MOLECULAR GENETICS OF VARICELLA ZOSTER VIRUS

水痘-带状疱疹病毒（VZV）引起水痘和带状疱疹。我们有 开发了一种系统，通过分离组织来产生重组VZV 用四个重叠的VZV粘粒DNA培养细胞。重组VZV 将有助于研究VZV基因在感染中的作用。 突变已经被工程化到产生VZV突变体的cosmoprotein中， 不能表达病毒胸苷酸合成酶，糖蛋白V 核糖核苷酸还原酶和ORF 10基因。删除的病毒 核糖核苷酸还原酶基因的生长速度比野生型病毒慢 在体外，比亲本病毒对阿昔洛韦更敏感。而 HSV-1的ORF 10同源物，VP 16，是HSV-1复制所必需的。 在细胞培养中，VZV的ORF 10缺失不会导致VZV的细胞功能受损。 体外生长选定的VZV突变体将用于几内亚 猪，以确定病毒是否已经失去建立 中枢神经系统的潜伏期 急诊大肠杆菌β-半乳糖苷酶基因，已被插入VZV 基因组，产生的病毒产生的空斑，染色蓝色与X 加豚鼠正在接种这种病毒，以确定 能够支持病毒复制的细胞类型 建立潜伏期。单纯疱疹病毒（HSV）糖蛋白D (gD)编码主要中和抗原的基因已被插入到 VZV和所得病毒在表面上表达高水平的HSV gD 感染的细胞。用该VZV突变体接种豚鼠 导致产生了针对HSV-1的中和抗体。当 用活的单纯疱疹病毒攻击动物， 与对照动物相比，单纯疱疹感染的严重程度。 E.表达人Fab片段的大肠杆菌， 识别VZV已经产生并筛选了中和活性 对抗VZV

Varicella-zoster virus (VZV) causes chickenpox and herpes zoster. We have developed a system to generate recombinant VZV by transfecting tissue culture cells with four overlapping VZV cosmid DNAs. The recombinant VZV will be useful to study the role of individual VZV genes in infection. Mutations have been engineered into cosmids that yield VZV mutants that are unable to express the viral thymidylate synthetase, glycoprotein V ribonucleotide reductase, and ORF10 genes. Virus deleted for the ribonucleotide reductase gene grows at a slower rate than wild-type virus in vitro and is more sensitive to acyclovir than parental virus. While the HSV-1 homolog of ORF10, VP16, is essential for replication of HSV-1 in cell culture, deletion of ORF10 from VZV does not result in impaired growth in vitro. Selected VZV mutants will be used to inoculate guinea pigs to determine if the viruses have lost the ability to establish latency in the central nervous system. The E. coli beta-galactosidase gene, has been inserted into the VZV genome, and the resultant virus produces plaques that stain blue with X gal. Guinea pigs are being inoculated with this virus to determine the types of cells that are able to support viral replication and establishment of latency. The herpes simplex virus (HSV) glycoprotein D (gD) gene, encoding a major neutralizing antigen, has been inserted into VZV and the resulting virus expresses high levels of HSV gD on the surface of infected cells. Inoculation of guinea pigs with this VZV mutant resulted in development of neutralizing antibodies to HSV-1. When the animals were challenged with live herpes simplex virus they showed reduced severity of herpes simplex infection compared with control animals. Two combinatorial libraries of E. coli expressing human Fab fragments that recognize VZV have been generated and screened for neutralizing activity against VZV.