2016年药学论文—阴沟肠杆菌质粒介导耐喹诺酮类药物的机制研究.docVIP

精品文档欢迎来主页查询 更多精品文档,欢迎来我主页查询 阴沟肠杆菌质粒介导耐喹诺酮类药物的机制研究 【摘要】 目的 了解深圳地区阴沟肠杆菌中qnrA基因的流行情况、基因定位及其介导喹诺酮耐药性产生的机制。方法 收集临床分离的阴沟肠杆菌45株，采用PCR法结合测序的技术筛查qnrA基因，大质粒提取技术、Southern杂交和接合传递试验进行质粒定位，琼脂对倍稀释法进行药物敏感性检测。结果 45株阴沟肠杆菌中，7株PCR法及测序证实为qnrA基因。7株菌中6株菌所携质粒被成功提取并进行Southern杂交，qnrA基因定位于80～200kb大小的低拷贝数天然质粒上。4株菌成功进行接合传递试验，使接合子对环丙沙星的MIC值提高了32～64倍。结论 质粒介导的喹诺酮耐药基因qnrA在深圳地区阴沟肠杆菌中具有较高的流行率，可能是导致阴沟肠杆菌对喹诺酮类抗菌药获得性耐药的重要原因。 【关键词】 qnrA基因 阴沟肠杆菌 喹诺酮 耐药 质粒 Plasmidmediated quinolone resistance in clinical isolates ABSTRACT Objective To investigate the prevalence and location of qnrA gene in clinical isolates of Enterobacter cloacae and its function in the development of quinolone resistance. Method Fortyfive Enterobacter cloacae were screened for the qnrA gene by PCR and then conformed with direct sequencing. Large plasmid extraction and Southern hybridization were done to determine the location of qnrA. Conjugation experiments were done to determine the transferability and its effects in the development of quinolone resistance. Results qnrA were detected in 7 (15.6%) of 45 strains. The qnrA sequence matched that of NCBI GenBank with 100%. Plasmid extraction and Southern hybridization were succeed in 6 stains. The qnrA were located in plasmids in size of 80～200kb. Conjugation experiments were done in 4 strains. Transconjugants had 32～64fold increases in the MIC of ciprofloxacin relative that of the recipient but were still susceptible to ciprofloxacin according to CLSI stardard. Conclusion Transferable plasmidmediated quinolone resistance associated with qnrA were high prevalent in clinical isolates of Enterobacter cloacae from Shenzhen city and may contribute to the quinolone resistance of thiese bacteria. KEY WORDS qnrA; Enterobacter cloacae; Quinolone; Drug resistance; Plasmid 1998年，Martinez等首次证实肺炎克雷伯菌对喹诺酮类抗菌药存在质粒介导的耐药机制，并将该耐药基因命名为qnr(现命名为qnrA)［1］。之后我国学者王明贵等研究发现qnrA基因在耐喹诺酮类抗菌药大肠埃希菌中的检出率高达7.7%，提示qnrA基因在介导肠杆菌科细菌耐药机制方面可能发挥着重要作用，引起广泛关注［2］。 目前阴沟肠杆菌已成为医院内感染的重要病原菌，且对喹诺酮类抗菌药的耐药性迅速上升，给临床抗感染治疗带来严峻挑战［3，4