肺炎克雷伯菌多粘菌素耐药基因mcr-8的水平传播及适应性机制

多粘菌素是治疗多重耐药细菌感染的最后一道防线，而质粒介导多粘菌素耐药基因(mcr)的发现，给全球公共健康带来巨大挑战。申请人前期研究结果表明肺炎克雷伯菌是mcr-8的主要宿主，且高毒力环境源mcr-8阳性肺炎克雷伯菌对公共健康存在威胁。但目前的研究局限于流行病学的筛查，对mcr-8传播机制及其与肺炎克雷伯菌的适应性关系尚不清楚。本项目拟采用分子生物学技术、高通量测序和生物信息学分析工具等研究方法，分析携带mcr-8肺炎克雷伯菌的分子特征和基因侧翼环境；采用质粒荧光标记、接合转移法、转录测序等方法揭示mcr-8水平传播效率及分子机制；通过克隆载体构建和细菌竞争模型，解析mcr-8基因与宿主菌在进化过程中的适应性变化及其潜在分子机制。本研究成果为更好的理解和应对细菌耐药性问题提供理论基础，为制定应对多粘菌素耐药策略提供科学依据。

Colistin is considered as a “last line of defense” against multidrug-resistant bacterial infection, however, the discovery of mobile colistin resistance genes (mcr), has become a significant challenge to global public health. Our preliminary results show that Klebsiella pneumoniae is the main bacterial host for mcr-8 gene and highly virulent mcr-8-positive K. pneumoniae strains(MCRPKP) were identified from environment samples, posing a great public health concern. However, current studies are limited to epidemiological screening for mcr-8, but the transmission mechanism of mcr-8-carrying plasmids and its fitness cost to K. pneumoniae remain unclear. Therefore, we analyse the genetic characteristics of MCRPKP strains and genetic environment surrounding to mcr-8 gene by applying high-throughput sequencing and bioinformatics analysis tools. We will use florescent-labelled plasmid, conjugation experiments and transcriptomics, to investigate the efficiency and mechanisms of horizontal transfer of mcr-8 plasmids. In addition, the changes of MCRPKP fitness and its potential mechanism during evolution experiments are explored via applying a series of experimental-modelling systems including molecular cloning technique and bacterial competition model. Taken together, the proposed project will provide theoretical basis in understanding and tackling the problems causes by antibiotic resistance, and also provide scientific evidence for developing better strategies against colistin resistance.