【作者】 吴春阳；

【导师】 顾国浩；

【作者基本信息】 苏州大学 ， 临床检验诊断学（专业学位）， 2013， 硕士

【摘要】 目的探讨头孢哌酮-舒巴坦等13种抗菌药物对鲍曼不动杆菌(Acinetobacterbaumannii,AB)的体外抗菌活性，为临床合理用药提供实验依据；了解AB主动外排系统和外膜蛋白基因的表达变化特点，分析其在介导细菌泛耐药中的作用。方法收集苏州大学附属第一医院微生物室分离自临床感染患者的100株非重复AB菌株，全部经法国生物梅里埃公司的VITEKⅡCompact和手工试验重新鉴定确定为AB。第一部分采用K-B法和琼脂二倍稀释法对100株AB进行13种抗菌药物体外抗菌活性的检测，观察AB的耐药状况并分析该组菌的耐药模式。第二部分PCR扩增鲍曼不动杆菌中主动外排系统（AdeABC、AdeIJK）和外膜蛋白（CarO）的基因，通过琼脂糖凝胶电泳观察结果，初步探讨鲍曼不动杆菌临床分离株中外排泵的分布情况；实时荧光定量RT-PCR检测外排泵（AdeABC、AdeIJK）基因的mRNA表达水平，了解外排泵基因的表达情况，分析耐药株和敏感株在基因表达方面的差异。结果第一部分临床常用的抗生素（如头孢他啶、哌拉西林、丁胺卡那、环丙沙星等）作用均较差，米诺环素、多黏菌素B和头孢哌酮-舒巴坦对AB有相对较好的抗菌活性，亚胺培南的作用相对差一些；100株AB中有75株泛耐药株，MIC测定的结果显示米诺环素、多粘菌素B和头孢哌酮-舒巴坦的耐药率分别为88%、59.15%、0%和21.13%，MIC值的几何均数分别为25.16（耐药）、5.85（中介）、1.06（敏感）和29.3（中介）。第二部分敏感和泛耐药两组间adeB基因的PCR及RT-PCR检测结果比较均有统计学意义，而adeJ基因RT-PCR检测两组间无显著差异；98.57%的泛耐药菌株检测到carO基因。结论AB临床分离株的耐药情况严重；米诺环素、多黏菌素B、头孢哌酮-舒巴坦对临床分离的泛耐药AB具有良好的抗菌活性，可作为治疗选择药物，而碳青霉烯类抗生素亚胺培南的抗菌作用较差；AB临床分离株adeB和adeJ基因检出率较高，且adeB基因在AB泛耐药株和少部分敏感株中表达量明显增加；AdeABC、AdeIJK主动外排系统及其基因表达量的增加在AB泛耐药性的形成中具有一定的作用。更多还原

【Abstract】 Objective To determine the in vitro activities of13antimicrobials includingCefoperazone-sulbactam against clinical isolates of Acinetobacter baumannii and provideexperimental evidences for rational use of antimicrobial agents. To investigate the functionthat induce pan-drug resistance and gene expression of active efflux pump and outermembrane protein in Acinetobacter baumannii.Methods A total of100non-repeated strains of Acinetobacter baumannii wereisolated from clinical specimens in The First Affiliated Hospital of Suchoow Universityand all of them had been re-identified by manual test and VITEK ⅡCompact ofBio-Merieux.First part The susceptibilities of100strains clinical Acinetobacter baumannii wasdetermined by K-B and2-fold agar dilution method followed by NCCLS recommendations．Second part The genes adeB、adeJ and carO were amplified by PCR and the resultswere observed by agarose gel electrophoresis．Reverse transcriptase polymerase chainreaction （RT-PCR） was used to measure the mRNA expression levels of active effluxpump AdeABC、AdeIJK.Results First part The antibacterial activity of antibiotic frequently-used is poorerthan minocyline、polymixin B and sulperazone. There are75pan-drug resistant strains inall100strains, their resistant rate to Imipenem、minocyline、polymixin B and sulperazoneare88%、59.15%、0%and21.13%while the geometrical mean of MICs are25.16（resistance）、5.85（medium）、1.06（sensitive）和29.3（medium）.Second part The test result of adeB gene including PCR and RT-PCR reveal thedifferences between resistant strains and sensitive strains have statistical significant whilethe expression level of adeJ gene between the two groups have no significant difference; The carO gene was detected in98.57%pan-drug resistant strains.Conclusions The antibiotic resistance is very serious while minocyline、polymixin Band sulperazone with considerable antibacterial activities may be the choices formulti-drug resistant isolates infections of Acinetobacter baumannii．The positive rate ofadeB and adeJ gene is high in Acinetobacter baumannii. Besides, the expression level ofadeB gene obviously increased in pan-drug resistant strains and some sensitive strains.Theactive efflux pump AdeABC play a role in pan-drug resistance of Acinetobacter baumannii更多还原