High-throughput Screenings to Identify Host Receptors for Staphylococcal Adhesins

描述（由申请方提供）：葡萄球菌引起的感染性疾病范围很广，从轻微的皮肤感染到严重的疾病，如败血症、肺炎和心内膜炎。虽然金黄色葡萄球菌是迄今为止最致命的菌种，但也发现表皮葡萄球菌和路邓葡萄球菌主要在免疫功能低下的个体中引起严重感染。过去，剧毒的S.金黄色葡萄球菌菌株局限于医院环境，主要影响免疫功能低下的患者或具有潜在共病的个体。不幸的是，类似的菌株最近出现在社区中，导致其他健康个体严重感染。表皮葡萄球菌和路邓葡萄球菌在较小程度上和病理学上略有不同，似乎遵循相同的模式。这些菌株中的大多数已经进化为对抗生素具有耐药性，这导致了世界范围的流行病，并给医疗保健系统带来了巨大的负担。因此，疫苗或鉴定用于治疗这些感染的新型或替代疗法的必要性已成为未满足且迫切的医疗需求。 尽管付出了巨大的努力，葡萄球菌感染的分子机制仍不完全清楚，但它似乎涉及大量的毒力因子，如粘附素，毒素和免疫调节剂。迄今为止，粘附到宿主组织被认为是感染的最重要阶段，并且在入侵生物体的存活中起着至关重要的作用。最有可能的是，毒素和免疫调节剂的合成在疾病过程的后期发挥作用，并取决于感染的第一步。因此，负责粘附的蛋白质长期以来被认为是药物和疫苗开发的靶点。 在葡萄球菌以及其他革兰氏阳性菌中，与宿主组织的粘附主要由细胞壁锚定（CWA）蛋白家族介导，该蛋白家族被称为微生物表面成分识别粘附基质分子（MSCRAP4）。这些蛋白质已显示a）介导对宿主蛋白质的粘附，B）参与自聚集和c）调节配体蛋白质的生物学。此外，MSCRA在动物感染模型中被证明是毒力因子，因为缺乏所有这些分子的突变体不能引起疾病。用重组MSCRA疫苗接种或抑制其功能可显著预防疾病。基于这些观察结果，人们普遍认为，MSCRA配体的鉴定将有助于更好地了解导致感染的分子事件，并将帮助我们设计有效的疫苗或靶向治疗药物用于疾病预防和治疗。 为此，本研究旨在建立一种高通量的噬菌体展示方法来鉴定MSCRA的配体。这项工作也将作为一个可行性研究，以确定这种技术是否适合大规模的配体发现细菌细胞表面蛋白。 公共卫生相关性：金黄色葡萄球菌是美国血液、下呼吸道、皮肤和软组织感染的主要原因，每年有近10万美国人死亡。我们的研究旨在更好地了解葡萄球菌感染，并为针对这种疾病的靶向治疗奠定基础。

DESCRIPTION (provided by applicant): Staphylococci cause a wide spectrum of infectious diseases spanning from rather minor skin infections to serious conditions such as sepsis, pneumonia and endocarditis. Although Staphylococcus aureus is by far the most virulent of species, Staphylococcus epidermidis and Staphylococcus lugdunensis have also been found to cause severe infections mostly in immunocompromised individuals. In the past, highly virulent S. aureus strains were confined to the hospital environment mostly affecting immunocompromised patients or individuals with underlying co-morbidities. Unfortunately, similar strains have recently emerged in the community causing severe infections in otherwise healthy individuals. To a lesser extent and slightly different pathology, Staphylococcus epidermidis and Staphylococcus lugdunensis seem to follow the same pattern. Most of these strains have evolved to be resistant to antibiotics, which resulted in a worldwide epidemic and an enormous burden for the healthcare system. Therefore, the necessity of a vaccine or identification of novel or alternative therapies for the treatment of these infections has become an unmet and pressing medical need. Despite intense efforts, the molecular mechanism of staphylococcal infections is incompletely understood, but it seemingly involves a large number of virulence factors such as adhesins, toxins and immunomodulators. To date, adhesion to host tissues is considered the foremost stage of infection and plays a vital role in the survival of the invading organism. Most likely, the synthesis of toxins and immunomodulators plays a role later in the disease process and depends on the first step of infection. Hence, proteins responsible for adherence have long been recognized as targets for drug and vaccine development. In staphylococci, as well as other Gram-positive bacteria, adhesion to host tissues is mostly mediated by a family of cell-wall-anchored (CWA) proteins referred to us microbial surface components recognizing adhesive matrix molecules (MSCRAMMs). These proteins have been shown to a) mediate adherence to host proteins, b) participate in self-aggregation and c) modulate the biology of the ligand protein. Furthermore, MSCRAMMs have been shown to be virulence factors in animal models of infection since a mutant lacking all these molecules fails to cause disease. Vaccination with recombinant MSCRAMMs or inhibition of their functions resulted in significant protection against disease. Based on these observations, it is widely agreed that identification of ligands for MSCRAMMs would contribute to a better understanding of molecular events leading to infection and would help us design effective vaccine or targeted therapeutics for disease prevention and treatment. To that end, this proposed research aims to establish a high-throughput phage display methodology to identify ligands for MSCRAMMs. This work will also serve as a feasibility study to determine if this technique is suitable for large-scale ligand discovery for bacterial cell surface proteins. PUBLIC HEALTH RELEVANCE: Staphylococcus aureus is the leading cause of bloodstream, lower respiratory tract, skin and soft tissue infections in the United States with an annual mortality approaching 100,000 Americans lives. Our research aims to better understand staphylococcal infection and to set the grounds for targeted therapies against this disease.