Comparative investigation of the genealogy of hepatitis B virus

尽管自20世纪80年代以来已经可获得针对B型肝炎的有效疫苗，但全世界每年有超过800，000人死于B型肝炎病毒（HBV;嗜肝DNA病毒科）感染。在过去十年中描述了几种动物嗜肝DNA病毒，包括能够感染人肝细胞的蝙蝠HBV（TMBHBV），表明存在人畜共患HBV，并表明HBV的宏观进化史比以前认为的更复杂。灵长类动物中HBV的增加可能与NTCP受体分子进入肝细胞的使用密切相关。然而，我们的初步数据表明，（i）NTCP的使用范围超出了灵长类嗜肝DNA病毒和（ii）物种障碍可能涉及另一个（共）受体和宿主因素，除了NTCP。COHBV加入了两个PI，这两个PI一直在协同研究来自大量宿主的新型HBV，结合了计算机模拟、进化和体外分析。COMBHBV的中心假设是，B型肝炎是一种古老的人畜共患病，HBV祖先感染灵长类干细胞谱系的先决条件是在啮齿动物和蝙蝠等小型哺乳动物中进化的。COHBV将受益于众多的领先发现，包括来自全球数千只动物的完成实地工作和筛选的五种新型嗜肝DNA病毒。这些主要发现包括来自非洲和拉丁美洲的与潜在的人畜共患TMBHBV遗传相关的新蝙蝠HBV，以及来自非洲的能够与人类NTCP结合的新啮齿动物HBV。COHBV的总体目标是解开哺乳动物嗜肝DNA病毒的谱系，并调查进入和复制因子在多大程度上有助于嗜肝DNA病毒的宿主范围。这些目标将在三个工作包（WP）内实现。在WP 1中，我们将描述嗜肝DNA病毒的基因组，并建立体外研究新嗜肝DNA病毒所需的反向遗传学系统。在WP 2中，我们将通过将评估适应性进化的计算机模拟方法与评估细胞结合、进入和cccDNA形成和功能的体外方法相结合，来表征与啮齿动物和蝙蝠嗜肝DNA病毒进入和复制相关的宿主易感因素。来自WP 2的数据将用于WP 3，通过贝叶斯框架中的进化重建以及对原代细胞和表达必需宿主易感因子的选定哺乳动物细胞系中嗜肝DNA病毒宿主范围的病毒和细胞相关元素的实验评估来解开嗜肝DNA病毒谱系。COHBV将为确定潜在的非NTCP依赖的进入途径和重建嗜肝DNA病毒谱系提供可持续的基础。此外，新的啮齿动物HBV可能为一个易于处理的小鼠感染模型铺平道路，以确定HBV发病机制和治疗的关键方面，其有限的理解阻碍了迄今为止慢性B型肝炎的治愈。

Although efficient vaccines against hepatitis B have been available since the 1980s, over 800,000 people die yearly worldwide in consequence of hepatitis B virus (HBV; family Hepadnaviridae) infection. Several animal hepadnaviruses were described over the last decade, including a bat HBV (TMBHBV) capable of infecting human hepatocytes, suggesting zoonotic HBVs exist and indicating that the macro-evolutionary history of HBV is more complex than previously thought. The rise of HBV in primates may be closely linked to usage of the NTCP receptor molecule for hepatocyte entry. However, our preliminary data indicate that (i) NTCP usage expands beyond primate hepadnaviruses and (ii) species barriers may involve another (co-)receptor and host factors besides the NTCP. COMHBV joins two PIs that have been working in synergy on novel HBVs from a plethora of hosts combining in silico, evolutionary and in vitro analyses. COMHBV’s central hypothesis is that hepatitis B is an ancient zoonosis and that the preconditions of HBV ancestors to infect the primate stem lineage evolved in small mammals like rodents and bats. COMHBV will benefit from numerous lead findings, including five novel hepadnaviruses originating from completed fieldwork and screening of thousands of animals globally. Those lead findings include new bat HBVs from Africa and Latin America genetically related to the potentially zoonotic TMBHBV and a new rodent HBV from Africa capable of binding to the human NTCP. The overall goal of COMHBV is to unravel the genealogy of mammalian hepadnaviruses and to investigate to which extent entry and replication factors contribute to the hepadnaviral host range. Those aims will be achieved within three work packages (WP). In WP1, we will characterize hepadnaviral genomes and build the reverse genetics systems necessary to study the new hepadnaviruses in vitro. In WP2, we will characterize host susceptibility factors associated with rodent and bat hepadnaviral entry and replication by combining in silico approaches assessing adaptive evolution with in vitro approaches assessing cell binding, entry and cccDNA formation and function. Data from WP2 will be used in WP3 to unravel the hepadnavirus genealogy by evolutionary reconstructions in a Bayesian framework and by experimental assessments of virus- and cell-related elements underlying the hepadnaviral host range in primary cells and in selected mammalian cell lines expressing essential host susceptibility factors. COMHBV will provide sustainable ground to identify potential NTCP-independent entry pathways and reconstruct the hepadnaviral genealogy. Additionally, the new rodent HBV may pave ways for a tractable murine infection model to determine key aspects of HBV pathogenesis and treatment whose limited understanding prevents cure of chronic hepatitis B so far.