Genome defense and acquired antibiotic resistance in Enterococcus faecalis and Enterococcus faecium

 性状（由申请方提供）：粪肠球菌和E.屎肠球菌是正常的人肠道定植菌和机会致病菌，是美国医院获得性感染的主要原因。E. faecalis和E.粪便导致医院获得性感染。这些医院肠球菌是多重耐药的，相对于大肠杆菌肠球菌菌株具有显著扩展的基因组。基因组扩增是由于获得移动的遗传元件（MGE），例如质粒和噬菌体。有趣的是，医院肠球菌缺乏CRISPR-Cas系统。CRISPR-Cas是原核基因组防御系统，可作为过去MGE遭遇的分子记忆，并提供针对MGE的获得性免疫。这些结果表明，抗生素治疗无意中选择了肠球菌获得MGE的能力增强。指导这项研究的中心假设是，受损的基因组防御与对MGE的强选择相结合导致临床环境中流行的多重耐药、基因组扩增的肠球菌的出现。在由NIAID K22研究学者发展奖支持的初步工作中，我们证明了CRISPR-Cas系统在大肠杆菌中的作用。粪球菌是获得抗生素耐药性的障碍。该提案中的研究调查了在E医院发生的独特孤儿CRISPR基因座的功能。粪球菌菌株，称为CRISPR 2，其调节这些菌株的抗微生物应激反应（Aim 1）。我们已经观察到，获得抗生素抗性质粒的Escherosal E。粪肠球菌导致含质粒细胞中的分子记忆丧失（CRISPR缺失）。将研究质粒逃避宿主CRISPR-Cas防御并诱导分子记忆丧失的机制（目的2）。最后，我们调查的假设，医院肠球菌是功能失调的“自我”与非自我识别，即基因组修饰防御（目的3）。

 DESCRIPTION (provided by applicant): Enterococcus faecalis and E. faecium are normal human intestinal tract colonizers and opportunistic pathogens that are among the leading causes of hospital-acquired infections in the US. Specific phylogenetic lineages of E. faecalis and E. faecium cause hospital-acquired infections. These hospital enterococci are multidrug-resistant and possess dramatically expanded genomes relative to commensal enterococcal strains. Genome expansion is due to the acquisition of mobile genetic elements (MGEs) such as plasmids and phage. Interestingly, hospital enterococci lack CRISPR-Cas systems. CRISPR-Cas are prokaryotic genome defense systems that serve as molecular memories of past MGE encounters and that provide acquired immunity against MGEs. These results suggest that antibiotic treatment inadvertently selects for enterococci with enhanced abilities to acquire MGEs. The central hypothesis guiding this research is that compromised genome defense combined with strong selection for MGEs leads to the emergence of multidrug-resistant, genome- expanded enterococci endemic to clinical environments. In preliminary work supported by a NIAID K22 Research Scholar Development Award, we demonstrated that a CRISPR-Cas system in commensal E. faecalis acts as an impediment to the acquisition of antibiotic resistance. The research in this proposal investigates the function of a unique orphan CRISPR locus occurring in hospital E. faecalis strains, called CRISPR2, which regulates the antimicrobial stress response of those strains (Aim 1). We have observed that acquisition of an antibiotic resistance plasmid by commensal E. faecalis leads to molecular memory loss (CRISPR deletion) in plasmid-containing cells over time. Mechanisms by which the plasmid evades host CRISPR-Cas defense and induces molecular memory loss will be investigated (Aim 2). Finally, we investigate the hypothesis that hospital enterococci are dysfunctional for `self' versus non-self recognition; i.e. genome modification defense (Aim 3).