鲍曼不动杆菌替加环素耐药机制研究

发布时间：2018-06-28 03:50

  本文选题：鲍曼不动杆菌 + 替加环素　； 参考：《浙江大学》2015年博士论文

【摘要】：多重耐药鲍曼不动杆菌已成为临床抗感染治疗的难题,替加环素对其具有很好的抗菌活性。替加环素具有抗菌谱广、抗菌活性强、不良反应小,应用不受患者年龄、性别、病情的限制等优点,使其在治疗多重耐药鲍曼不动杆菌中倍受关注。然而,抗生素应用的同时往往伴随着细菌耐药的产生。自2005年美国FDA批准替加环素应用临床以来,2006年就分离到了替加环素敏感性降低的鲍曼不动杆菌菌株。目前越来越多的替加环素不敏感菌株或耐药菌株不断在临床主要病原菌中被分离。替加环素耐药或不敏感菌株中最常见的是鲍曼不动杆菌。本研究主要分三部分,逐步深入研究鲍曼不动杆菌替加环素耐药机制。 第一部分主要通过PCR、荧光定量PCR、外排泵抑制剂抑制表型检测、突变检测等方法分析了RND外排泵对多重耐药鲍曼不动杆菌替加环素耐药机制的影响,结果显示：在替加环素耐药和敏感菌株中,外排泵基因adeB、adeG和adeJ广泛存在。替加环素耐药菌株外排泵表达水平明显高于替加环素敏感菌株,主要表现为AdeABC高表达,是由双组分调节系统adeS中ISAba-1插入突变所致。具有外排泵抑制剂抑制表型菌株约占替加环素不敏感菌株(MIC≥4mg/L)的65.6%。 第二部分通过在鲍曼不动杆菌标准菌株ATCC17978中查找并克隆tetX基因的同源基因——A1S3356基因,在鲍曼不动杆菌中研究A1S3356基因对四环素类抗生素和替加环素耐药机制的影响。结果显示在A1S3356过表达菌株中未检测到四环素类抗生素和替加环素的敏感性发生明显变化,说明在鲍曼不动杆菌中A1S3356不介导四环素类抗生素和替加环素耐药。 第三部分,利用替加环素连续体外诱导试验,诱导鲍曼不动杆菌标准菌株ATCC19606,获得替加环素耐药菌株19606-T8；利用二代高通量测序技术进行全基因组测序,通过比较基因组学和比较转录组学,以及基因回补、功能验证试验等,力求探讨替加环素耐药新机制以及替加环素对鲍曼不动杆菌的影响。研究结果显示：trm基因缺失突变导致替加环素敏感性降低,野生型trm基因能够回复替加环素敏感性。因此,除RND外排泵外,trm基因作为新机制在替加环素耐药过程中发挥重要作用。同时在替加环素的作用下,鲍曼不动杆菌的一些耐药基因、外排泵相关基因、膜孔蛋白和转录因子等表达水平明显增高,以及代谢相关基因表达水平也发生改变等。
[Abstract]:Multidrug resistant Acinetobacter baumannii has become a difficult problem in clinical anti-infective therapy, and tegicycline has good antibacterial activity against it. Tegicycline has the advantages of wide antibacterial spectrum, strong antibacterial activity, small adverse reaction, and its application is not limited by age, sex and condition of the patient, so it has attracted much attention in the treatment of multidrug resistant Acinetobacter baumannii. However, antibiotic use is often accompanied by bacterial resistance. Acinetobacter baumannii strain with low sensitivity to tegicycline was isolated in 2006 since FDA approved the clinical application of tegicycline in 2005. At present, more and more tegacycline insensitive or resistant strains have been isolated from the main clinical pathogens. Acinetobacter baumannii is the most common strain of tegicycline-resistant or insensitive strains. This study was divided into three parts, and the mechanism of tegacycline resistance of Acinetobacter baumannii was studied step by step. In the first part, the effects of RND efflux pump on multidrug resistance of Acinetobacter baumannii were analyzed by means of PCR, fluorescence quantitative PCR, inhibition phenotype of efflux pump inhibitor, mutation detection and so on. The results showed that the efflux pump gene adeBnadeG and adeJ were widely present in tegacycline resistant and sensitive strains. The expression level of efflux pump in tegacycline resistant strains was significantly higher than that in tegacycline sensitive strains. The high expression of AdeABC was mainly caused by the insertion mutation of ISAba-1 in the two-component regulatory system (adeS). The strain with efflux pump inhibitors accounted for 65.6% of tegacin-insensitive strains (MIC 鈮，

本文编号：2076614