肠杆菌科细菌多粘菌素耐药及传播机制研究

Enterobacteriaceae are important clinical pathogens. Multi-drug resistant (MDR) and extensively-drug resistant (XDR) Enterobacteriaceae, especially carbapenem-resistance Enterobacteriaceae (CRE) are dramatically increased, making colistin the last-resort antibiotic for treating infections caused by these resistant pathogens. However, colistin resistance has increased rapidly in recent years, leading to the plight of no drugs available for clinicians. Present studies, including our previous studies, suggest that colistin resistance is caused by the two-component systems of PmrA/B and PhoP/Q mediated by chromosome and plasmid-mediated gene mcr that regulates the modification of lipopolysaccharide (LPS). However, there are still a lot of colistin resistant strains that can not be explained by these existing mechanisms, and the resistance genes’ capacity of transmission and characteristics have not been elucidated. We are going to use filter-mating experiments, high-throughput whole genome sequencing, comparative genomics, genes knockout and complementation tests to clarify the mechanisms of colistin resistance and transfer of Enterobacteriaceae, the function of mcr gene in the formation of high-level resistance to colistin based on our previous studies. We hope to reveal the mechanisms of colistin resistance in Enterobacteriaceae systematacially, clarify the mechanisms and capacity of transfer of mcr gene as well as the function of mcr gene in the formation of high-level resistance to colistin in Enterobacteriaceae. It is expected to find new mechanisms of resistance and transfer and finally find effective prevention and control measures to colistin resistant Enterobacteriaceae.

肠杆菌科细菌是临床重要病原菌。多重耐药、广泛耐药肠杆菌科细菌迅速增加，尤其是碳青霉烯耐药肠杆菌科细菌的流行，使多粘菌素成为治疗该耐药菌感染的最后一道防线。近年来，多粘菌素耐药性迅速增加，使人类面临无药可用的困境。现有研究（包括本课题组前期研究）发现其对多粘菌素耐药主要由染色体介导的双组分调控系统PmrA/B和PhoP/Q和质粒介导的mcr基因调节脂多糖修饰所致。但仍有不少耐药株现有机制不能解释，其耐药基因传播能力和特征仍未阐明。本研究拟在原有研究基础上通过接合试验、高通量全基因组测序、比较基因组学等方法，系统研究多粘菌素耐药机制、耐药传播机制及携带mcr基因对多粘菌素高耐药形成的作用等。系统阐明我国肠杆菌科细菌对多粘菌素耐药的确切机制；明确多粘菌素耐药基因传播机制和传播能力，揭示mcr基因在多粘菌素高耐药菌株形成中的作用；极可能发现新的耐药和传播机制，为我国多粘菌素耐药防控提供科学依据。

多黏菌素是临床治疗多重耐药肠杆菌科细菌最重要的抗菌药物之一，但是近年来，多黏菌素耐药率有逐年缓慢上升的趋势，而其耐药机制仍未完全阐明。本研究通过高通量全基因组测序、比较基因组学、基因敲除与回补等方法，阐明了肠杆菌科细菌多黏菌素耐药机制，其中大肠埃希菌以质粒介导的多黏菌素耐药基因mcr-1基因为主，肺炎克雷伯菌和肠杆菌属细菌则以染色体介导的耐药机制为主，并且发现了可疑的肺炎克雷伯菌多黏菌素耐药相关新基因blaCDP和citG基因；通过接合实验、全基因组测序和分析等方法，明确携带mcr-1质粒可在细菌种内及种间进行水平传播，并进一步通过构建系统发育树和质粒序列分析等，阐明mcr-1基因和mcr-9基因主要通过质粒进行传播，而mcr-10基因则主要通过转座元件进行传播；同时，通过基因克隆、比较基因组学等方法，明确mcr-1基因对大肠埃希菌多黏菌素高水平耐药形成具有促进作用，而在肺炎克雷伯菌中没有类似作用。通过本研究，系统阐明了我国肠杆菌科细菌的多黏菌素耐药机制、mcr基因的传播机制及其在肠杆菌科细菌多黏菌素高水平耐药形成中所起的作用，为我国多黏菌素耐药肠杆菌科细菌防控提供切实有力的科学依据。