-resistant clones of this organism.     

贵州省2株牛种布鲁氏菌分子流行病学特征分析

目的 分析贵州省牛种布鲁氏菌分子流行病学特征,为牛种布鲁氏菌病疫情的防控提供科学依据。方法 采用BCSP31-PCR及AMOS-PCR技术对经传统方法鉴定为牛种布鲁氏菌的2株分离株进行属/种复核,采用基于布鲁氏菌rpoB基因的单核苷酸多态性分析了解其rpoB基因型,采用MLVA-16技术进行MLVA分型,了解其与国内牛种布鲁氏菌的聚类关系,采用MLST技术分析其序列型及其与国内菌株间的遗传进化关系。结果 2株分离株经BCSP31-PCR和AMOS-PCR技术被鉴定为布鲁氏菌属细菌,未能明确其具体种型;其rpoB基因型相同;MLVA分型与新疆牛种布鲁氏菌共享同一MLVA-16型(4-5-3-12-2-2-3-1-6-43-8-5-5-5-3-3); MLST分析鉴定2株菌均为ST2型,最小间距图显示2株菌与布鲁氏菌属内的牛种布鲁氏菌遗传距离最近,属同一个遗传复合体。结论 贵州省2株牛种布鲁氏菌分离株间的遗传距离近,与新疆牛种3型布鲁氏菌为同一MLVA型,提示分离株引起的感染可能为外省输入。     

Molecular characteristics of serotype 3 Streptococcus pneumoniae isolates among community-acquired pneumonia patients in Japan

In order to understand the spread of the erythromycin-resistant serotype 3 Streptococcus pneumoniae clone in Japan, we have assessed the molecular characteristics of this clone. Among 156 S. pneumoniae isolates recovered from adults with community-acquired pneumonia between 2003 and 2005, 42 were serotype 3 and 40 were sequence type (ST) 180/Netherlands(3)-31 by multilocus sequence typing. Thirty-eight of the 40 ST 180 isolates had acquired resistance to erythromycin via the ermB gene. Although the ermB-positive ST180 clone isolates were more susceptible to penicillin and trimethoprim-sulfamethoxazole than ermB-positive non-ST180 isolates and contained a less mutated pbp1a or pbp2b gene, without a mefA gene, the ST180 clone was highly prevalent among ermB-positive isolates. Routine surveillance for the ST180 S. pneumoniae clone may soon become necessary.     

中国不可分群脑膜炎奈瑟菌的分子分型分析

目的了解我国血清不可分群(non-serogroupable,NG)脑膜炎奈瑟菌分子分型特征。方法选取我国2005-2011年分离的104株血清不可分群脑膜炎奈瑟菌作为研究对象,采用多位点序列分型(MLST)和聚合酶链反应(PCR)基因分群(A、B、C、W、E、X、Y、Z、H、I、L、K、cnl)方法,检测其ST序列群及基因群。结果 104株不可分群脑膜炎奈瑟菌中,基因群分布为荚膜基因缺失菌株(capsule null locus,cnl)(36株)、B群(20株)、C群(14株)、W群(9株)、E群(9株)、X群(3株)和Y群(3株),余下10株未鉴定出基因群;未发现其他基因群菌株。MLST分型将104株NG菌株分为45种ST型,其中14种为新的ST型;17种ST型可归为7个序列群(65株);cnl菌株全属于ST-198序列群,B群和C群菌株以ST-4821为优势序列群,W群以ST-11和ST-174序列群为主。结论我国NG脑膜炎奈瑟菌具有基因多态性,其中ST-198序列群全部为cnl菌株。     

Changes of serotype and genotype in Streptococcus pneumoniae isolates from a Korean hospital in 2007

We investigated the change in clones and serotypes of Streptococcus pneumoniae isolates in a Korean tertiary-care hospital. Serotypes of S. pneumoniae isolates were determined by the capsular quellung method, and in vitro susceptibility testing was performed by broth microdilution method. Multilocus sequence typing was performed to determine the genotypes of the S. pneumoniae isolates. The erm(B) and mef(A) genes in erythromycin-resistant isolates were also detected using the duplex polymerase chain reaction method. During the 2 periods assayed (1998–2000 and 2007), 7-valent pneumococcal conjugate vaccine (PCV7) serotypes decreased significantly from 58.3% to 30.9% (P = 0.001). Especially, serotypes 19F and 23F decreased significantly from 31.7% to 8.5% (P < 0.0001) and from 20.0% to 7.4% (P = 0.021), respectively. In contrast to the other PCV7 serotypes, serotype 14 coupled with CC554 emerged in 2007, which may indicate no effect of PCV7 against serotype 14 isolates from Korea and the possibility of a different subtype. Of the non- PCV7 serotypes, serotype 19A increased from 8.3% to 14.9% (P = 0.227) and serotype 15 increased from 0% to 8.5% (P = 0.023). The increase of serotype 19A was due to the expansion of a preexisting clone with serotype 19A, ST320. However, S. pneumoniae isolates of serotype 15 showed diverse STs. Our data may provide helpful information in local vaccine serotype expansion or replacement in Korea.     

内蒙古布鲁氏菌临床分离株多位点序列分型研究

目的探讨临床分离布鲁氏菌的种群结构和遗传进化特征。方法布鲁氏菌标准参考菌株羊种16M,牛种544A和猪种1 330 S作为对照菌株,采用多位点序列分型(Multiple locus sequence typing,MLST)方法对内蒙古地区分离的116株羊种布鲁氏菌进行分析,确定待测菌株的序列型(STs),用软件BioNumerics(Version 5.0)构建菌株的最小生成树。结果116株羊种布鲁氏菌全部为ST8型,9个MLST位点的变异完全相同,分离株种群结构单一;羊种菌的生物型与ST型无明显关联,ST8型菌系该地区的主要流行菌种,并与内蒙古地区先前分离羊种布鲁氏菌进化程度相同,且有密切的亲缘关系。结论羊种布鲁氏菌的序列分型(ST)与常规生物分型存在差异,ST8型菌可能具有较强的环境适应性和致病性。MLST是一种较好的羊布鲁氏菌种群和进化关系研究方法,但更适合于具有较大时间跨度和地理分布菌株间的流行病学调查。     

四川自贡地区健康人群中艾伯特埃希菌的分离鉴定

目的了解长期从事鸡鸭等家禽养殖及屠宰人员等健康人群艾伯特埃希菌的携带情况。方法收集家禽养殖、屠宰人群及其他健康人粪便,经EC肉汤增菌后PCR检测eae基因,阳性样品接种麦康凯琼脂,挑选不发酵乳糖菌落,通过16SrDNA序列分析和多位点序列分型(MLST)对菌株进行鉴定。分离株进行紧密粘附素亚型和cdtB亚型分析,并通过脉冲场凝胶电泳(PFGE)分析与动物来源艾伯特埃希菌菌株间的相关性。结果从189份从事鸡鸭宰杀人员的粪便中,分离到2株艾伯特埃希菌,从58份其他健康人群粪便中分离到1株菌,而138份家禽养殖场人员粪便中未分离到菌株。3株菌株的紧密粘附素亚型分别为sigma,iota 2,nu,cdtB亚型均为Ⅱ/Ⅲ/Ⅴ亚型。PFGE显示三株菌之间的带型相似性小于80%,并且与鸡鸭等来源的艾伯特埃希菌带型不同。结论从事鸡鸭屠宰等健康人员中存在一定程度的艾伯特埃希菌携带率,但其与家禽携带艾伯特埃希菌的关系,仍需进一步研究。     

山东省动物源沙门氏菌MLST和血清分型与分布研究

目的比较动物源沙门氏菌多位点序列分型(MLST)与血清分型的差别,获得动物源沙门氏菌在山东省的分布规律。方法从山东部分地区分离78株鸡源沙门氏菌、56株鸭源沙门氏菌和20株猪源的沙门氏菌,PCR扩增七段沙门氏菌内部保守的片段序列进行多位点序列分型,同时用玻片凝集法分析其血清型。结果血清分析结果表明,鸡源沙门氏菌检出6种血清型,其中肠炎沙门氏菌占88.5%、印第安纳沙门氏菌占5.1%、汤卜逊沙门氏菌占2.6%、鼠伤寒沙门氏菌占1.3%、山夫登堡沙门氏菌占1.3%、阿格玛沙门氏菌占1.3%;鸭源沙门氏菌检出2种血清型,其中肠炎沙门氏菌占67.9%、鼠伤寒沙门氏菌占32.1%;猪源沙门氏菌检出3种血清型,其中鼠伤寒沙门氏菌占65%、德贝儿沙门氏菌占20%、肠炎沙门氏菌占15%。通过MLST分型,鸡源检出7种ST型:ST11、ST19、ST26、ST128、ST14、ST17和New1;鸭源检出3种ST型:ST11、ST19和New2;猪源检出3种:ST34、ST40和ST3007。结论整体来看,分离菌的血清型和ST型数量比较接近,二者分型能力相近。