深圳地区2002-2008年副溶血弧菌分子特征研究

目的了解深圳地区副溶血弧菌主要血清型和分子分型以及毒力因子分布,分析新O3:K6型克隆群与国际流行株的进化关系.方法对2002-2008年深圳地区1005株副溶血弧菌临床分离株进行血清型分型;采用荧光PCR方法对68种不同血清型的28 1株副溶血弧菌进行tlh、toxR、tdh和trh毒力基因检测;对281株菌株进行orf8检测;采用脉冲场凝胶电泳(PFGE)对不同分离时间的5种主要血清型菌株进行分子分型,根据有代表性的分子分型图谱选取82株副溶血弧菌菌株进行GS-PCR检测,并选取60株副溶血弧菌菌株进行toxRS基因测序;进而对41株O3:K6和O1:K25副溶血弧菌菌株进行多位点序列分型(MLST)分析,采用eBURST软件分析进化关系.结果1005株菌共得到79种不同的血清型,主要血清型为O3:K6、O4:K8、O1:K25、O1:KUT、O4:K68、O1:K56和O9:K44,所占比例分别为57.9%、8.16%、5.27%、5.87%、1.39%、1.39%和0.99%.243株为tlh+、toxR+、tdh+、trh-,37株为tlh+、toxR+、tdh-和trh-,1株tlh+、toxR+、tdh+和trh+.35株O3:K6型菌株的MLST序列类型为ST3,是同源复合体CC3,O4:K8型副溶血弧菌的序列类型为ST189,无同源复合体.结论深圳地区主要流行的副溶血弧菌为tdh+、trh-的O3:K6、O4:K8和O1:K25菌株,2006年后出现O1:K56、O9:K44、O3:K29、O4:K9新的血清型;其中O3:K6血清型属于新的O3:K6克隆群,与其他国家流行的副溶血弧菌属于同一克隆群,同时也存在新、旧克隆群.血清的多样性和新旧克隆群的存在是造成深圳地区副溶血弧菌腹泻病高发态势的主要原因之一.     

基于不同方法的大肠埃希菌分型效果评价

目的 评价并验证成簇规律间隔短回文重复序列（CRISPRs）、血清学和多位点测序分型（MLST）方法对大肠埃希菌分型的效果。方法 使用CRISPRsFinder分析大肠埃希菌全基因组序列的CRISPRs间隔序列,使用在线软件SeroTypeFinder和MLST获得血清型和序列型（ST）;采用PCR扩增并分析349株大肠埃希菌CRISPRs,使用CRISPRs间隔序列预测血清型和ST,并比较血清学和MLST分型结果。结果 将I-E型CRISPR/Cas、I-F型CRISPR/Cas和CRISPR3-4分别命名CT-Ⅰ、CT-Ⅱ和CT-Ⅲ。根据CRISPRs间隔序列构成和排列进一步进行分型,203株大肠埃希菌被分为79个CT型别,76个血清型和66个ST。CRISPRs分型的区分能力最强,辛普森指数为0.936。CRISPRs和血清学的关联程度最高,调整兰德指数为0.908。CRISPRs型能进一步区分相同血清或ST产志贺毒素的大肠埃希菌［O157∶H7（ST11）、O104∶H4（ST678）和O26∶H11（ST21）］菌株。扩增实验室菌株的CRISPR1、CRISPR2、CRISPR3、CRISPR4和CRISPR3-4,检出率分别为81.1%、94.5%、1.4%、1.4%和4.6%;根据CRISPRs间隔序列预测O157∶H7（ST11）和ST131准确率分别为95.0%和100.0%。结论 基于CRISPRs的大肠埃希菌的分子分型方法呈现较好的分型效果和临床应用效果,预期可以成为大肠埃希菌分型的重要分子标志物。     

铜绿假单胞菌耐药率与多位点序列分型研究

目的研究铜绿假单胞菌的耐药率与分子流行病学规律,明确其进化特征,为临床抗菌药物的合理应用提供依据。方法 2010年6月-2012年10月对168株铜绿假单胞菌开展体外药物敏感性试验与多位点序列分型(MLST)研究,并应用BioNumerics与START2软件进行生物信息学分析,以揭示其流行趋势。结果 MLST方案中选取的7个管家基因GC含量分布达60.0%~70.0%;各等位基因数分布于4~10个,aroE的等位基因数只有4个,而acsA的等位基因数有10个,trpE的多态性位点最多,为16个;168株铜绿假单胞菌对除多黏菌素外的抗菌药物耐药率为16.3%~35.2%;MLST结果显示,全部菌株共形成20个ST型,其中ST244有26株、ST986有22株、ST597有16株、ST441有13株,上述4个ST型均属于克隆复合体CC244,此外,研究还发现5个新ST型。结论铜绿假单胞菌对临床常用抗菌药物的耐药率较高,呈现多药耐药趋势;铜绿假单胞菌的传播趋势以散发为主,其进化速度较快,但在医院抗菌药物选择压力的作用下也易形成典型的克隆复合体,目前需要重点监控以CC244为代表的铜绿假单胞菌的流行,以防其在医院内大规模暴发与流行。     

浙江省永康地区多重耐药鲍曼不动菌的分子分型研究

目的分析永康市第一人民医院临床分离的30株多重耐药鲍曼不动杆菌的同源性分布情况及亲缘进化关系,明确永康市第一人民医院多重耐药鲍曼不动杆菌的基因分型。方法收集永康市第一人民医院临床分离的30株多重耐药鲍曼不动杆菌,利用限制性内切酶ApaI酶切进行脉冲场凝胶电泳(PFGE)分析,利用BioNumerics软件进行聚类分析反映菌株间的同源性分布情况;通过多位点序列分型(MLST)方法反映菌株间的进化关系,经过数据共享及比较,进行基因分型。结果通过PFGE技术将30株(其中3株菌无PFGE结果)多重耐药鲍曼不动杆菌分为4个脉冲群,其中脉冲群A为主要流行克隆;经MLST分析,永康市第一人民医院感染多重耐药鲍曼不动杆菌主要流行克隆为ST208,且得到一个新的序列型。结论经PFGE和MLST分析,均得到了一个主要流行克隆。永康市第一人民医院的主要流行株ST208与世界上的主要流行克隆具有高度的同源性。     

马鞍山地区耐甲氧西林金黄色葡萄球菌的分子特征研究

目的 了解马鞍山地区耐甲氧西林金黄色葡萄球菌(金葡菌)(MRSA)肠毒素、溶血素分布情况、菌株克隆群关系及其耐药性。方法 采用全自动酶联荧光免疫系统和PCR技术分别检测肠毒素和溶血素基因分布;选择金葡菌的7个管家基因作为目的基因,对34株MRSA和3株甲氧西林敏感金葡菌(MSSA)进行多位点序列分型(MLST),然后与网上数据库比对,获得序列型(ST),根据eBURST的ST进行亲缘性分析;采用琼脂稀释法检测MRSA对12种抗生素的耐药情况。结果 210株金葡菌肠毒素阳性率为50.9%,溶血素基因携带率为97.1%,其中51株MRSA全部含有溶血素基因。34株MRSA有10个ST,以ST239为主(47.1%,16/34),其次为ST5(17.6%,6/34);3株MSSA的ST为ST188、ST1281和ST7。17株患者来源菌株分为6个ST,以ST239为主(35.3%,6/17),其次为ST5(29.4%,5/17);20株食品来源菌株有9个ST别,以ST239为主(45.0%,9/20),其次为ST7(15.0%,3/20)。ST585、ST630以及ST239的亲缘关系较近,其他ST之间亲缘关系较远。除万古霉素外,所有菌株对10种抗生素有不同程度的耐药。结论 金葡菌溶血素普遍存在;ST239为马鞍山地区MRSA的主要优势菌株,各ST间亲缘关系较远。     

四川盆地结核分枝杆菌群体遗传学特征研究

目的 探讨四川盆地(重庆、四川地区)流行的结核分枝杆菌(MTB)群体遗传学特征。方法 对413株源自四川盆地的MTB进行长序列多态性(LSP)谱系鉴定, 采用χ²检验分析不同谱系MTB在人群中的分布差异。对菌株进行15位点数目可变串联重复序列基因分型, 评价各位点的分辨力。构建N-J树分析其系统进化;构建最小跨度树以模拟群体菌株的遗传结构;采用分子方差分析不同谱系MTB的群体间遗传分化;采用贝叶斯模型计算菌株最近共祖年代。结果 通过LSP可将413株MTB分为两大谱系, 其中北京谱系占56.2%(232/413), 欧美谱系占43.8%(181/413)。两谱系MTB在人群中的分布(性别、年龄、民族以及菌株耐药性)差异无统计学意义(P>0.05)。北京谱系MTB的N-J树呈放射状分布, 最小跨度树中有72.4%(168/232)的菌株被划分到同一“克隆复合群”;欧美谱系MTB的N-J树呈枝状分布, 最小跨度树中呈现出多个“克隆复合群”。北京谱系MTB在该地区有显著的群体间遗传分化(F_ST=0.018 91, P<0.05), 而欧美谱系MTB群体间遗传分化不显著(F_ST=0.005 19, P>0.05)。欧美谱系MTB中最大菌株复合群的最近共祖年代为723(95%CI:517～946)年。结论 北京谱系和欧美谱系MTB在四川盆地以“互相对抗”形式流行;两谱系MTB的群体间遗传分化存在差异;欧美谱系MTB的侵入可能与一次历史事件(约720年前的战争)有关。     

山东省某地产超广谱β-内酰胺酶大肠埃希菌耐药情况及多位点序列分型

目的 分析山东省某生猪养殖县农村居民粪便中分离的产超广谱β-内酰胺酶（ESBLs）大肠埃希菌耐药情况和多位点序列分型。方法 采用琼脂稀释法对2016年7月收集的山东省某生猪养殖县农村居民新鲜粪便中分离的360株产ESBLs大肠埃希菌进行药物敏感性试验;对CTX-M、TEM、SHV等β-内酰胺酶基因进行PCR扩增,采用毛细管电泳筛选阳性菌株;采用多位点序列分型（MLST）方法进行分子分型,采用eBURST v3.0软件进行聚类分析。结果 360株产ESBLs大肠埃希菌对头孢噻肟、四环素、复方新诺明及氟苯尼考的耐药率较高,分别为100.0%（360/360）、82.2%（296/360）、81.1%（292/360）、80.3%（289/360）。CTX-M基因检出率为99.2%（357/360）,以CTX-M-9组、CTX-M-1组为主,分别占35.6%（128/360）和24.4%（88/360）;TEM基因检出率为26.9%（97/360）。MLST共得到132个ST型别,相对优势ST型为ST10,占12.5%（45/360）;聚类分析显示CC10为最主要的克隆群,包括39个ST型,占41.1%（148/360）。结论 该农村地区产ESBLs大肠埃希菌对头孢噻肟、四环素、复方新诺明、氟苯尼考的耐药情况较严重,存在以CC10为主的小范围聚集性,可能存在动物与人类之间的传播。     

贵州与中国大陆其它地区空肠弯曲菌MLST特征的比较分析

目的 了解贵州省鸡源空肠弯曲菌的分子流行特征,阐明各地菌株间的亲缘关系和进化特征。方法 在贵州省随机抽取7个大型养鸡场,共采集210份鸡的新鲜粪便样本,分离培养出38株空肠弯曲菌,采用多位点序列分型对分离株的序列型及克隆复合体进行构成和聚类分析,与PubMLST数据库中来自国内的分离株分子特征进行比较。结果 贵州省38株鸡源空肠弯曲菌分为23种ST型和8个克隆复合体,其中9种是目前中国大陆地区已有的ST型,2个等位基因和8种序列型为新发现的。ST型构成比前四位分别是ST - 45、ST - 161、ST - 354、ST - 4 324,与数据库中已有的6株贵州来源菌株相比,不存在共同的ST型。优势克隆复合体（CC - 45、CC - 354、CC - 52）与数据库中鸡源分离株的前三位克隆复合体没有重叠。菌株遗传进化树显示,本研究中属于优势克隆复合体的18株菌与国内部分人源、猪源、野生鸟源、鹅源分离株有较近的遗传距离。结论 贵州省鸡源空肠弯曲菌具有高度遗传多样性,在一定程度上表现出地域流行特征。序列型与国内华东地区的分离株有较高的交叉性,部分分离株与国内人、野生鸟、鹅等来源分离株遗传距离近,应加强对不同地区、不同宿主来源空肠弯曲菌的动态监测。     

广东省2014年食源性副溶血弧菌病原学特征分析

目的 了解2014年广东省食源性副溶血弧菌分离株的血清型别、抗生素敏感性、毒力基因携带情况以及分子分型特征。方法 对60株副溶血弧菌分离株进行血清分型、抗生素敏感性试验及检测耐热直接溶血素基因(tdh)和耐热相关溶血素基因(trh),并进行脉冲场凝胶电泳(PFGE)和多位点序列(MLST)分型。结果 60株分离株可分为13个血清型,主要型别为O3:K6、O4:K8、O1:K36和O4:KUT;对氨苄西林、磺胺复合物和头孢噻吩的耐药率分别为100.0%、43.3%和28.3%,有56.7%(34/60)的菌株对2类及以上抗生素同时耐药,2株菌对3类抗生素同时耐受。毒力基因PCR检测发现,有63.3%(38/60)的分离株为tdh⁺trh^-菌株,仅1株为tdh⁺trh+菌株。分子分型显示,经NotⅠ酶消化后,60株副溶血弧菌可产生48个PFGE谱型,可分为3个聚类(Cluster A、B和C)。其中Cluster B的菌株主要分离自食源性疾病监测中心散发病例,血清型以O3:K6为主,PFGE谱型的相似度在62.6%～100.0%;Cluster C主要是由4株O4:K8型菌株组成,谱型相似度为56.7%～62.5%。60株菌MLST分型可分为26个ST型,其中33株菌为ST-3型,主要是O3:K6和O1:K36菌株;4株O4:K8菌株聚集成另一个不同于ST-3型的相对优势的菌群。结论 2014年广东省副溶血弧菌菌型多样性可能是疾病高发的原因之一,O4:K8型菌株的分子特征与其他优势血清型别明显不同,应高度警惕该型菌株引起暴发的可能。     

类鼻疽伯克霍尔德菌跨洲际流行株的同源性分析及历史溯源

目的 比较不同年代分离自澳大利亚、中国海南地区7株ST562型类鼻疽伯克霍尔德菌（类鼻疽伯克菌）的同源性,并分析其传播来源。方法 利用生物信息学方法提取ST562型类鼻疽伯克菌澳大利亚临床株MSHR5858、中国海南历史菌株350105基因组中Spe Ⅰ限制性酶切片段指纹谱、多位点可变数目串联重复序列多态性指纹（MLVA-4）等遗传特征,并与近年分离的5株海南ST562型临床株的脉冲场凝胶电泳（PFGE,Spe Ⅰ酶切）、MLVA-4分子分型结果相比较。同时分析MSHR5858与350105在基因组水平的共线性及同源性。结果 5株海南ST562型临床株的PFGE带型相同（相似度>97%）且与澳大利亚临床株MSHR5858的Spe Ⅰ限制性带型一致;澳洲临床株（MSHR5858）与海南历史菌株（350105）基因组中Spe Ⅰ限制片段数分别为31和34,其中31个片段的长度一致。5株海南ST562临床株的MLVA-4型别各不相同,但HPPH43（MLVA-4指纹谱：10,8,10,8）与MSHR5858（10,8,8,6）在2341 k、1788 k两个位点重复数相同;HK003（11,8,15,7）、HK061（11,8,17,7）与历史菌株350105（11,8,11,8）在此二位点重复数也相同。此外,350105与MSHR5858在基因组水平具有良好共线性,共有基因占绝大部分,提示来源一致。结论 分离的7株ST562型类鼻疽伯克菌（包括中国海南临床分离株、历史菌株及澳大利亚临床分离株）遗传特征一致,且ST562型近年流行株与历史菌株350105可能具有同源关系。     

Development of a multilocus sequence typing (MLST) scheme for Mannheimia haemolytica and assessment of the population structure of isolates obtained from cattle and sheep

Mannheimia haemolytica is an important veterinary pathogen affecting cattle and sheep. Previous typing methods, including restriction enzyme analysis, pulsed-field gel electrophoresis and multilocus enzyme electrophoresis (MLEE) have indicated a clonal population structure. Multilocus sequence typing (MLST) has now almost replaced MLEE and is a definitive and portable typing method, allowing global data exchange. The purpose of this study was to develop a MLST scheme for M. haemolytica. A collection of isolates from 10 countries, including the type strain and reference strains for all recognized serotypes were included in the study. Partial sequences of the housekeeping genes adk, aroE, deoD, gapDH, gnd, mdh and zwf were used to define the MLST scheme. The 95 isolates demonstrated 34 different sequence types (ST) of which 19 were connected in three clonal complexes (CC). ST1 constituted more than one-third of the isolates and was most frequently demonstrated among isolates from bovine sources. The analysis indicated a common evolutionary origin of 33 isolates from the French alps, collected from domestic and wild animals and demonstrating several related STs. An analysis of 17 isolates from the USA demonstrated the same ST in 14 of the isolates. In conclusion, an unambiguous typing scheme is presented for M. haemolytica and results obtained confirm previous observations of a clonal population of the organism.     

昆明地区2018-2020年腹泻患者中艰难梭菌感染特征分析

目的对2018-2020年昆明地区腹泻患者中艰难梭菌感染特征进行分析, 为后续监测和防治提供数据支持。方法收集2018-2020年云南省4家哨点医院腹泻患者粪便标本共388份, 使用实时荧光定量PCR方法进行艰难梭菌粪便毒素基因检测, 对结果阳性的粪便标本进行菌株的分离, 用基质辅助激光解析电离飞行时间质谱鉴定菌株。提取分离菌株的基因组DNA进行多位点序列分型(MLST)。分析毒素阳性和菌株分离阳性与患者的临床特征以及艰难梭菌阳性与其他病原共感染的情况。结果 388份粪便标本中, 艰难梭菌内参tpi基因阳性标本47份, 总阳性率为12.11%。其中, 非产毒艰难梭菌4份(8.51%), 产毒艰难梭菌43份(91.49%)。47份阳性标本分离得到18株艰难梭菌, 阳性标本的分离率为38.30%。其中tcdA、tcdB、tcdC、tcdR和tcdE基因均为阳性的菌株14株。18株艰难梭菌的二元毒素均为阴性。所有分离菌株的MLST结果共形成10种序列型(ST), 其中ST37型5株(27.78%);ST129、ST3、ST54和ST2型各2株;ST35、ST532、ST48、ST27和ST3...     

Identification of a dominant Chlamydia trachomatis strain in patients attending sexual transmitted infection clinic and female sex workers in Tunisia using a high resolution typing method

BackgroundThe distribution of Chlamydia trachomatis genotypes in Tunisia was previously studied using the reverse hybridization method. In this study, we used multilocus sequence typing (MLST) to describe Chlamydia trachomatis genetic diversity among heterosexual populations in Tunisia. The obtained sequence types (STs) were compared with those from a heterosexual population from Amsterdam, the Netherlands.MethodsClinical Tunisian patients and female sex workers provided 107 Chlamydia trachomatis positive samples that were used for MLST. Samples from 256 heterosexuals visiting the Amsterdam STI clinic were included as a reference group. Six highly variable genetic regions including the ompA gene were amplified and sequenced. The ST numbers were derived from a Chlamydia typing database (http://mlstdb.uu.se) and used to draw minimum spanning trees.ResultsompA sequencing detected 7 genotypes among the Tunisian populations of which genotype E was the most prevalent (66.3%). This genotype E resolved into 23 different STs and among these the ST3 was predominant (53.5%). MLST displayed 43 STs, of which 28 (65%) were new in the database. Minimum spanning tree analysis of all Tunisian samples identified 4 clusters of which one formed a clonal cluster with samples presenting the most prevalent ST3. When comparing samples from the Tunisian and Dutch populations in one minimum spanning tree, there was little overlap between the Chlamydia trachomatis samples.ConclusionThe CT-hrMLST scheme allowed us to identify that the Tunisian distribution was dominated by one genotype E (ST3) strain which is also highly prevalent in many other countries worldwide.     

Novel Chlamydia trachomatis Strains in Heterosexual Sex Partners,Indianapolis, Indiana,USA

Chlamydia trachomatis causes a high number of sexually transmitted infections worldwide, but reproducible and precise strain typing to link partners is lacking. We evaluated multilocus sequence typing (MLST) for this purpose by detecting sequence types (STs) concordant for the ompA genotype, a single-locus typing standard. We tested samples collected during April 2000–October 2003 from members of established heterosexual partnerships (dyads) in the Indianapolis, Indiana, USA, area who self-reported being coital partners within the previous 30 days. C. trachomatis DNA from 28 dyads was tested by MLST; sequences were aligned and analyzed for ST and phylogenetic relationships. MLST detected 9 C. trachomatis STs, 4 unique to Indianapolis; STs were identical within each dyad. Thirteen unique strains were identified; 9 (32%) dyads harbored novel recombinant strains that phylogenetically clustered with strains comprising the recombinants. The high rate of novel C. trachomatis recombinants identified supports the use of MLST for transmission and strain diversity studies among at-risk populations.     

Reassessment of MLST schemes for Leptospira spp. typing worldwide

Leptospirosis is a neglected zoonosis of global importance. Several multilocus sequence typing (MLST) methods have been developed for Leptospira spp., the causative agent of leptospirosis.In this study we reassessed the most commonly used MLST schemes in a set of worldwide isolates, in order to select the loci that achieve the maximum power of discrimination for typing Leptospira spp. Global eBURST algorithm was used to detect clonal complexes among STs and phylogenetic relationships among concatenated and individual sequences were inferred through maximum likelihood (ML) analysis. The evaluation of 12 loci combined to type a subset of strains rendered 57 different STs. Seven of these loci were selected into a final scheme upon studying the number of alleles and polymorphisms, the typing efficiency, the discriminatory power and the ratio dN/dS per nucleotide site for each locus. This new 7-locus scheme was applied to a wider collection of worldwide strains. The ML tree constructed from concatenated sequences of the 7 loci identified 6 major clusters corresponding to 6 Leptospira species. Global eBURST established 8 CCs, which showed that genotypes were clearly related by geographic origin and host. ST52 and ST47, represented mostly by Argentinian isolates, grouped the higher number of isolates. These isolates were serotyped as serogroups Pomona and Icterohaemorrhagiae, showing a unidirectional correlation in which the isolates with the same ST belong to the same serogroup.In summary, this scheme combines the best loci from the most widely used MLST schemes for Leptospira spp. and supports worldwide strains classification. The Argentinian isolates exhibited congruence between allelic profile and serogroup, providing an alternative to serological methods.     

Genotyping of community-associated methicillin resistant Staphylococcus aureus (CA-MRSA) in a tertiary care centre in Mysore,South India: ST2371-SCCmec IV emerges as the major clone

The burden of community-associated methicillin resistant Staphylococcus aureus (CA-MRSA) is on the rise in population and clinical settings on account of the adaptability and virulence traits of this pathogen. We characterized 45 non-duplicate CA-MRSA strains implicated mainly in skin and soft tissue infections (SSTIs) in a tertiary care hospital in Mysore, South India. All the isolates were genotyped by staphylococcal cassette chromosome mec (SCCmec) typing, staphylococcal protein A (spa) typing, accessory gene regulator (agr) typing, and multi-locus sequence typing (MLST). Four sequence types (STs) belonging to three major clonal complexes (CCs) were identified among the isolates: CC22 (ST2371 and ST22), CC1 (ST772) and CC8 (ST8). The majority (53.3%) of the isolates was of the genotype ST2371-t852-SCCmec IV [sequence type-spa type-SCCmec type], followed by ST22-t852-SCCmec IV (22.2%), ST772-t657-SCCmec V (13.3%) and ST8-t008-SCCmec IV (11.1%). ST237I, a single locus variant of ST22 (EMRSA-15 clone), has not been reported previously from any of the Asian countries. Our study also documents for the first time, the appearance of ST8-SCCmec IV (USA300) strains in India. Representative strains of the STs were further analyzed by pulsed field gel electrophoresis (PFGE). agr typing detected type I or II alleles in the majority of the isolates. All the isolates were positive for the leukotoxin gene, pvl (Panton–Valentine leukocidin) and the staphylococcal enterotoxin gene cluster, egc. Interestingly, multidrug resistance (resistance to ⩾3 classes of non-beta-lactam antibiotics) was observed in 77.8% (n = 35) of the isolates. The highest (75.5%) resistance was recorded for ciprofloxacin, followed by erythromycin (53.3%), and quinupristin–dalfopristin (51.1%). Inducible clindamycin-resistance was identified in 37.7% of the isolates and it was attributed to the presence of erm(A), erm(C) and a combination of erm(A) and erm(C) genes. Isolates which showed a phenotypic pattern of M^R/L^S (macrolide-resistance/lincosamide-sensitivity) harbored the msr(A) gene. In conclusion, we report a high rate of multidrug resistance among Indian strains of CA-MRSA and the emergence of the lineages ST2371 and ST8 in India.     

A reliable combination method to identification and typing of epidemic and endemic clones among clinical isolates of Acinetobacter baumannii

The multi-drug resistant (MDR) Acinetobacter baumannii as an important nosocomial pathogen has emerged a global health concern in recent years. In this study, we applied three easier, faster, and cost-effective methods including PCR-based open reading frames (ORFs) typing, sequence typing of bla_OXA-51-like and RAPD-PCR method to rapid typing of A. baumannii strains. Taken together in the present study the results of ORFs typing, PCR-sequencing of bla_OXA-51-like genes and MLST sequence typing revealed there was a high prevalence (62%, 35/57) of ST2 as international and successful clone which detected among clinical isolates of multi-drug resistant A. baumannii with ORF pattern B and bla_OXA-66 gene. Only 7% (4/57) of MDR isolates belonged to ST1 with ORF pattern A and bla_OXA-69 gene. Interestingly, we detected singleton ST513 (32%, 18/57) that encoded bla_OXA-90 and showed the ORF pattern H as previously isolated in Middle East. Moreover, our data showed RAPD-PCR method can detect divergent strains of the STs. The Cl-1, Cl-2, Cl-3, Cl-4, Cl-10, Cl-11, Cl-12, Cl-13 and Cl-14 belonged to ST2. While the Cl-6, Cl-7, Cl-8 and Cl-9 belonged to ST513. Only Cl-5 belonged to ST1. It seems that the combination of these methods have more discriminatory than any method separately and could be effectively applied to rapid detection of the clonal complex (CC) of A. baumannii strains without performing of MLST or PFGE.     

Vibrio parahaemolyticus isolates from southeastern Chinese coast are genetically diverse with circulation of clonal complex 3 strains since 2002

Multilocus sequence typing (MLST) was used to examine the clonal relationship and genetic diversity of 71 Vibrio parahaemolyticus isolates from clinical and seafood-related sources in southeastern Chinese coast between 2002 and 2009. The tested isolates fell into 61 sequence types (STs). Of 17 clinical isolates, 7 belonged to ST3 of the pandemic clonal complex 3, with 3 strains isolated in 2002. Although there was no apparent clonal relationship found between clinical strains and those from seafood-related sources positive with pathogenic markers, there were clonal relationships between clinical strains from this study and those from environmental sources in other parts of China. Phylogenetic analysis showed that strains of 112 STs (61 STs from this study and 51 retrieved from PUBMLST database covering different continents) could be divided into four branches. The vast majority of our isolates and those from other countries were genetically diverse and clustered into two major branches of mixed distribution (of geographic origins and sample sources), whereas five STs representing six isolates split as two minor branches because of divergence of their recA genes, which had 80%-82% nucleotide identity to typical V. parahaemolyticus strains and 73.3%-76.9% identity to the CDS24 of a Vibrio sp. plasmid p23023, indicating that the recA gene might have recombined by lateral gene transfer. This was further supported by a high ratio of recombination to mutation (3.038) for recA. In conclusion, MLST with fully extractable database is a powerful system for analysis of clonal relationship for strains of a particular region in a national or global scale as well as between clinical and environmental or food-related strains.     

粪肠球菌临床株对泰利唑胺体外抗菌活性的研究

目的评价泰利唑胺体外抗粪肠球菌的活性,探讨泰利唑胺不敏感粪肠球菌的耐药机制及其多位点序列分型分布情况。方法收集2011年1月1日—2016年6月30日深圳市南山区人民医院临床分离的粪肠球菌菌株,使用自动化仪器法及微量肉汤稀释法对分离菌株的耐药性进行检测。应用聚合酶链式反应(PCR)方法检测恶唑烷酮类抗生素耐药基因的携带情况,并采用多位点序列分型(MLST)对分离菌株进行分型。结果共获得289株粪肠球菌,来源科室主要为外科(57.4%),标本来源主要为中段尿(126株,43.6%)。289株粪肠球菌对泰利唑胺敏感率为94.1%,对氨苄西林、呋喃西林和万古霉素有较高的敏感性(敏感率为97.9%～99.7%)。MLST结果显示,共分为47个ST型,优势ST分型为ST16和ST179,分别占29.1%(84株)和24.9%(72株),在泰利唑胺不敏感粪肠球菌中,ST16的比例高于ST179(P0.05)。共检出泰利唑胺不敏感粪肠球菌17株,其携带optrA基因比例高于敏感株。结论泰利唑胺对粪肠球菌的抗菌活性整体优于利奈唑胺,但对携带optrA基因的粪肠球菌则未显示出良好的抗菌活性。