A Rapid, Low-Cost Point of Care Diagnostic for detection of Zika virus RNA

用于检测寨卡病毒 RNA 的快速、低成本护理点诊断

• 批准号: 9255921
• 负责人: Sahar Usmani-Brown
• 金额: $ 22.5万
• 依托单位: L2 DIAGNOSTICS, LLC
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-02-01 至 2019-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9255921
• 关键词: Acute; Adult; Aedes; African; Amniocentesis; Area; Asians; Bathing; Biological Assay; Blood; Blood specimen; Caribbean region; Central America; Chikungunya virus; Clinical; Colombia; Communities; Complex; Congenital Abnormality; Culicidae; Dengue; Dengue Virus; Dependence; Detection; Development; Diagnosis; Diagnostic; Diagnostic tests; Disease; Disease Outbreaks; Distant; Epidemic; Epidemiologic Monitoring; Equipment; Evaluation; Exanthema; Eye; Female of child bearing age; Fetus; Fever; Flavivirus; Goals; Guillain-Barré Syndrome; Health; Hour; Human; Imagery; Laboratories; Lateral; Mediating; Methods; Microcephaly; Micronesia; Mothers; Newborn Infant; Nucleic Acids; Patients; Performance; Phase; Pregnancy; Pregnant Women; Prenatal care; Procedures; Public Health; Puerto Rico; Quantitative Reverse Transcriptase PCR; RNA; RNA Sequences; RNA-Directed DNA Polymerase; Reaction; Reporting; Research; Resources; Risk; SYBR Green I; Saliva; Sampling; Sensitivity and Specificity; Serologic tests; Serum; Small Business Innovation Research Grant; South America; Southeastern United States; Specificity; Specimen; System; Techniques; Technology; Temperature; Testing; Time; TimeLine; Ultrasonography; United States Dept. of Health and Human Services; Urine; Viral; Viral Genome; Virus; Virus Diseases; Water; West Nile virus; Woman; Work; Zika Virus; assay development; authority; base; chikungunya; clinical Diagnosis; clinical diagnostics; cost; cost effective; cross reactivity; design; diagnostic assay; experience; point of care; point-of-care diagnostics; pregnant; prenatal; prototype; sample collection; screening; transmission process; validation studies; viral RNA; viral detection

ABSTRACT Zika virus (ZIKV) has rapidly emerged and spread through South and Central America, the Caribbean, and Puerto Rico since its last outbreak in Micronesia in 2007. Transmitted by the mosquito Aedes sp, its forecasted spread will have a major impact on the Southeast U.S. Most ZIKV infections remain asymptomatic or present with non-specific rash and fever; therefore, they have been difficult to diagnose and report. However, two major health consequences appear to be associated with the ZIKV outbreak which sets it apart from other flaviviruses such as West Nile Virus and Dengue; namely; (a) transmission from an infected mother to fetus resulting in reports of microcephaly in fetuses; and, (b) Guillain-Barre syndrome (GBS) in adults. Several teams have now developed qRT-PCR assays to detect ZIKV. However such tests are relatively expensive, require well-equipped laboratories with specialized equipment, and the procedure takes at least 3 hours to finish. There is an urgent need for a rapid, sensitive, specific and economical diagnostic test for ZIKV. Such an assay could be routinely used in resource-poor settings as well as in doctors' offices, including as part of regular prenatal care. Therefore, the goal of the SBIR Phase I project is to use loop mediated isothermal nucleic acid amplification (LAMP) to develop a rapid, sensitive point of care diagnostic for ZIKV. This technology is of low complexity, requiring only a water bath. Colorimetric results are visible to the naked eye in one hour or less. We will use serial dilutions of ZIKV and other viruses (including West Nile virus, Dengue viruses, and Chikungunya virus) spiked in human blood, saliva, and urine to determine the assay's sensitivity and specificity. Based on our laboratory's extensive previous experience with LAMP assay development, we forecast a lower limit of detection of 10-100 viral genomes, with very high specificity. In addition to rapid, colorimetric ZIKV detection, we will also develop a rapid, lateral flow assay to facilitate the differential analysis with other flaviviruses. The development of rapid point-of-care assays will reduce the dependence on central laboratory testing facilities for epidemiologic surveillance and clinical diagnosis, a key advantage in the resource-poor areas where the epidemic is currently prevalent. Further, we anticipate that the availability of a rapid test will result in the incorporation of ZIKV testing into the sustainable, routine evaluation of women who are pregnant or anticipating pregnancy, as well as their partners.

摘要 寨卡病毒（ZIKV）迅速出现并蔓延到南美洲和中美洲，加勒比地区， 自2007年密克罗尼西亚上一次爆发以来，波多黎各就一直有疫情。由蚊媒伊蚊传播， 传播将对美国东南部产生重大影响，大多数ZIKV感染仍无症状或存在 非特异性皮疹和发热;因此，它们很难诊断和报告。然而两 主要的健康后果似乎与ZIKV爆发有关，这使其有别于其他疾病。 黄病毒，如西尼罗河病毒和登革热;即;（a）从受感染的母亲传播给胎儿 导致报道胎儿中的小头畸形;和（B）成人中的格林-巴利综合征（GBS）。几 研究小组现在已经开发出qRT-PCR检测方法来检测ZIKV。然而，这种测试相对昂贵， 需要配备专业设备的设备齐全的实验室，并且程序至少需要3小时， 说完迫切需要一种快速、灵敏、特异和经济的ZIKV诊断测试。 这样的测定可以在资源贫乏的环境中以及在医生办公室中常规使用，包括作为 定期的产前护理因此，SBIR第一阶段项目的目标是使用环介导等温 本发明涉及一种用于核酸扩增（LAMP）的方法，以开发用于ZIKV的快速、灵敏的护理点诊断。这 该技术的复杂性低，仅需要水浴。比色结果是肉眼可见的， 我们将使用ZIKV和其他病毒（包括西尼罗河病毒、登革热病毒和登革热病毒）的系列稀释液， 病毒和基孔肯雅病毒）掺入人体血液，唾液和尿液中，以确定检测的灵敏度 和特异性。基于我们实验室在LAMP检测开发方面的丰富经验，我们 预测10-100个病毒基因组的检测下限，具有非常高的特异性。除了快速， 除了ZIKV的比色检测外，我们还将开发一种快速的侧流测定法，以促进差异分析 与其他黄病毒。快速床旁检测的发展将减少对中心检测的依赖。 流行病学监测和临床诊断的实验室检测设施， 目前流行病流行的资源贫乏地区。此外，我们预计， 快速测试将导致ZIKV测试纳入可持续的常规评估， 怀孕或预期怀孕，以及他们的伴侣。