Determinants of Enteric Calicivirus Infection.

肠道杯状病毒感染的决定因素。

• 批准号: 10442726
• 负责人: TIBOR FARKAS
• 金额: $ 37.32万
• 依托单位: TEXAS A&M AGRILIFE RESEARCH
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-07-01 至 2025-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10442726
• 关键词: Acute; Apical; B-Lymphocytes; Bile fluid; Binding Sites; Biodiversity; Biological; Biological Models; Biopsy; Blood Group Antigens; CRISPR/Cas technology; Calicivirus; Calicivirus Infections; Cell Culture System; Cell Culture Techniques; Cell Line; Cell surface; Cessation of life; Consumption; Coxsackie Viruses; Data; Development; Diarrhea; Disease; Enteral; Enterobacter; Enterocytes; Environment; Epithelium; Exhibits; Extracellular Domain; Fever; Gastroenteritis; Genetic; Goals; Healthcare; Hospitalization; Human; In Vitro; Infection; Integration Host Factors; Intervention; Intestines; Laboratories; Lamina Propria; M cell; Macaca; Maps; Mediating; Membrane; Modeling; Norovirus; Pathway interactions; Peripheral Blood Mononuclear Cell; Play; Positioning Attribute; Predisposition; Prevention strategy; Primates; Protein Isoforms; Public Health; Reproducibility; Research; Rhesus; Role; Serotyping; Site; Surface; System; Testing; Tight Junctions; Time; United States; Viral; Virus; Work; Zoonoses; adenovirus receptor; antigen binding; bile salts; comparative; cross-species transmission; enteric virus infection; genome wide screen; glycosylation; improved; infected B cell; insight; intestinal epithelium; monolayer; mutant; nonhuman primate; novel; permissiveness; polarized cell; receptor; societal costs

PROJECT SUMMARY Human norovirus (HuNoV) gastroenteritis is a significant public health burden worldwide. The lack of a robust and reproducible HuNoV cell culture system still limits our ability to study fundamental aspects of HuNoV infection. While several recent breakthroughs increased our ability to propagate HuNoVs in vitro (e.g. B cell and human enteroid cultures), these systems are not robust enough, can be time consuming and expensive or hard to replicate. The surrogate models available for HuNoV research do not necessarily reflect essential biological features of HuNoVs and their natural host. In this proposal we will use our novel rhesus enteric caliciviruses (ReCV) model that reflects the biological features and diversity of HuNoVs to identify host determinants of infection. Zoonotic/interspecies transmission of ReCVs and HuNoVs between human and non- human primate hosts suggests evolutionary conservation of shared factors of host susceptibility between the two genera. Here we seek to identify host determinants of enteric calicivirus infection using CRISPR-Cas9 genome wide screening. We recently identified a functional ReCV entry receptor that is necessary for ReCV permissiveness in cell culture. We hypothesize that the ReCV entry receptor is also involved in HuNoV infections. This hypothesis will be tested in the following specific aims. Aim 1. Characterize receptor mediated ReCV infection. Aim 2: Comparative characterization of HuNoV and ReCV infections in enteroid and B cell cultures. In the first aim we will dissect the role of HBGAs in ReCV receptor mediated entry by using ReCV isolates with disctinct HBGA binding, cell lines expressing different HBGAs and/or the receptor, Enterobacter SENG-6 EPS and different synthetic HBGAs. We will also evaluate the role of the different transmembrane isoforms of the receptor in infection and map the ReCV interaction site and importance of glycosylation. In the second aim, we will evaluate the role of ReCV entry receptor in HuNoV infections in enteroid and B cell cultures and identify other cell surface components playing a role in HuNoV infections. We will also evaluate the mechanism of bile or bile salts in promoting HuNoV infections and the role of M cells in infections of polarized cells. Our long-term goal is to identify and characterize viral and host determinants of ReCV and HuNoV infections and to develop novel intervention/prevention strategies. Our proposal uses a novel enteric calicivirus model to understand viral entry and the role of bile and HBGAs in enteric viral infections. Our findings with ReCVs may be directly transferable to HuNoV infection. The major significance of this project is the identification of determinants of susceptibility to enteric calicivirus infection that can lead to improved HuNoV cell culture systems and new intervention/prevention strategies.

项目总结 人类诺如病毒(HuNoV)胃肠炎是全球范围内严重的公共卫生负担。缺乏一个健壮的 而可复制的HuNov细胞培养系统仍然限制了我们研究HuNov基本方面的能力 感染。虽然最近的几项突破提高了我们在体外繁殖HuNov的能力(例如B细胞 和人类肠道培养)，这些系统不够健壮，可能会耗时和昂贵 很难复制。可用于HuNoV研究的替代模型不一定反映基本的 HuNov及其自然寄主的生物学特性。在这项提案中，我们将使用我们的新型恒河猴肠道 反映HuNov生物学特性和多样性的杯状病毒(ReCV)模型用于识别宿主 感染的决定因素。人与非人之间ReCVs和HuNov的人畜共患/种间传播 人类灵长类宿主提示宿主易感性的共同因素在进化上是保守的 两个属。在这里，我们试图使用CRISPR-Cas9来识别肠道杯状病毒感染的宿主决定因素 全基因组筛查。我们最近发现了一种具有功能的Recv进入受体，这是Recv所必需的 细胞培养中的放任性。我们推测，ReCV进入受体也参与了HuNov 感染。这一假设将在以下具体目标中得到检验。目的1.鉴定受体介导型 新冠状病毒感染。目的2：肠样和B细胞中人新城疫病毒和轮状病毒感染的比较特征 文化。在第一个目标中，我们将剖析HBGA在ReCV受体介导的ReCV进入中的作用 HBGA结合不同的分离株、表达不同HBGA和/或受体的细胞系肠杆菌 Seng-6 EPS和不同的合成HBGA。我们还将评估不同跨膜蛋白的作用 并绘制了ReCV相互作用的位置和糖基化的重要性。在 第二个目的，我们将评估ReCV进入受体在肠道和B细胞感染HuNov中的作用 培养并确定在HuNov感染中起作用的其他细胞表面成分。我们还将评估 胆汁或胆盐促进人新城疫病毒感染的机制及M细胞在人新城疫感染中的作用 极化细胞。我们的长期目标是识别和表征ReCV和Rcv的病毒和宿主决定因素 新病毒感染和开发新的干预/预防策略。我们的方案使用了一种新的肠道 杯状病毒模型，以了解病毒进入和胆汁和HBGA在肠道病毒感染中的作用。我们的 ReCVs的结果可能直接转移到HuNov感染上。这个项目的主要意义是 肠道杯状病毒感染易感性决定因素的鉴定 HuNov细胞培养系统和新的干预/预防策略。