Molecular mechanisms of lytic bacteriophage infection of enterococci

项目总结 近几十年来，有效的抗生素疗法迅速减少，而多药耐药的情况有所增加。 耐药性(MDR)细菌感染。一种常见的耐多药病原体是革兰氏阳性肠道细菌 对多种抗生素耐药的粪肠球菌，包括诸如 万古霉素。耐多药肠球菌可在接受广谱抗生素治疗的患者的肠道内扩张 心理治疗。这种扩张可导致粪肠球菌移位到血液中，败血症，并进一步脱落。 从而使这些医院获得性感染永久化。对抗MDR的一个潜在战略 肠球菌是利用噬菌体(噬菌体)。噬菌体是感染和杀死细菌的病毒，具有高度的 专一性。噬菌体感染依赖于与细菌细胞表面的结合，将噬菌体DNA喷射到 细菌细胞，噬菌体基因组的复制，以及病毒颗粒从细胞中释放。利用噬菌体作为 治疗引发了类似于抗生素使用的担忧；细菌将对感染产生抵抗力，从而减少 治疗效果。因此，在噬菌体能够成为标准的临床治疗方法之前，我们必须清楚地了解 噬菌体感染细菌的具体机制以及细菌对噬菌体感染的反应。至 为了开始阐明噬菌体在感染过程中使用的机制，我用裂解噬菌体攻击粪肠球菌 发现肠球菌多糖抗原可用于对不同来源的E. 粪便菌株，包括噬菌体不能成功感染的菌株。除此之外，我还展示了 粪肠球菌中的移动质粒限制了噬菌体的感染。这个项目的目标是填补关键 我们对噬菌体-细菌相互作用的基本知识的差距，并提供对 影响噬菌体感染的转归，这将为噬菌体治疗的应用提供有益的认识。这 该项目包括以下两个具体目标：目标1：确定促进DNA进入的受体 依赖于EPA的噬菌体。这将通过补充生化分析来解决，以确定 促进噬菌体DNA进入粪肠球菌的受体。目标2：确定内源性疾病的遗传基础 限制肠球菌噬菌体感染的质粒。在这里，我将使用遗传方法来识别一种 编码在移动质粒上的新型肠球菌抗噬菌体限制机制。这些实验将 揭示肠球菌噬菌体识别和感染的新机制 最终帮助开发知情的噬菌体疗法。通过这样做，我的结果将扩展我们的基本 关于噬菌体感染期间肠球菌细胞生物学的知识，并可能确定新的目标 肠球菌的抗病治疗。

PROJECT SUMMARY In recent decades there has been a rapid decline in effective antibiotic therapies and an increase in multidrug- resistant (MDR) bacterial infections. One common MDR pathogen is the Gram-positive intestinal bacterium Enterococcus faecalis which resists many antibiotics including “last-line-of-defense” drugs such as vancomycin. MDR enterococci can expand in the intestine in individuals undergoing broad-spectrum antibiotic therapy. This expansion can lead to E. faecalis translocation to the bloodstream, sepsis, and further shedding of the bacterium, thus perpetuating these hospital-acquired infections. One potential strategy to combat MDR enterococci is the use of bacteriophages (phages). Phages are viruses that infect and kill bacteria with high specificity. Phage infection relies on binding to the bacterial cell surface, ejection of phage DNA into the bacterial cell, replication of the phage genome, and viral particle release from the cell. The use of phages as therapeutics raises concern similar to antibiotic use; bacteria will become resistant to infection, diminishing therapeutic efficacy. Thus, before phages can become a standard clinical therapy, we must clearly understand the specific mechanisms used by phages to infect bacteria and how bacteria respond to phage infection. To begin to elucidate the mechanisms used by phages during infection, I challenged E. faecalis with lytic phages and found that the enterococcal polysaccharide antigen is used for initial phage adsorption to various E. faecalis strains, including strains that the phage cannot successfully infect. Along with this, I have shown that the presence of a mobile plasmid in E. faecalis restricts phage infection. The goals of this project are to fill key gaps in our basic knowledge of phage-bacteria interactions and to provide insights into the mechanisms that influence the outcomes of phage infection, which will be beneficial knowledge for applied phage therapies. This project encompasses the following two Specific Aims: Aim 1: Identify the receptor that promotes DNA entry of Epa-dependent phages. This will be addressed through complementary biochemical assays to identify the receptor that promotes phage DNA entry into E. faecalis. Aim 2: Define the genetic basis of endogenous plasmids in restricting enterococcal phage infection. Here, I will use genetic approaches to identify a novel, enterococcal anti-phage restriction mechanism encoded on a mobile plasmid. These experiments will reveal new insights into the mechanisms that dictate enterococcal phage recognition and infection that will ultimately aid in the development of informed phage therapies. In doing so, my results will expand our basic knowledge surrounding enterococcal cell biology during phage infection and potentially identify new targets for anti-enterococcal therapies.