Viral Diversity and Pathogenicity in Mucosal Respiratory and Gastrointestinal Disease

项目摘要 人诺如病毒（HuNoV）是NIAID B类病原体，是全球急性胃肠炎的主要原因， 呼吸道合胞病毒（RSV）是C类病原体，是全球主要的呼吸道病原体， 孩子HuNoV感染导致所有年龄组的显著急性发病率， 免疫功能低下的癌症和移植患者，以及幼儿和老年人的死亡。RSV是 是儿童下呼吸道发病率和死亡率的主要原因， 免疫功能低下者、合并症患者和老年人的疾病和死亡。疾病负担 这些粘膜病原体的经济影响刺激了疫苗开发的广泛努力。 随着疫苗预计在未来5年内为这两种病原体，这个预许可窗口，加上我们的 最近开发了使用干细胞衍生的人肠道成功培养HuNoVs的系统 类器官（HIO）和最近对人肺类器官（HLO）中RSV感染的描述提供了理想的 在相关人类文化系统中应用分析基因组学和功能研究的窗口， 关于HuNoV和RSV多样性、进化和毒力的问题。我们建议的研究仔细利用 涵盖HuNoV和RSV疾病谱的选定临床标本，包括：（i）急性胃肠炎 在两个人群（美国和香港）的免疫功能正常儿童中，（ii）急性呼吸道感染， CDC赞助的新疫苗监测网络中的儿童，（iii）控制良好的志愿者HuNoV （iv）慢性HuNoV感染或急性RSV感染的移植和免疫功能低下患者 感染，（v）硝唑尼特（NTZ）治疗慢性HuNoV的首个随机双盲临床试验 在移植受者中，以及（vi）在第一个RSV-F疫苗（Novavax）的III期试验中的妇女和婴儿， 健康的孕妇，以防止疾病在其婴儿。除了病原体靶向测序，我们 将使用高通量、整合测序来评估生态位在病毒发病机制中的作用 分析临床样品的细菌组和病毒组的方法。全长（FL）基因组分析 HIEs中的HuNoV和功能研究将提供有关不同人群中病毒多样性的关键信息 随着时间的推移，在不同的宿主中，基因型内和基因型间重组体的存在， 出现对NTZ具有治疗抗性的突变体。RSV的FL基因组分析和RSV的功能研究 HLO和其他细胞系将建立RSV疾病严重程度的特异性病毒特征， 免疫压力和免疫治疗剂对病毒基因组的影响，包括抗原位点特异性 基序以及B和T细胞表位的变化。科学界将从大规模的 HuNoV和RSV的FL基因组数据库，以及在新的人类培养物中的功能研究， 在疫苗接种前全面解决病毒基因组特征、微生物生态学和宿主反应 导论.识别生物标志物、病毒和宿主靶标将指导治疗和疗法的开发。

PROJECT SUMMARY Human norovirus (HuNoV), a NIAID Category B pathogen, is the leading cause of acute gastroenteritis globally, and respiratory syncytial virus (RSV), a Category C pathogen, is the major global respiratory pathogen of children. HuNoV infections result in significant acute morbidity in all age groups, chronic disease in immunocompromised cancer and transplant patients, and death in young children and older adults. RSV is the leading cause of lower respiratory tract morbidity and mortality among children, and contributes significantly to illness and death in the immunocompromised, those with co-morbidities and older adults. The burden of disease and economic impact of these mucosal pathogens have stimulated extensive efforts for vaccine development. With vaccines anticipated in the next 5 years for both pathogens, this pre-licensure window, together with our recent development of a successful cultivation system for HuNoVs using stem cell-derived human intestinal organoids (HIOs) and the recent description of RSV infections in human lung organoids (HLOs) provide an ideal window to apply analytical genomics and functional studies in relevant human culture systems to address key questions about HuNoV and RSV diversity, evolution and virulence. Our proposed studies utilize carefully selected clinical specimens covering the spectrum of HuNoV and RSV illness, including: (i) acute gastroenteritis in immunocompetent children from two populations (USA and Hong Kong), (ii) acute respiratory infections in children in the CDC sponsored New Vaccine Surveillance Network, (iii) well-controlled volunteer HuNoV challenge studies, (iv) transplant and immunocompromised patients with chronic HuNoV infection or acute RSV infection, (v) the first randomized double-blind clinical trial of nitazoxanide (NTZ) for treatment of chronic HuNoV in transplant recipients, and (vi) women and infants in the first phase III trial of an RSV-F vaccine (Novavax) in healthy pregnant women for preventing disease in their infants. In addition to pathogen-targeted sequencing, we will evaluate the role of the ecological niche in viral pathogenesis using high throughput, integrated sequencing approaches to profile the bacteriome and virome of clinical samples. Full-length (FL) genomic analyses of HuNoV and functional studies in HIEs will provide critical information on virus diversity in different populations and over time in diverse hosts, the presence of intragenotypic and intergenotypic recombinants, and the appearance of mutants with treatment resistance to NTZ. FL genomic analyses of RSV and functional studies in HLOs and other cell lines will establish specific viral signatures of RSV disease severity, enable understanding the effects of immune pressure and immunotherapeutics on the viral genome including antigenic site specific motifs, and changes in B and T cell epitopes. The scientific community will benefit immensely from large databases of FL genomes for HuNoV and RSV, and functional studies in novel human cultures that comprehensively address viral genomic signatures, microbial ecology and host responses before vaccine introduction. Identifying biomarkers, viral and host targets will guide treatment and development of therapeutics.