Lateral flow assay for detecting colonization by Streptococcus agalactiae

用于检测无乳链球菌定植的侧流测定

• 批准号: 7481800
• 负责人: YING LI
• 金额: $ 20.1万
• 依托单位: BIOHELIX CORPORATION
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-04-01 至 2009-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7481800
• 关键词: Accounting; Agglutination; Antibiotics; Antibodies; Bathing; Binding; Biological Assay; Biotin; Birth; Buffers; Caring; Carrots - dietary; Centers for Disease Control and Prevention (U.S.); Clinical; Clinical Research; DNA; Detection; Devices; Diagnostic; Disease; Economics; Equilibrium; Fever; Filtration; Fluorescein-5-isothiocyanate; Goals; Haptens; Health Care Costs; Heating; Hour; Human; Incidence; Incubated; Invasive; Knowledge; Laboratories; Lateral; Latex; Latex Particles; Lead; Liquid substance; Localized; Manufacturer Name; Materials Testing; Medical center; Membrane; Meningitis; Methods; Microspheres; Molecular; Morbidity - disease rate; Mothers; Multi-Institutional Clinical Trial; National Institute of Allergy and Infectious Disease; Newborn Infant; Nucleic Acid Amplification Tests; Nucleic Acids; Performance; Perinatal Infection; Phase; Physicians; Plastics; Pneumonia; Polymerase Chain Reaction; Pregnancy; Premature Birth; Premature Rupture Fetal Membranes; Preparation; Process; Public Health; Published Comment; Reaction; Reagent; Recording of previous events; Risk; Risk Factors; Rupture; Sales; Sampling; Score; Screening procedure; Sensitivity and Specificity; Sepsis; Site; Solutions; Specificity; Specimen; Standards of Weights and Measures; Step Tests; Streptavidin; Streptococcal Infections; Streptococcus Group B; Swab; System; Term Birth; Testing; Time; Training; Tube; United States; United States Food and Drug Administration; Universities; Vagina; Water; Week; Woman; antimicrobial; base; cost; design; desire; early onset; helicase; instrumentation; internal control; migration; mortality; neonate; prevent; prophylactic; rapid detection

DESCRIPTION (provided by applicant): Streptococcus agalactiae, a.k.a. group B streptococcus (GBS), is one of the leading causes of morbidity, and mortality among newborns. We propose to develop a molecular assay for GBS using our proprietary helicase dependent amplification (HDA), and a lateral flow strip embedded in a device that encloses the reaction vessel to prevent contamination of the laboratory by amplicons. HDA is similar to the polymerase chain reaction (PCR) in that it uses two primers to exponentially amplify nucleic acids. Like PCR, HDA assays can use a competitive internal control; i.e., a template DNA of known concentration spiked into the raw sample that can be amplified by the same primers as the analyte, but detected separately. Unlike PCR, HDA is entirely isothermal, and thus does not require costly thermocyclers. In fact we routinely use water baths to perform assays. The objectives for Phase I are to: 1) Validate the sensitivity and specificity of the assay with a panel of DNA samples from near neighbor strains, as well as strains commonly found in the sampling sites. 2) Evaluate the performance of the filtration sample preparation method proposed in the body with clinical specimens from Dr. Gary Procop, University of Miami Medical Center (UMMC). 3) Develop a clinical plan & a pre-IDE for review by the Food and Drug Administration (FDA) for a multi-site clinical study to seek regulatory clearance for sale of the assay for human diagnostics. The milestone for this Phase I is to deliver a pre-IDE to FDA. Comments from FDA will be used to design a clinical study during Phase II. We believe our proposed assay system can obtained the moderate degree of complexity designation from the FDA. Indeed, to obtain such a designation, a system manufacturer must obtain a score of 12 of less when answering 7 questions covering: user knowledge, user training, reagent preparation, operational steps, testing materials, troubleshooting, and result interpretation. The answer to these 7 questions can receive a score of 1 or 3. We estimate our proposed system could get a score of 11. Considering our assay system will not require costly instrumentation and plastic disposables, we believe we can offer a useful alternative to the Xpert GBS(tm), if we can obtain FDA clearance for our test. Support from NIAID is essential to reaching this goal. PUBLIC HEALTH RELEVANCE: Group B streptococcus (GBS) infection of newborns usually takes place in the birth canal and can lead to sepsis, pneumonia, and meningitis. GBS infection of neonates is estimated to cost roughly $300 million in additional healthcare costs. The current "standard of care" is to screen all women at 35 to 37 weeks of gestation, and to eradicate GBS in colonized women. Despite this aggressive approach, invasive early-onset disease incidence has only declined by 34% in the US. Risk factors for early-onset invasive GBS in the newborns are: rupture of membranes more than 18 h before delivery, a febrile mother during labor, preterm delivery, and a history of GBS disease in a previous delivery. Current practice it to prescribe prophylactic antibiotics to mothers with the aforementioned risk profile, even in the absence of evidence of colonization by GBS. Premature rupture of membranes (PROM) occurs in approximately one third of preterm births and it increases the potential for perinatal infection because of its association with brief latency between membrane rupture, and delivery. Preterm pregnancies only represent 7% to 11% of births, but account for 32% to 38% of early-onset GBS disease because the mother was not tested prior to labor. CDC estimates that there are 500,000 pre-term births, and 143,000 births from mothers that received no pre-natal care in the United States. When culture is performed at the time of delivery, it seldom provides a sufficiently rapid turnaround time to influence the decision of whether or not to initiate antimicrobial therapy. A rapid test is clearly needed to test women just before delivery. Unfortunately, current rapid test options are insensitive, too slow, or too costly. Some culture-based tests allow for relatively rapid detection of GBS colonization in heavily colonized women. For example, the FDA cleared StrepB Carrot Broth" (Hardy Diagnostics, Santa Maria, CA) can detect heavy GBS colonization in as little as 6 hours after inoculation of the culture. FDA cleared probe based nucleic acid tests to detect GBS grown in cluture are also commercially available. Unfortunately, 6 hours is too long in many situations. Cepheid's GeneXpert System has obtained FDA clearance for use in laboratories cleared for performing test with "a moderate degree of complexity". This test can yield results in less than 1 hour, but it is far more costly than the slower alternatives. Some physicians have questioned the economic justification for the test. We propose to offer a lower cost molecular assay with an intermediate assay time of ~1 to 2 hours that should also be eligible for being categorized as a moderate degree of complexity assay by FDA.

描述（由申请人提供）：无乳链球菌，又名 B 族链球菌 (GBS)，是新生儿发病和死亡的主要原因之一。我们建议使用我们专有的解旋酶依赖性扩增 (HDA) 和嵌入反应容器的装置中的侧流条来开发 GBS 分子测定，以防止扩增子污染实验室。 HDA 与聚合酶链式反应 (PCR) 类似，它使用两个引物以指数方式扩增核酸。与 PCR 一样，HDA 检测可以使用竞争性内部对照；即，将已知浓度的模板 DNA 添加到原始样品中，可以通过与分析物相同的引物进行扩增，但单独检测。与 PCR 不同，HDA 是完全等温的，因此不需要昂贵的热循环仪。事实上，我们通常使用水浴进行测定。第一阶段的目标是： 1) 使用一组来自邻近菌株以及采样点常见菌株的 DNA 样本验证测定的灵敏度和特异性。 2) 使用迈阿密大学医学中心 (UMMC) Gary Procop 博士的临床标本评估体内提出的过滤样品制备方法的性能。 3) 制定临床计划和预 IDE，供美国食品和药物管理局 (FDA) 审查，进行多中心临床研究，以寻求监管部门批准销售用于人体诊断的检测方法。第一阶段的里程碑是向 FDA 提供 pre-IDE。 FDA 的意见将用于设计第二阶段的临床研究。我们相信我们提出的检测系统可以获得 FDA 的中等复杂度指定。事实上，要获得这样的称号，系统制造商在回答 7 个问题时必须获得 12 分或更低的分数，这些问题包括：用户知识、用户培训、试剂制备、操作步骤、测试材料、故障排除和结果解释。这 7 个问题的答案可以获得 1 或 3 分。我们估计我们提出的系统可以获得 11 分。考虑到我们的测定系统不需要昂贵的仪器和塑料一次性用品，我们相信我们可以提供一种有用的替代方案Xpert GBS(tm)，如果我们能够获得 FDA 的测试许可。 NIAID 的支持对于实现这一目标至关重要。 公共卫生相关性：新生儿 B 族链球菌 (GBS) 感染通常发生在产道中，可导致败血症、肺炎和脑膜炎。据估计，新生儿 GBS 感染将导致约 3 亿美元的额外医疗费用。目前的“护理标准”是在妊娠 35 至 37 周时对所有女性进行筛查，并根除殖民女性中的 GBS。尽管采取了这种积极的方法，美国侵袭性早发性疾病的发病率仅下降了 34%。新生儿早发侵袭性GBS的危险因素有：产前18小时以上胎膜破裂、临产时母亲发热、早产、既往分娩有GBS病史。目前的实践是，即使没有 GBS 定植的证据，也为具有上述风险状况的母亲开预防性抗生素。大约三分之一的早产会发生胎膜早破（PROM），它增加了围产期感染的可能性，因为它与胎膜破裂和分娩之间的短暂潜伏期有关。早产仅占出生的 7% 至 11%，但却占早发性 GBS 疾病的 32% 至 38%，因为母亲在临产前未接受检查。 CDC估计，美国有50万例早产，其中14.3万例是由未接受产前护理的母亲分娩的。当在分娩时进行培养时，它很少提供足够快的周转时间来影响是否开始抗菌治疗的决定。显然需要在分娩前对女性进行快速检测。不幸的是，当前的快速测试选项不敏感、太慢或成本太高。一些基于培养的测试可以相对快速地检测严重定植女性中的 GBS 定植情况。例如，FDA 批准的 StrepB Carrot Broth”（Hardy Diagnostics，圣玛丽亚，加利福尼亚州）可以在接种培养物后短短 6 小时内检测到大量 GBS 定植。 FDA 批准的基于探针的核酸测试可检测培养物中生长的 GBS不幸的是，Cepheid 的 GeneXpert 系统在许多情况下都太长了，可用于进行“中等复杂程度”测试的实验室。不到 1 小时，但它比较慢的替代方案成本更高，一些医生质疑该测试的经济合理性，我们建议提供一种成本较低的分子检测，中间检测时间也应该为 1 至 2 小时。有资格被 FDA 归类为中等复杂程度的检测。