A Reverse Genetics Platform That Spans the Zika Virus Family Tree.

Zika virus (ZIKV), a mosquito-borne flavivirus discovered in 1947, has only recently caused large outbreaks and emerged as a significant human pathogen. In 2015, ZIKV was detected in Brazil, and the resulting epidemic has spread throughout the Western Hemisphere. Severe complications from ZIKV infection include neurological disorders such as Guillain-Barré syndrome in adults and a variety of fetal abnormalities, including microcephaly, blindness, placental insufficiency, and fetal demise. There is an urgent need for tools and reagents to study the pathogenesis of epidemic ZIKV and for testing vaccines and antivirals. Using a reverse genetics platform, we generated six ZIKV infectious clones and derivative viruses representing diverse temporal and geographic origins. These include three versions of MR766, the prototype 1947 strain (with and without a glycosylation site in the envelope protein), and H/PF/2013, a 2013 human isolate from French Polynesia representative of the virus introduced to Brazil. In the course of synthesizing a clone of a circulating Brazilian strain, phylogenetic studies identified two distinct ZIKV clades in Brazil. We reconstructed viable clones of strains SPH2015 and BeH819015, representing ancestral members of each clade. We assessed recombinant virus replication, binding to monoclonal antibodies, and virulence in mice. This panel of molecular clones and recombinant virus isolates will enable targeted studies of viral determinants of pathogenesis, adaptation, and evolution, as well as the rational attenuation of contemporary outbreak strains to facilitate the design of vaccines and therapeutics. Viral emergence is a poorly understood process as evidenced by the sudden emergence of Zika virus in Latin America and the Caribbean. Malleable reagents that both predate and span an expanding epidemic are key to understanding the virologic determinants that regulate pathogenesis and transmission. We have generated representative cDNA molecular clones and recombinant viruses that span the known ZIKV family tree, including early Brazilian isolates. Recombinant viruses replicated efficiently in cell culture and were pathogenic in immunodeficient mice, providing a genetic platform for rational vaccine and therapeutic design.

寨卡病毒（ZIKV）是一种于1947年发现的蚊媒黄病毒，直到近期才引发大规模疫情，并成为一种重要的人类病原体。2015年，寨卡病毒在巴西被检测到，由此引发的疫情已在西半球蔓延。寨卡病毒感染的严重并发症包括神经系统疾病，如成人中的吉兰 - 巴雷综合征，以及多种胎儿异常，包括小头畸形、失明、胎盘功能不全和胎儿死亡。迫切需要相关工具和试剂来研究流行的寨卡病毒的发病机制，并用于疫苗和抗病毒药物的测试。利用反向遗传学平台，我们构建了6个寨卡病毒感染性克隆以及衍生病毒，它们代表了不同的时间和地理来源。其中包括3个版本的MR766（1947年的原型毒株，其包膜蛋白上有无糖基化位点），以及H/PF/2013（2013年从法属波利尼西亚分离的一株人源病毒，代表了传入巴西的病毒）。在合成一株巴西流行毒株的克隆过程中，系统发育研究发现巴西存在两个不同的寨卡病毒分支。我们重建了SPH2015和BeH819015毒株的有活性克隆，它们分别代表每个分支的原始成员。我们评估了重组病毒的复制情况、与单克隆抗体的结合情况以及在小鼠中的致病性。这一组分子克隆和重组病毒分离株将有助于针对发病机制、适应性和进化的病毒决定因素进行靶向研究，以及对当代疫情毒株进行合理减毒，以促进疫苗和治疗方法的设计。 病毒的出现是一个鲜为人知的过程，寨卡病毒在拉丁美洲和加勒比地区的突然出现就证明了这一点。能够追溯到疫情之前并涵盖不断扩大的疫情的可变试剂，是了解调控发病机制和传播的病毒学决定因素的关键。我们已经构建了具有代表性的cDNA分子克隆和重组病毒，它们涵盖了已知的寨卡病毒家族树，包括早期的巴西分离株。重组病毒在细胞培养中高效复制，并在免疫缺陷小鼠中具有致病性，为合理的疫苗和治疗设计提供了一个遗传学平台。