High-throughput Screenings to Identify Host Receptors for Staphylococcal Adhesins

描述(申请人提供)：葡萄球菌引起的传染病范围很广，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。虽然到目前为止，金黄色葡萄球菌是毒力最强的物种，但也发现表皮葡萄球菌和鲁顿葡萄球菌也会导致严重感染，主要发生在免疫功能低下的人身上。在过去，高毒力的金黄色葡萄球菌菌株仅限于医院环境中，主要影响免疫功能低下的患者或有潜在共病的个人。不幸的是，最近在该社区出现了类似的菌株，在其他健康的人中造成了严重感染。在较小程度和略有不同的病理情况下，表皮葡萄球菌和鲁顿葡萄球菌似乎遵循相同的模式。这些菌株中的大多数已经进化到对抗生素产生抗药性，这导致了全球流行，并给医疗系统带来了巨大的负担。因此，治疗这些感染的疫苗或确定新的或替代疗法的必要性已成为一个未得到满足的紧迫的医疗需求。 尽管进行了大量的努力，但葡萄球菌感染的分子机制仍不完全清楚，但它似乎涉及到大量的毒力因素，如粘附素、毒素和免疫调节剂。到目前为止，与宿主组织的黏附被认为是感染的最重要阶段，对入侵生物的生存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，负责黏附的蛋白质长期以来一直被认为是药物和疫苗开发的靶标。 在葡萄球菌和其他革兰氏阳性细菌中，与宿主组织的黏附主要是由细胞壁锚定(CWA)蛋白家族介导的，CWA蛋白是指识别黏附基质分子(MSCRAMM)的微生物表面成分。这些蛋白质已被证明a)介导与宿主蛋白的黏附，b)参与自我聚集，c)调节配基蛋白质的生物学。此外，MSCRAMM已被证明是感染动物模型中的毒力因素，因为缺乏所有这些分子的突变不会导致疾病。接种重组MSCRAMM或抑制其功能可显著预防疾病。基于这些观察，人们普遍认为，确定MSCRAMM的配体将有助于更好地了解导致感染的分子事件，并将有助于我们设计有效的疫苗或靶向治疗方法来预防和治疗疾病。 为此，这项拟议的研究旨在建立一种高通量噬菌体展示方法来识别MSCRAMM的配体。这项工作也将作为一项可行性研究，以确定该技术是否适合于大规模发现细菌细胞表面蛋白的配基。 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.