MOLECULAR DIAGNOSTIC TESTS FOR SIMIAN HERPESVIRUSES

描述(申请人提供)：检测和诊断的能力 非人灵长类病毒的感染远远不是最理想的，在某些情况下 根本不存在。随着无特定病原体猴群的发展，它 对猴子病毒最敏感的测试将是至关重要的 可用来发展和监测这样的殖民地。该计划的总体目标 拟议的研究是开发猕猴BV的基本分子数据和 为了利用这些结果来开发更敏感的诊断测试， 比目前可用的更具体和更安全。要实现这些目标 将进行以下实验：1)指标细胞系 将被构建用于表达GFP的病毒分离的诊断用途 当被BV感染时，使BV病毒分离检测变得更加敏感 和更安全的程序。2)BV(ABV)致死缺失突变株将是 通过删除必要的结构基因并构建 补充ABV可以在其中复制的细胞系。此系统(ABV+ 补充细胞系)将使BV抗原和DNA的制备大量 更安全。3)通过对猕猴的评估，确定主要的BV免疫原 对单个BV蛋白的免疫反应。此信息将允许 高效疫苗和重组抗原的研制 诊断性测试。4)BV特异性糖蛋白基因的表达 一种可诱导的哺乳动物表达系统，所表达的蛋白用于 发展免疫检测方法。5)将开发一种TaqMan检测方法，可以检测到 并区分单纯疱疹病毒-1、单纯疱疹病毒-2 (HSV2)和BV。这将提供一种极其灵敏的诊断手段 人类BV病例。成功完成拟议的实验将 提供多种试剂，以更安全的方式检测 临床标本中的传染性BV，更安全的BV抗原制备 诊断试验，用于检测的更敏感和特异的血清试验 猕猴和HSV阳性人类血清中的抗BV抗体，以及基础 有关可能导致BV感染的抗原决定簇的信息 对感染的保护性免疫。

DESCRIPTION (Provided by applicant): The capability to detect and diagnose infections by non-human primate viruses is far less than optimal and in cases non-existent. As Specific Pathogen Free monkey colonies are developed, it will be essential that maximally sensitive tests for monkey viruses are available to develop and monitor such colonies. The overall goal of the proposed research is to develop basic molecular data for BV of macaques and to utilize these results to develop diagnostic tests that are more sensitive, more specific and safer than those currently available. To achieve these goals, the following experiments will be performed: 1) An indicator cell line will be constructed for diagnostic use in virus isolation which expresses GFP when infected by BV, making virus isolation testing for BV a more sensitive and safer procedure. 2) A lethal-deletion mutant strain of BV (ABV) will be constructed by deleting an essential structural gene and constructing a complementing cell line in which ABV can replicate. This system (ABV + complementing cell line) will make preparation of BV antigens and DNA much safer. 3) Major BV immunogens will be identified by assessing the macaque immune response to individual BV proteins. This information will allow development of effective vaccines and recombinant antigens for use in diagnostic tests. 4) Specific glycoprotein genes of BV will be expressed using an inducible mammalian expression system, and the expressed proteins used to develop immunciassays. 5) A TaqMan assay will be developed that can detect and differentiate herpes simplex virus-1 (HSV I), herpes simplex virus-2 (HSV2) and BV. This will provide an extremely sensitive means of diagnosing human BV cases. Successful completion of the proposed experiments will provide a number of reagents that will allow much safer detection of infectious BV in clinical samples, safer preparation of BV antigen for diagnostic tests, more sensitive and specific serological tests for detection of anti-BV antibodies in both macaque and HSV-positive human sera, and basic information regarding the antigenic determinants of BV which may induce protective immunity against infection.