Identification of Host and Viral Determinants of Human Norovirus B cell Infection

摘要 人类诺如病毒是儿童严重腹泻和胃肠炎暴发的主要全球原因。 然而，治疗和预防策略的发展受到了严重限制，因为无法 在培养中繁殖这些病毒。我们最近发现诺如病毒在体外和体内感染B细胞， 他们利用共生细菌作为感染的辅助因素。这些新颖的发现直接导致了我们的 开发第一个人类诺沃克病毒培养系统，这是一个将给诺沃克病毒研究带来革命性变化的工具。 我们现在独一无二地准备识别影响人类的寄主限制和易感因素 诺如病毒感染B细胞以及B细胞感染的病毒决定因素。我们假设这是 信息将揭示关键的病毒-宿主相互作用，这些信息可以指导新疗法的开发， 候选疫苗，以及增强的诊断策略。在具体目标1中，我们将利用互补 有针对性和不偏不倚的方法，以制定一套全面的主机因素集，以限制或 促进人诺如病毒感染B细胞。在具体目标2中，将采取多管齐下的办法 确定体外复制适合性的病毒决定因素。策略包括病毒在中国的连续传播 并测试一组不同的人类诺沃克病毒基因型别的B细胞感染性。我们还将 构建人诺如病毒反向遗传学系统，以测试适应性突变的表型效应。这些 研究不仅将为人类感染B细胞的宿主和病毒决定因素提供重要的洞察力 诺如病毒，但它们也应该在第二代培养系统的开发中达到顶峰，以支持 广泛的病毒基因型别的强大感染。

SUMMARY Human noroviruses are a leading global cause of severe childhood diarrhea and gastroenteritis outbreaks. However, development of therapeutic and preventative strategies has been severely limited by the inability to propagate these viruses in culture. We recently discovered that noroviruses infect B cells in vitro and in vivo, and that they utilize commensal bacteria as a co-factor for infection. These novel findings led directly to our development of the first human norovirus cultivation system, a tool that will revolutionize norovirus research. We are now uniquely poised to identify host restriction and susceptibility factors influencing human norovirus infection of B cells as well as viral determinants of B cell infection. We hypothesize that this information will reveal key virus-host interactions that can guide the development of novel therapeutics, vaccine candidates, and enhanced diagnostic strategies. In Specific Aim 1, we will utilize complementary targeted and unbiased approaches to develop a comprehensive set of host factors that either restrict or promote human norovirus infection of B cells. In Specific Aim 2, a multi-pronged approach will be undertaken to identify viral determinants of in vitro replicative fitness. Strategies include serial passaging of virus in permissive cells and testing a diverse set of human norovirus genotypes for B cell infectivity. We will also construct a human norovirus reverse genetics system to test adaptive mutations for phenotypic effects. These studies will not only provide critical insight into host and viral determinants of B cell infection by human noroviruses but they should also culminate in the development of 2nd generation culture systems supporting robust infection of a broad panel of viral genotypes.