DEVELOPMENT OF Q PCR ASSAY FOR DETECTION OF ENTERIC CALICIVIRUSES

用于检测肠道杯状病毒的 Q PCR 检测方法的开发

• 批准号: 8358112
• 负责人: KAROL SESTAK
• 金额: $ 3.72万
• 依托单位: TULANE UNIVERSITY OF LOUISIANA
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-05-01 至 2012-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8358112
• 关键词: Adolescent; Animals; Antibodies; Antigens; Biological; Biological Assay; Biopsy; Blood Group Antigens; Calicivirus; Detection; Development; Diarrhea; Duodenum; Enteral; Epidemiology; Fever; Funding; Gastroenteritis; Genetic Variation; Grant; Housing; Human; Immunity; Infection; Infection Control; Macaca; Macaca mulatta; Modeling; National Center for Research Resources; Norovirus; Pathogenesis; Primates; Principal Investigator; RNA; Reporting; Research; Research Infrastructure; Resources; Sampling; Serum; Source; System; United States National Institutes of Health; Virus; Virus Shedding; antigen binding; base; cost; nonhuman primate; positional cloning

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Recently we reported the discovery of a cultivable calicivirus (Tulane virus; TV) isolated from a rhesus macaque. Based on preliminary results that show close similarities between TVs and noroviruses (NVs) in respect to their genetic diversity, epidemiology and histo-blood group antigen (HBGA) binding, plus the availability of in-house reverse genetics system, we are pursuing the development of a TV-based non-human primate surrogate model for human NV gastroenteritis. To support this effort in this study, we developed a qRT-PCR with the objective to enumerate TV RNA load in biological samples through the course of controlled infections of rhesus macaques. A TV was used to inoculate serologically pre-screened juvenile rhesus macaques to induce experimental infection. Additional macaques were inoculated with rhesus- and human-derived NoVs. While TV-inoculated animals developed infection characterized by virus shedding, fever and diarrhea, only asymptomatic infection was noted in NoV-inoculated macaques. A remarkable increase in virus-neutralizing serum antibodies was observed in TV-inoculated animals. Presence of TV antigens was visualized in duodenum biopsies obtained at post-inoculation day three. These results suggest that TV calicivirus might be used to study the pathogenesis and immunity of enteric caliciviruses.

该子项目是利用资源的众多研究子项目之一 由 NIH/NCRR 资助的中心拨款提供。子项目的主要支持 并且子项目的主要研究者可能是由其他来源提供的， 包括其他 NIH 来源。 子项目可能列出的总成本 代表子项目使用的中心基础设施的估计数量， NCRR 赠款不直接向子项目或子项目工作人员提供资金。 最近，我们报道了从恒河猴中分离出的可培养杯状病毒（杜兰病毒；TV）。初步结果显示 TV 和诺如病毒 (NV) 在遗传多样性、流行病学和组织血型抗原 (HBGA) 结合方面非常相似，加上内部反向遗传学系统的可用性，我们正在开发基于 TV 的人类 NV 胃肠炎非人灵长类动物替代模型。为了支持本研究的这项工作，我们开发了一种 qRT-PCR，目的是通过恒河猴的受控感染过程来计算生物样本中的 TV RNA 负载。使用电视对经过血清学预筛选的幼年恒河猴进行接种，以诱导实验性感染。其他猕猴接种了恒河猴和人类来源的 NoV。虽然接种 TV 的动物出现了以病毒脱落、发烧和腹泻为特征的感染，但接种 NoV 的猕猴仅出现无症状感染。在电视接种的动物中观察到病毒中和血清抗体显着增加。在接种后第三天获得的十二指肠活组织检查中可见TV抗原的存在。这些结果表明电视杯状病毒可用于研究肠道杯状病毒的发病机制和免疫。