Virology Core

项目总结/摘要 由鼻病毒（rheinoviruses，RV）引起的呼吸道感染在呼吸道疾病的发生、加重和发展过程中起着重要作用。 儿童和成人哮喘的慢性化。该计划的重点是确定致病 与病毒生物学（细胞受体）、宿主（遗传学、免疫反应）和 环境（农业生活方式，微生物组），导致气道功能的生理变化，并决定 房车引起的疾病的严重程度。项目一继续在包括农场和非农场的出生队列中进行研究 家庭，而项目二的重点是确定RV-C之间的相互作用，与更多的 儿童严重喘息性疾病和宿主气道上皮细胞。这两个项目都将使用病毒 分子诊断分析，需要专门制备纯化的RV和其他分子试剂 例如抗体和cDNA克隆，它们对于获得可重复的结果是必需的。病毒学核心是 为项目I提供灵敏、准确的分子检测采样 鉴定所有常见呼吸道病毒（RVP）和细菌（宏基因组）的策略和测定 测序）。任务包括使用分子技术分配RV物种和类型（RT-PCR和 测序）用于临床样品中检测到的病毒。需要全面的病毒诊断来评估 每种病毒和RV类型在呼吸道疾病发病机制中的作用。病毒载量将是 通过RT-qPCR在来自项目I的人类受试者的RV阳性鼻样品亚组中定量， 随后的病毒分离、全基因组测序、cDNA克隆和用于项目II。芯会 分离、培养和表征用于项目II人气道上皮细胞的原代培养物，其中 将测试病毒和微生物分离株之间的相互作用。此外，该核心将支持体外 通过提供（1）所选RV的纯化储备液（感染性和灭活）进行项目I-II的实验 血清型/毒株，（2）RV特异性抗体，（3）RV-A、RV-B和RV-C分离株的cDNA克隆， 重组病毒生产，（4）进行病毒RNA定量（qRT-PCR）和RV感染性（噬斑 和TCID 50）测定，并测定抗RV血清中和抗体滴度。核心还将继续 为UW AADCRC计划提供使用这些试剂和病毒诊断检测的专业知识 研究人员，并与其他NIH资助和国际研究人员合作。

PROJECT SUMMARY/ABSTRACT Respiratory infections induced by rhinoviruses (RV) play important roles in development, exacerbation and chronicity of asthma in both children and adults. This program is focused on identifying pathogenic mechanisms related to the virus biology (cellular receptor), host (genetics, immune response), and environment (farming lifestyle, microbiome) that lead to physiologic changes in airway function and determine the severity of RV-induced illnesses. Project I continues studies in a birth cohort involving farm and non-farm families, while the emphasis of Project II is on determining interactions between RV-C, associated with more severe wheezing illnesses in children, and host airway epithelial cells. Each of the two projects will use viral molecular diagnostic assays and require specialized preparations of purified RVs and other molecular reagents such as antibodies and cDNA clones that are essential for obtaining reproducible results. The Virology Core is established to provide Project I with a repertoire of sensitive and accurate molecular detection sampling strategies and assays to identify all common respiratory viruses (RVP) and bacteria (metagenomic sequencing). Tasks including using molecular techniques to assign RV species and types (RT-PCR and sequencing) for viruses detected in clinical samples. Comprehensive viral diagnostics are needed to evaluate the role of each virus species and RV type in pathogenesis of respiratory illnesses. The viral loads will be quantified by RT-qPCR in a subset of RV-positive nasal samples from human subjects of Project I for subsequent virus isolation, complete genome sequencing, cDNA cloning and use in Project II. The core will isolate, grow and characterize primary cultures of human airway epithelial cells for use in Project II, in which interactions between viruses and microbial isolates will be tested. In addition, this core will support the in vitro experiments of Projects I-II by supplying (1) purified stocks (infectious and inactivated) of selected RV serotypes/strains, (2) RV-specific antibodies, (3) cDNA clones of RV-A, RV-B and RV-C isolates for recombinant virus production, (4) performing viral RNA quantification (qRT-PCR) and RV infectivity (plaque and TCID50) assays, and determining anti-RV serum neutralizing antibody titers. The Core will also continue to provide expertise for the use of these reagents and viral diagnostic assays to the UW AADCRC program investigators, and to collaborate with other NIH-funded and international investigators.