Novel Determinants of Streptococcus Pyogenes Virulence and Protective Immunity in the Primate Oropharynx: A Genome-wide Strategy

灵长类口咽部化脓性链球菌毒力和保护性免疫的新决定因素：全基因组策略

• 批准号: 10387431
• 负责人: James MALLORY Musser
• 金额: $ 44万
• 依托单位: METHODIST HOSPITAL RESEARCH INSTITUTE
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-06-01 至 2022-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10387431
• 关键词: Acute; Address; Animal Model; Antibiotics; Biology; Candidate Disease Gene; Clinical; Complement; Consultations; Data; Disease; England; Epidemic; Equus caballus; Event; Female genitalia; Funding; Gene Deletion; Genes; Genomics; Goals; Gold; Grant; Human; Immunity; Immunization; Individual; Infection; Knowledge; Libraries; Macaca fascicularis; Membrane Proteins; Modeling; Molecular; Monobactams; Myositis; National Institute of Allergy and Infectious Disease; Necrotizing fasciitis; Organism; Oropharyngeal; Pathogenesis; Pharyngeal structure; Pharyngitis; Predisposition; Primates; Process; Proteins; Public Health; Publications; Recording of previous events; Research; Respiratory Tract Diseases; Role; Saliva; Septicemia; Serotyping; Severities; Site; Specimen; Streptococcus; Streptococcus equi; Streptococcus pyogenes; Study models; Technology; Testing; Upper Respiratory Infections; Upper respiratory tract; Vaccination; Vaccine Antigen; Vaccine Research; Vaccines; Virulence; Virulence Factors; Work; base; experience; fitness; gene product; genetic information; genome sequencing; genome wide screen; genome-wide; genome-wide analysis; human disease; improved; in vivo; innovation; mouse model; multiple myeloma M Protein; mutant; nonhuman primate; novel; novel vaccines; pathogen; pathogenic bacteria; prevent; public health relevance; reproductive tract; success; vaccine candidate; vaccine development; vaccinology

PROJECT SUMMARY The human bacterial pathogen Streptococcus pyogenes (group A streptococcus, GAS) causes more than 600 million cases of pharyngitis annually worldwide. Despite 100 years of effort, this organism has successfully defied attempts to create a vaccine that protects humans from pharyngitis and invasive infections. We believe a new strategy is warranted and necessary. The proposed research seeks to discover, on a genome-wide scale using a strategy not previously applied to GAS vaccinology, novel GAS genes contributing to pharyngitis, and exploit this information for vaccine research efforts. It is our thesis that this knowledge gap has severely limited the ability to fully understand the virulence determinants at work in the upper respiratory tract (URT) and create a protective GAS vaccine. We will address these issues by combining our recent successes applying transposon- directed insertion site sequencing (TraDIS) technology to GAS in several settings, together with our 19 years of productively using a cynomolgus macaque (nonhuman primate, NHP) model of pharyngitis, in vaccinology research. We will first use TraDIS to conduct in vivo genome-wide screens to systematically identify serotype M89 GAS genes required for colonization, clinical disease, and persistence in the URT of NHPs. These data will complement and enrich analogous information we have already generated for serotype M1 and M28 GAS. Our central hypothesis is that defining the GAS genes that contribute to fitness in the NHP URT will significantly improve our understanding of the molecular processes occurring in this niche, thereby filling a massive knowledge gap and leading to new strategies for creating a vaccine that protects against GAS pharyngitis. We propose the following three specific aims: Specific Aim 1: Exploit TraDIS for genome- wide identification of serotype M89 GAS genes required for colonization, acute clinical disease, and persistence (i.e., fitness) in the URT of NHPs. Specific Aim 2: Use our combined M1, M28, and M89 TraDIS URT screen data to generate isogenic gene-deletion mutant strains and validate the importance of additional selected candidate genes in causing GAS pharyngitis in NHPs. Specific Aim 3: Determine if vaccination of NHPs with proteins encoded by genes identified in Specific Aims 1 and 2 confers protective immunity against experimental pharyngitis caused by homologous (serotype M1) and heterologous (serotype M28) M protein serotypes of GAS. The proposed line of research will exploit and significantly expand studies funded by an R21 grant, and our innovative and successful application of TraDIS to GAS, our long history of GAS pharyngitis and vaccinology studies using NHPs, our analyses of human specimens, and our contributions to understanding the basic biology of GAS molecular pathogenesis and genomics. In essence, this research represents a new way forward for GAS vaccine efforts. Important to note, an analogous strategy has been used to identify four proteins present in a new vaccine that protects horses against “strangles,” a severe URT disease caused by the closely related pathogen, Streptococcus equi.

项目概要 人类细菌病原体化脓性链球菌（A 族链球菌，GAS）可引起 600 多种疾病 尽管经过 100 年的努力，全世界每年有 100 万例咽炎病例，但这种微生物已经成功地抵抗了这一挑战。 我们相信有一种新的疫苗可以保护人类免受咽炎和侵袭性感染。 拟议的研究旨在利用全基因组范围进行发现。 以前未应用于 GAS 疫苗学的策略、导致咽炎的新 GAS 基因以及利用 我们的论点是，这种知识差距严重限制了疫苗研究工作的信息。 能够充分了解上呼吸道 (URT) 中起作用的毒力决定因素并创建 我们将结合我们最近应用转座子的成功来解决这些问题。 定向插入位点测序 (TraDIS) 技术在多种环境下应用于 GAS，以及我们 19 年的经验 在疫苗学中有效地使用食蟹猴（非人灵长类动物，NHP）咽炎模型 我们将首先使用TraDIS进行体内全基因组筛选，以系统地鉴定血清型。 NHP 的定植、临床疾病和 URT 持久性所需的 M89 GAS 基因。 补充和丰富我们已经为 M1 和 M28 GAS 血清型生成的类似信息。 中心假设是，定义有助于 NHP URT 健康的 GAS 基因将 显着提高我们对这个利基中发生的分子过程的理解，从而 填补了巨大的知识空白，并制定了新的策略来创造一种可预防感染的疫苗 我们提出以下三个具体目标： 具体目标 1：利用 TraDIS 进行基因组分析。 广泛鉴定定植、急性临床疾病和持久性所需的血清型 M89 GAS 基因 （即健身）在 NHP 的 URT 中 具体目标 2：使用我们组合的 M1、M28 和 M89 TraDIS URT 屏幕。 数据产生等基因基因缺失突变株并验证额外选择的重要性 具体目标 3：确定 NHP 是否接种了 GAS 咽炎的候选基因。 具体目标 1 和 2 中确定的基因编码的蛋白质赋予针对实验的保护性免疫力 由 GAS 的同源（血清型 M1）和异源（血清型 M28）M 蛋白血清型引起的咽炎。 拟议的研究方向将利用和扩大由 R21 拨款资助的研究，我们的 TraDIS 在 GAS 上的创新和成功应用，我们在 GAS 咽炎和疫苗学方面的悠久历史 使用 NHP 进行的研究、我们对人类标本的分析以及我们对理解基础生物学的贡献 从本质上讲，这项研究代表了 GAS 的新发展方向。 值得注意的是，类似的策略已被用来识别存在的四种蛋白质。 在一种新疫苗中，可以保护马免受“勒死”的侵害，这是一种由密切接触者引起的严重 URT 疾病 相关病原体，马链球菌。