【摘要】 目的检测临床分离的肺炎克雷伯菌在体外形成生物被膜后产超广谱β-内酰胺酶(Extended-Spec-trumβ-Laetamases,ESBLs)的情况,分析及研究其耐药性和耐药基因的分型情况。方法采用改良平板法在体外建立肺炎克雷伯菌生物被膜模型,用三维试验确认产ESBLs菌株,用K-B法进行药敏试验,用聚合酶链反应(Polymer-ase chain reaction,PCR)进行blaSHV、blaTEM和blaCTX-M基因扩增,产物分别克隆入pMD18-T载体后测定其核苷酸序列,分析其基因亚型。结果临床筛选出的60株ESBLs阴性肺炎克雷伯菌有46株在体外成功建立了生物被膜模型,并有9株产生了ESBLs表型。产酶后菌株的耐药性明显高于产酶前。PCR结果显示9株细菌均携带SHV基因,有4株同时携带TEM基因,没有检出携带CTX-M基因的菌株。9株细菌的SHV基因分别属于SHV-5、SHV-12和SHV-28亚型。4株携带TEM基因的细菌均为TEM-1亚型。结论生物被膜的形成能够诱导肺炎克雷伯菌产生ESBLs。本实验中检出的产ESBLs的基因型都是由SHV-1突变产生的。生物被膜的形成和产生ESBLs的协同作用是生物被膜肺炎克雷伯菌耐药性增强的主要原因之一。更多还原

【Abstract】 Objective To investigate the extended-spectrum β-lactamases(ESBLs) production in biofilm-forming Klebsiella pneumoniae and analyze the drug resistance and genotyping in that condition.Method The in vitro model of Klebsiella pneumoniae biofilm was established by modified plating method.ESBLs-producing strains were confirmed by three dimensional test.Antimicrobial susceptibility testing was performed using Kirby-Bauer method.The blaSHV,blaTEM and blaCTX-M genes of ESBLs were amplified by PCR,then sequenced and analyzed after cloned into pMD18-T vector respectively.Result 46 of 60 ESBLs negative Klebsiella pneumoniae strains formed biofilm.9 strains of ESBLs phenotype were identified,which were highly-resistant.Genotypes showed that all the 9 strains carried SHV gene,among which 4 strains carried TEM gene while none carried CTX-M gene.The 9 SHV genes belonged to SHV-5,SHV-12 and SHV-28 respectively,while all the 4 TEM genes belonged to TEM-1.Conclusion Klebsiella pneumoniae biofilm formation could induce ESBLs production.The subtypes of ESBLs in this study mutated from SHV-1.The synergetic effect of ESBLs and biofilm was one of the main reasons that Klebsiella pneumoniae was highly resistant to antibiotics.更多还原