Virology Core

病毒学核心

• 批准号: 9464814
• 负责人: Yury A Bochkov
• 金额: $ 14.79万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/9464814
• 关键词: Adult; Air; Allergens; Antibodies; Antibody titer measurement; Asthma; Bacteria; Biological Assay; Biology; Birth; Catalytic RNA; Child; Chronic; Clinical; Cloning; Collection; Complementary DNA; Consult; Consultations; Cytomegalovirus; Detection; Development; Diagnostic; Environment; Epithelial Cells; Family; Farming environment; Flow Cytometry; Funding; Genetic; Hepatitis Delta Virus; Hospitalization; Human; Human Resources; Immune response; In Vitro; Infant; International; Lead; Life Style; Liquid substance; Measures; Metagenomics; Microspheres; Molecular; Molecular Biology; Monoclonal Antibodies; Nose; Pathogenesis; Pathogenicity; Physiological; Plaque Assay; Play; Preparation; Production; Quantitative Reverse Transcriptase PCR; RNA; Reagent; Recombinants; Reporter; Reproducibility of Results; Research; Research Personnel; Respiratory Tract Infections; Reverse Transcriptase Polymerase Chain Reaction; Rhinovirus; Role; Sampling; Serotyping; Serum; Severities; Severity of illness; Shipping; Specificity; Specimen; Structure; Techniques; Testing; Transfection; United States National Institutes of Health; Viral; Viral Load result; Viral Respiratory Tract Infection; Virus; Virus Replication; Wheezing; airway obstruction; base; cohort; cost; diagnostic assay; experimental study; genome sequencing; human subject; metagenomic sequencing; microbial; microbiome; molecular diagnostics; monolayer; neutralizing antibody; next generation sequencing; pathogen; plasmid DNA; prevent; programs; promoter; receptor; recombinant virus; respiratory; respiratory virus; viral RNA; virology

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.

项目概要/摘要 鼻病毒 (RV) 引起的呼吸道感染在发展、恶化和恶化过程中发挥着重要作用。 儿童和成人的慢性哮喘。该计划的重点是识别致病菌 与病毒生物学（细胞受体）、宿主（遗传学、免疫反应）和 导致气道功能生理变化并决定的环境（农业生活方式、微生物组） RV 引起的疾病的严重程度。项目 I 继续对涉及农场和非农场的出生队列进行研究 家庭，而项目 II 的重点是确定 RV-C 之间的相互作用，与更多相关 儿童严重喘息疾病和宿主气道上皮细胞。这两个项目都将使用病毒式传播 分子诊断测定，需要专门制备纯化的 RV 和其他分子试剂 例如对于获得可重复结果至关重要的抗体和 cDNA 克隆。病毒学的核心是 成立的目的是为项目 I 提供灵敏、准确的分子检测采样方案 识别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来分配 RV 物种和类型（RT-PCR 和 测序）用于临床样本中检测到的病毒。需要全面的病毒诊断来评估 每种病毒种类和 RV 类型在呼吸道疾病发病机制中的作用。病毒载量将为 通过 RT-qPCR 对项目 I 人类受试者的 RV 阳性鼻腔样本子集进行定量 随后进行病毒分离、全基因组测序、cDNA 克隆并在项目 II 中使用。核心将 分离、培养和表征人类气道上皮细胞的原代培养物，用于项目 II，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 的纯化库存（感染性和灭活）来进行项目 I-II 的实验 血清型/菌株，(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（空斑 和 TCID50) 测定，并测定抗 RV 血清中和抗体滴度。核心也将继续 为 UW AADCRC 项目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 资助的和国际研究人员合作。