新生儿病区产ESBLs大肠埃希菌整合子及其耐药性分析

目的 了解新生儿病区产ESBLs大肠埃希菌整合子的携带情况及其耐药性.方法 采用K-B琼脂扩散法对56株产ESBLs大肠埃希菌进行药敏试验;应用PCR法检测Ⅰ类、Ⅱ类和Ⅲ类整合子;以肠杆菌科重复序列-聚合酶链式反应(ERIC-PCR)进行基因分型.结果 56株产ESBLs大肠埃希菌的Ⅰ类整合子检出率为60.7％,未检出Ⅱ类和Ⅲ类整合子;菌株对庆大霉素、环丙沙星、左氧氟沙星、复方新诺明、头孢唑林、氨曲南、头孢他啶的耐药率差异有统计学意义(P＜0.05),阳性菌株的耐药率高于阴性菌株;56株大肠埃希菌分为45种基因型.结论 Ⅰ类整合子广泛存在于新生儿病区产ESBLs大肠埃希菌并与其耐药性相关.     

大肠埃希菌耐药性水平传播实验研究

目的研究重症监护病房（ICU）患者标本中分离的大肠埃希菌的耐药情况以及耐药性水平传播的实验研究。方法采取双纸片法（K-B）检测细菌的耐药性;产超广谱β-内酰胺酶（ESBLs）大肠埃希菌为供体菌,耐利福平大肠埃希菌（对其他抗生素敏感）作为受体菌进行接合实验;采用聚合酶链反应（PCR）技术扩增整合子和耐药基因。结果30株大肠埃希菌中产ESBLs菌株检出率为46．7％;接合培养后,接合菌携带23kb和25kb大质粒,而无供体菌中一系列小质粒;供体菌和接合菌均携带I型整合子。结论大肠埃希菌耐药性严重,且呈多重耐药性;产ESBLs菌株可通过质粒和整合子将耐药基因转移给敏感菌,导致耐药性传播。     

产ESBLs大肠埃希菌整合子及其相关基因盒的研究

目的检测产超广谱β-内酰胺酶（ESBLs）大肠埃希菌中整合子的整合酶及插入的相关基因盒情况,分析整合子对细菌耐药性的影响。方法采用K-B琼脂扩散法对45株临床分离的产ESBLs大肠埃希菌进行药敏试验;应用PCR法检测45株产ESBLs大肠埃希菌Ⅰ类、Ⅱ类和Ⅲ类整合子;对Ⅰ类整合子阳性菌进行整合子相关基因盒检测。结果45株菌中有27株（60．0％）含有Ⅰ类整合子,没有检测到Ⅱ类和Ⅲ类整合子阳性菌。在Ⅰ类整合子阳性菌中,有23株携带Ⅰ类整合子相关基因盒（85．2％）,5种不同的基因盒图谱,片段大小在600～2322bp,分离自同一科室的部分菌株携带大小相同的基因盒;Ⅰ类整合子阳性菌株的耐药率高于整合子阴性的菌株。结论Ⅰ类整合子及整合子相关基因盒在产ESBLs大肠埃希菌株中分布广泛,整合子在细菌耐药中发挥作用。     

临床产ESBLs大肠埃希菌整合子检测及基因型分析

了解临床分离的产ESBLs大肠埃希菌(ESBLs-producing Escherichia coli,ESBL-EC)中Ⅰ、Ⅱ、Ⅲ类整合子及ESBL-EC基因型的分布。收集2014年1月至12月某三甲医院住院患者分离的大肠埃希菌,经全自动细菌分析系统鉴定并检测其对临床常用抗菌药物的耐药性,双纸片协同试验确定ESBL-EC,采用聚合酶链反应(PCR)对整合子基因和ESBLs基因进行检测。K-B法比较整合子阳性菌株与阴性菌株的耐药率。结果发现,98株临床非重复ESBL-EC对青霉素类、头孢菌素类、喹诺酮类、单环酰胺类和庆大霉素耐药率均大于50%,对妥布霉素耐药率为31.62%,对呋喃妥因、阿莫西林/克拉维酸和阿米卡星较敏感,分别为11.11%、13.4%和6.12%;对碳青霉烯类抗菌素、替加环素和哌拉西林/他唑巴坦敏感率为100%。98株菌中检出47株含Ⅰ类整合子(47.96%),3株含Ⅱ类整合子(3.06%),所有菌株中有1株同时含Ⅰ类和Ⅱ类整合子,未检出Ⅲ类整合子。整合子阳性菌株对四环素和复方新诺明的耐药率高于整合子阴性菌株(P0.05)。98株菌中β-内酰胺酶基因TEM、CTX-M-9、CTX-M-1、CTX-M-2和SHV阳性率分别为62.24%、53.06%、32.65%、4.08%和3.06%,ESBL-EC基因分型分布以TEM合并CTX-M-9型(共30株)最多见,占30.61%。结果表明,Ⅰ类整合子在产ESBLs大肠埃希菌中分布广泛,本研究尚不足以证明整合子的存在可影响ESBL-EC菌株抗生素耐药水平。同时携带TEM和CTX-M-9基因是安徽医科大学解放军174临床学院产ESBLs大肠埃希菌临床耐药菌株产生的主要原因。     

ESBLs大肠埃希菌和肺炎克雷伯杆菌整合子分布及其与ESBLs基因关系研究

目的研究整合子在院内感染ESBLs大肠埃希菌和肺炎克雷伯杆菌中的分布及其与细菌耐药的相关性,探讨Ⅰ类整合子在ESBLs基因水平转移中的作用及其与ESBLs基因型的关系。方法运用K-B法,纸片扩散法,PCR,接合传递试验、套式PCR、质粒谱分析及DNA测序研究携带耐药基因的Ⅰ类整合子与耐药播散的关系。结果产ESBLs和非产ESBLs菌株中Ⅰ类整合酶扩增阳性例数分别是70例（占66.7%）和22例（占22.4%）,2组整合子阳性率差异有统计学意义（P〈0.01）。整合子阳性组中ESBLs、多重耐药菌均明显高于阴性组。产ESBLs菌株中blaSHV、blaCTX和blaTEM基因与整合子经质粒共同转移的频率分别是79.3%、58.5%和45.1%。结论整合子在产ESBLs大肠埃希菌和肺炎克雷伯杆菌中的分布明显高于非产ESBLs菌株,主要为Ⅰ类整合子,并参与产ESBLs菌株多重耐药性的形成,其中blaSHV基因型与整合子经质粒共同转移的频率高于其他两类基因,提示SHV型酶在浙南地区具有更强的扩散优势。     

泌尿道感染大肠埃希菌耐药性分析

目的了解医院泌尿道感染大肠埃希菌产ESBLs的发生率和耐药情况。方法据NCCLS文件,产ESBLs细菌的检测用双纸片确认法。抗生素敏感测定采用K—B琼脂扩散法。结果在检出的176株大肠埃希菌中．ESBLs阳性的菌株有53例,ESBLs检出率为30．1％。检出的产ESBLs细菌对所有青霉素类抗生素产生耐药,产ESBLs对3代头孢类抗生紊头孢曲松、头孢噻肟的耐药率〉81％,对磺胺类、喹诺酮类也在71％以上。对亚胺培南全部敏感。结论医院泌尿道大肠埃希菌产ESBLs发生率较高,且对多种抗生素呈交叉耐药和多重耐药．应做好常规检测和监测．指导临床合理使用抗生素。     

整合子介导的肠致病性大肠埃希菌多重耐药研究

目的了解肠致病性大肠埃希菌（EPEC）多重耐药菌株中整合酶基因的携带情况,研究整合子与抗生素多重耐药的相关性。方法使用血清学的方法对EPEC进行初筛,用PCR扩增EPEC毒力基因（eae,EAF,bfpA）进行确证。对确证为EPEC的细菌DNA进行提取,使用PCR方法对整合酶基因及在整合子中插入的基因盒进行扩增。EPEC药敏试验采用K-B琼脂扩散法。结果在34株EPEC中,ESBL为14株,其中在lI株ESBL阳性细菌中扩增出整合子I整合酶片段,在20株ESBL阴性细菌中,有7株扩增出相应的片段。在这所有的34株细菌中未检出整合子Ⅱ和Ⅲ。结论I类整合子在肠致病性大肠埃希菌多重耐药菌株中最常见,是导致细菌多重耐药的一个重要因素,合理用药,控制耐药基因的传播是当前医学面临的一个重要问题     

外科手术切口感染大肠埃希菌的耐药性分析

目的分析外科手术切口中大肠埃希菌的感染率及耐药性,为临床合理用药提供科学依据。方法对2009年6月至2010年9月外科患者手术切口标本中分离的大肠埃希菌耐药情况进行分析,并对产超广谱β-内酰胺酶（ESBLs）菌株进行表型确证试验。结果 140株大肠埃希菌中,产ESBLs菌株的检出率为42.14%;产ESBLs菌株对一～四代头孢菌素、广谱青霉素、氟哇诺酮类及氨基糖苷类抗菌药物具有较高的耐药率;非产ESBLs菌株对青霉素类以外的抗菌药物具有较高的敏感率;产ESBLs菌株对大多数抗菌药物的耐药率明显高于非产ESBLs菌株（P〈0.05）。结论外科手术切口术后大肠埃希菌的感染较严重,且产ESBLs菌株多药耐药明显,临床应加强监测与控制。     

肿瘤患者医院感染产ESBLS大肠埃希菌调查及基因分析

目的探讨肿瘤患者医院感染产超广谱β-内酰胺酶(extended spectrum beta-lactamase,ESBLs)大肠埃希菌的临床特点,并分析其基因型特征。方法对2014年3月~2016年10月我院肿瘤患者498株临床标本中分离出的338株产ESBLs大肠埃希菌的临床分布进行分析,并比较产与非产ESBLs菌株对19种抗菌药物的耐药率,同时随机抽取338株中的60株应用聚合酶链反应(polymerase chain reaction,PCR)分别扩增ESBLs基因(TEM、CTX-M和SHV)型别。结果 498株大肠埃希菌中产ESBLs 338株(67.87%),标本类型以尿液标本为多(101株,29.88%),临床科室分布中妇瘤科最多(94株,27.81%)。产与非产ESBLs大肠埃希菌对亚胺培南及美罗培南耐敏感性高(耐药率为零),而产ESBLs大肠埃希菌的耐药率大多数均高于非产ESBLs大肠埃希菌,差异均有统计学意义(P0.05)。PCR扩增后基因测序显示,55株为TEM(91.67%),42株为CTX-M1(70.00%),42株为CTX-M13(70.00%),24株为SHV(40.00%),其中携带2种或2种以上耐药基因的有53株(88.33%)。结论本研究肿瘤患者医院感染产ESBLs大肠埃希菌主要标本以尿液标本为主,重点分布在妇瘤科;基因型以TEM、CTX-M1、CTX-M13型为主,并呈现多重耐药趋势。     

社区与医院感染中肺炎克雷伯杆菌和大肠埃希菌耐药性分析

目的探讨社区和医院感染中肺炎克雷伯杆菌和大肠埃希菌产ESBLs的情况及耐药特性。方法采用体外扩散确证试验检测ESBLs,同时用Micro scan wat RA way-40系统全自动细菌鉴定/药敏分析仪及K-B琼脂扩散法进行细菌鉴定和体外药敏试验。结果社区感染标本中分离出肺炎克雷伯杆菌79株,产ESBLs20株,阳性率为25.3%,大肠埃希菌177株,产ESBLs27株,阳性率为15.3%;医院感染标本中分离出肺炎克雷伯杆菌82株,产ES-BLs33株,阳性率为40.2%,大肠埃希菌135株,产ESBLs42株,阳性率为31.1%,社区与医院感染菌株产ESBLs比较差异均有统计学意义(P均<0.05);ESBLs阳性菌株对多种抗生素耐药,其耐药性明显高于ESBLs阴性菌株。结论肺炎克雷伯杆菌和大肠埃希菌产ESBLs菌株在临床分离率较高,医院感染标本要显著高于社区感染标本,并且对多种抗生素具有高度耐药性,产ESBLs菌株耐药性显著高于不产ESBLs菌株,临床上应加强对ESBLs的控制,以防感染流行。     

Distribution of class 1 integrons among enteropathogenic Escherichia coli

The aim of this study was to investigate the incidence of and resistance gene content of class 1 integrons among enteropathogenic Escherichia coli (EPEC) and non-EPEC and to investigate intraspecies genetic diversity of EPEC strains isolated from children with diarrhea in Iran. Twenty-eight EPEC and 16 non-EPEC strains isolated from children with diarrhea were tested for the presence of a class 1 integron associated integrase gene (int1). Sequence analysis was performed to identify the resistance gene content of integrons. Genetic diversity and cluster analysis of EPEC isolates were also investigated using enterobacterial repetitive intergenic concensus-polymerase chain reaction (ERIC-PCR) fingerprinting. Twenty-three (82%) EPEC isolates and 11 (68.7%) non-EPEC isolates harbored the int1 gene specific to the conserved integrase region of class 1 integrons. Sequence analysis revealed the dominance of dfrA and aadA gene cassettes among the isolates of both groups. ERIC-PCR fingerprinting of EPEC isolates revealed a high diversity among these isolates. The widespread distribution of 2 resistance gene families (dfrA and aadA) among both groups of EPEC and non-EPEC isolates indicates the significance of integrons in antibiotic resistance transfer among these bacteria. Furthermore, clonal diversity of EPEC isolates harbouring a class 1 integron also suggests the circulation of these mobile elements among a diverse population of EPEC in this country.     

Molecular characterization of antibiotic resistant Escherichia coli strains isolated from tap and spring waters in a coastal region in Turkey

A hundred and seventeen antibiotic-resistant Escherichia coli strains were isolated from public tap and spring waters which were polluted by fecal coliforms. There were no significant differences between two water sources as to the coliform pollution level (p> 0.05). All E. coli isolates were detected to be resistant to one or more antibiotics tested. Nearly 42% of the isolates showed multiresistant phenotype. Three (2.5%) of these isolates contained class 1 integron. Sequencing analysis of variable regions of the class 1 integrons showed two gene cassette arrays, dfr1-aadA1 and dhfrA17-aadA5. Resistance to ampicillin, tetracycline or trimethoprim-sulfamethoxazole was transferable according to the results of conjugation experiments. The rate of tetracycline resistance was 15%. tet(A)-mediated tetracycline resistance was widespread among tetracycline-resistant E. coli isolates. Genotyping by BOX-polymerase chain reaction (BOX-PCR) showed that some of the strains were epidemiologically related. This is the first report on the prevalence and characterization of class 1 integron-containing E. coli isolates of environmental origin in Turkey.     

上呼吸道分离鲍曼不动杆菌1 型整合子的分离与鉴定

【目的】研究I型整合子的结构特征,探讨其与细菌多重耐药之间的相关性。【方法】收集2008年至2009年广州呼吸疾病研究所上呼吸道分离的187株鲍曼不动杆菌,应用K-B纸片扩散法检测耐药性,采用聚合酶链式反应进行I型整合子整合酶基因的检测;扩增整合子的可变区,应用DNA测序技术分析I型整合子基因结构。【结果】I型整合子的阳性率达53.4%。共七种1型整合子基因盒被鉴定,其中首次发现报道一种新的整合子(GenBank:HQ322622)。可变区主要编码氨基糖苷类药物的耐药基因。20种抗菌素耐药的结果均表明携带Ⅰ型整合子的鲍曼不动杆菌耐药率较不携带I型整合子的鲍曼不动杆菌的耐药率明显增高。整合子与鲍曼不动杆菌的多重耐药表型具有密切相关性。【结论】I类整合子相关耐药基因在本院临床分离鲍曼不动杆菌中分布较广泛。整合子在鲍曼不动杆菌耐药性的形成和播散中具有重要作用。     

重症监护病房产ESBLs菌的基因分型

目的了解深圳市人民医院重症监护病房分离菌超广谱β-内酰胺酶（ESBLs）的检出率及其基因型分布情况。方法收集来自重症监护病房大肠埃希菌和肺炎克雷伯菌分离株48株,采用CLSI推荐的表型确证方法筛选出ESBLs株,并利用PCR及DNA测序法分析产酶菌株的ESBL基因型。结果（1）48分离株菌中共检出产ESBLs菌24株,阳性率为50.0%。（2）产酶菌中93.8%（15/16）的大肠埃希菌和87.5%（7/8）的肺炎克雷伯菌分别检出CTX-M基因;其中72.7%（16/22）为CTX-M-14。6株肺炎克雷伯菌检出SHV基因,其中3株为SHV-11型,另3株为SHV-12型,6株含SHV基因的肺炎克雷伯菌中5株合并CTX-M基因。而所有大肠埃希菌株均未检出SHV基因。所有产酶菌中,分别有10株大肠埃希菌和2株肺炎克雷伯菌检出TEM-1基因,其中1株大肠埃希菌只检出TEM-1基因,未检出SHV型或CTX-M型基因。结论重症监护病房分离菌ESBLs检出率高,以CTX-M-14为主要基因型。     

Phylogenetic Distribution and Prevalence of Genes Encoding Class I Integrons and CTX-M-15 Extended-Spectrum β-Lactamases in Escherichia coli Isolates from Healthy Humans in Chandigarh,India

Escherichia coli is generally considered as a commensal inhabitant of gastrointestinal tract of humans and animals. The aim of this study was to gain insight on the distribution of phylotypes and presence of genes encoding integrons, extended β-lactamases and resistance to other antimicrobials in the commensal E. coli isolates from healthy adults in Chandigarh, India. PCR and DNA sequencing were used for phylogenetic classification, detections of integrase genes, gene cassettes within the integron and extended β-lactamases. The genetic structure of E. coli revealed a non-uniform distribution of isolates among the seven phylogenetic groups with significant representation of group A. Integron-encoded integrases were detected in 25 isolates with class 1 integron-encoded intI1 integrase being in the majority (22 isolates). The gene cassettes identified were those for trimethoprim, streptomycin, spectinomycin and streptothricin. The dfrA12-orfF-aadA2 was the most commonly found gene cassette in intI1 positive isolates. Phenotypic assay for screening the potential ESBL producers suggested 16 isolates to be ESBL producers. PCR detection using gene-specific primers showed that 15 out of these 16 ESBL-producing E. coli harboured the bla _CTX-M-15 gene. Furthermore, molecular studies helped in characterizing the genes responsible for tetracycline, chloramphenicol and sulphonamides resistance. Collectively, our study outlines the intra-species phylogenetic structure and highlights the prevalence of class 1 integron and bla _CTX-M-15 in commensal E. coli isolates of healthy adults in Chandigarh, India. Our findings further reinforce the relevance of commensal E. coli strains on the growing burden of antimicrobial resistance.     

I类整合子与产ESBLs肺炎克雷伯菌多重耐药关系的研究

目的了解产ESBLs肺炎克雷伯菌的整合子存在状况。方法用PCR方法扩增Ⅰ类整合酶基因,经电泳后检测扩增产物。结果72株产ESBLs肺炎克雷伯菌中检测出Ⅰ类整合子67株,检出率为93．0％,Ⅰ类整合子阳性菌对氨基糖苷类、喹诺酮类及头孢菌素类药物表现出较高的耐药,其多重耐药率明显高于Ⅰ类整合子阴性菌株（P〈0．05）。结论Ⅰ类整合子广泛地存在产ESBLs肺炎克雷伯菌中,Ⅰ类整合子对细菌多重耐药性的产生和传播起着重要作用。     

Analysis of integrons in clinical isolates of Escherichia coli in China during the last six years

A multiple PCR for the detection of the integrase genes of the three classes of integrons was carried out, and their gene cassettes were characterized in 111 clinical strains of Escherichia coli isolated in Guangzhou City, China during the last 6 years. IntI1 and intI2 genes were detected in 95 isolates (85.6%) and four isolates (3.6%), respectively. No intI3 gene was detected. Six different gene cassettes were found in these strains, and a high prevalence of dfr and aad genes was observed. The E. coli isolates that contained a 1664-bp amplicon of dfrA17-aadA5 in class 1 integron were found to be phylogenetically unrelated to each other by using the enterobacterial repetitive intergenic consensus PCR, as the cassette could be transferred to recipient strains, indicating that the gene cassettes might be disseminated in the clinical strains by a horizontal gene transfer. Therefore, it is important that guidelines for the prudent use of antimicrobial agents are adopted and surveillance programs are established.     

ICU病区大肠埃希菌中qnr基因的分布与耐药性的研究

目的调查温州医科大学附属第一医院ICU病区分离的大肠埃希菌基因的分布以及与耐药谱的关系,并初步探讨其在分子流行病学中的作用。方法收集2012年1-9月ICU病区分离的大肠埃希菌76株进行qnr基因检测,并通过DNA直接测序确定;分析qnr基因在ICU病区分离的大肠埃希菌的分布及其与耐药性的关系。结果根据PCR产物片段大小及测序分析,76株大肠埃希菌中共有qm基因阳性菌株46株,阳性率为60. 5% ;对阳性菌株进行DNA测序、BLAST比对,其中25株为qnrB基因,17株为qnrS基因阳性,12株基因阳性,未检测到qwC和qnrD基因。在46株qnr基因阳性菌株中有38株为产ESBL菌株,而在qnr阴性菌株中仅有5株ESBL阳性。结论该院ICU分离大肠埃希菌qnr基因携带严重,呈现出多重耐药性,多伴随呈现为产ESBL菌株。