脑膜炎奈瑟菌CC4821耐药优势克隆的形成机制与分子进化

Neisseria meningitides (Nm) is the pathogen of epidemic cerebrospinal meningitis. China is a country with a traditionally high incidence of invasive meningococcal disease (MD) due to Nm. Since 2003, serogroup C clonal complex (CC4821) isolates has attributed to multiple MD outbreaks in Anhui and the other provinces in China. The research group have collected Nm isolates in 1965-2013 in Shanghai. We found that the serogroup C/B Nm (MenC/B) clone (ChinaCC4821-R1-C/B) is the main epidemic clone with resistant to fluoroquinolones, genetically close to CC4821 isolates in the early stage in Shanghai of different phylogenetic group. What important changes happened in these early stage CC4821 strains which help to generate the predominant resistance clone? Is it because of the acquisition or variation of genes related to colonization, invasiveness, virulence, antigen proteins? What’s the genetic relationship between CC4821 isolates from China and those from abroad? The study will focus on the CC4821 isolates of the early stage and current stage, explore the key variation of genes and mechanism of the generation of predominant resistance clone which was highly prevalent in China by phylogenetic analysis, comparative genomics and recombination analysis. It will pose theoretical and practical significance for the vaccination, targeted therapy and surveillance and prevention strategies of meningococcal disease caused by new invasive Nm.

脑膜炎奈瑟菌是引起流脑的病原体。我国是流脑高发国家。自2003年起，C群CC4821克隆造成安徽等中国多地流脑暴发流行。申请人项目组前期收集上海地区1965-2013年的脑膜炎奈瑟菌，发现近年我国主要流行克隆为C/B群CC4821喹诺酮类耐药优势克隆（ChinaCC4821-R1-C/B），与上海早期CC4821菌株遗传关系接近，但属不同分组。早期CC4821菌株在进化过程中发生哪些关键变化形成耐药优势克隆？是否与定植、侵袭、毒力、抗原相关蛋白等基因的获得或变异相关？与最近国际上出现的CC4821菌株的遗传关系是怎样的？本项目拟对早期CC4821菌株与近年耐药优势克隆进行分组研究，通过系统进化分析、比较基因组学、重组分析等方法寻找关键变化基因，明确耐药优势克隆的形成机制，揭示其在我国广泛流行的内在原因，为防控脑膜炎奈瑟菌新毒株的播散流行、研发针对本地区流行株的疫苗及药物靶位提供参考依据。

ChinaCC4821-R1-C/B是近年引起我国流脑的主要优势克隆，其形成机制不清。项目对本地及国内外早期与当代CC4821菌株进行遗传进化研究，将CC4821菌株分为4个亚系，ChinaCC4821-R1-C/B耐药克隆属于L44.1亚系，均为当代分离株，侵袭性菌株比例显著高于其它亚系，具有独特的抗原、毒力与耐药分子特征。中国早年菌株与当代W群携带株构成L44.4亚系。国外CC4821菌株多聚于L44.3，构成独特的Europe-USA簇，对青霉素非敏感（PenNS）。发现126个水平转移（HGT）事件可能促使L44.1出现。L44.1中有299个独特的基因座（loci）与等位基因，近一半与代谢旁路相关，可影响脑膜炎球菌的致病性和毒力。这些独特的loci可能来自中国早期不同种系，如C群ST-9514簇的菌株，A群CC5和CC1脑膜炎菌株等。优势克隆的出现可能与这些独特的等位基因积聚有关，并造成其在系统发育树上与其它亚系分离。.发现近年本地PenNS脑膜炎球菌上升迅速，首次报道CC4821克隆近期获得对青霉素和头孢噻肟的耐药性，与水平获得编码PBP2关键氨基酸突变的penA基因片段有关。PBP2蛋白A311V和T483S突变导致对头孢噻肟耐药。此外，发现本地1株PenNS脑膜炎球菌携带新penA型—penA832，该菌是欧洲Y群CC23优势克隆的近亲，从本地乳糖奈瑟菌获取penA832而产生耐药。本地共栖奈瑟菌已成为脑膜炎球菌获取耐药基因的重要储库。.研究表明ChinaCC4821-R1-C/B优势克隆已向海外扩张，并不断进化获得新的耐药突变，尤为值得关注。绝大部分PenNS脑膜炎球菌未被我国目前常规计划免疫覆盖，现有疫苗对预防B群CC4821侵袭株的潜在效果不确定，目前仍亟需对脑膜炎球菌流行株变异的监测，研发针对我国流行株的疫苗产品，防控新毒株的播散流行。