Discriminating viral and bacterial meningitis infections with iDDS probes

使用 iDDS 探针区分病毒性和细菌性脑膜炎感染

• 批准号: 9253915
• 负责人: DAVID A SHAFER
• 金额: $ 24.26万
• 依托单位: GENETAG TECHNOLOGY, INC.
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-01-03 至 2017-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9253915
• 关键词: Antibiotics; Bacterial Meningitis; Biological Assay; Brain; Cerebrospinal Fluid; Characteristics; Clinical; Color; Custom; DNA; Data; Data Quality; Detection; Diagnosis; Diagnostic; Disease; Doctor of Philosophy; Enterococcus; Enterovirus; Etiology; FDA approved; Fluorescence; Freeze Drying; Generations; Genome; Goals; Government Agencies; Gram-Positive Bacteria; Haemophilus influenzae; Healthcare Systems; Hospitals; Hybrids; Individual; Infection; Inflammation; Joint Ventures; Label; Laboratories; Legal patent; Length of Stay; Manufacturer Name; Marketing; Membrane; Meningitis; Methods; Molecular; Neisseria meningitidis; Nucleic Acid Amplification Tests; Outcome; Parasites; Parechovirus; Patient Care; Patients; Performance; Pharmaceutical Preparations; Phase; Physicians; Preparation; Price; Property; Public Health; Reaction; Reagent; Regimen; Resistance; Risk; Sampling; Signal Transduction; Site; Specificity; Specimen; Spinal Cord; Streptococcus pneumoniae; Symptoms; System; Technology; Test Result; Testing; Time; Trichomonas vaginalis; University Hospitals; Vancomycin; Viral; Viral Genome; Viral meningitis; Virus; Virus Diseases; Water; Work; base; cost; cost effective; diagnostic assay; fluorophore; fungus; instrument; instrumentation; interest; microorganism; molecular diagnostics; mortality; multiplex detection; phase 1 study; phase 2 study; prevent; success; validation studies

Discriminating viral and bacterial meningitis infections with iDDS probes Confidential PI: Shafer, David A., PhD PROJECT SUMMARY Rapid and accurate molecular diagnostics assays empower physicians to make informed treatment decisions, especially when a disease state may result from variable causes that are indeterminable without such testing. Meningitis may be caused by infection with microorganisms such as viruses, gram-negative (GN) and gram- positive (GP) bacteria, fungi, or parasites. Most frequently, meningitis results from infection with non-polio enteroviruses (EV) and parechoviruses (PV), which typically self-resolve within 10 days. In contrast, bacterial meningitis has a high mortality rate that approaches 100% if not treated. Patients present with similar symptoms regardless of origin; thus, timely diagnosis of the causative agent is paramount for patient care. Patients are commonly treated with an `empiric' regimen of antibiotics until a definitive diagnosis can be made. Nucleic acid amplification tests (NAATs) reduce diagnosis turnaround times by up to several days compared to standard culturing methods, allowing shortened hospital stays, appropriate use of antibiotics, and a reduced financial burden to patients and the health care system. Currently, there are only two FDA-approved NAATs for diagnosing viral and/or bacterial meningitis. Unfortunately, these tests are only approved for use on the manufacturer's fully automated systems, the cost of which is often prohibitive. Further, these assays detect the highly variable viral genomes at a single target site. Collectively, these limitations highlight the need for an instrument-independent assay that can discriminate between viral and bacterial meningitis, with error-checking properties and confirmatory detection at redundant sites in the viral genomes. GeneTAG Technology, Inc. has developed error-checking DNA Detection Switch (iDDS) probes, which employ a fluorescent labeled probe and a slightly mismatched quencher-labeled antiprobe. In the absence of the intended target, the antiprobes hybridize to the probes, quenching their fluorescence and preventing off- target detection. Recently, we developed assays for detecting EV and PV employing two iDDS probes for each target, providing simultaneous 2-color signaling and automatic confirmation of positive or negative test results. We previously developed iDDS probes for GN and GP detection. Ultimately, we are interested developing an FDA-approved iDDS probe-based meningitis assay in a Sample-ReadyTM, lyophilized plate format that will parallel similar tests developed by our collaborator, BioGX Inc., where all reagents, primers, and probes are included, and the test would only require adding sample and molecular grade water. Aim 1 studies will focus on optimizing the dual-iDDS probe EV and PV assays to perform well in multiplex format with commercial EV and PV strains and patient cerebrospinal fluids, converting the multiplex test to Sample-ReadyTM format, and re- testing the Sample-ReadyTM assay with the original samples. A similar strategy will be employed in Aim 2 to prepare a Sample-ReadyTM GN/GP bacteria assay. Delivering a high fidelity, cost-effective meningitis assay should positively impact meningitis diagnostics and provide a direct benefit to public health.

IDDS探针鉴别病毒性脑膜炎和细菌性脑膜炎 项目总结 快速和准确的分子诊断分析使医生能够做出知情的治疗决定， 尤其是当疾病状态可能是由不能在没有这种测试的情况下确定的可变原因引起的情况下。 脑膜炎可能是由感染病毒、革兰氏阴性杆菌和革兰氏阴性杆菌等微生物引起的。 阳性(GP)细菌、真菌或寄生虫。最常见的脑膜炎是由感染非脊髓灰质炎引起的。 肠道病毒(EV)和细小病毒(PV)，通常在10天内自行溶解。相比之下，细菌 脑膜炎的死亡率很高，如果不治疗，死亡率接近100%。出现类似症状的患者 症状不分起因；因此，及时诊断病原体对病人的治疗至关重要。 在确诊之前，患者通常使用抗生素的“经验性”疗法进行治疗。 核酸扩增测试(NAAT)可将诊断周转时间最多缩短数天 标准培养方法，允许缩短住院时间，适当使用抗生素，并减少 给患者和医疗保健系统带来经济负担。目前，只有两种FDA批准的NAAT用于 诊断病毒性和/或细菌性脑膜炎。不幸的是，这些测试仅被批准用于 制造商的全自动化系统，其成本往往令人望而却步。此外，这些化验检测到 在单一靶点的高度可变的病毒基因组。总而言之，这些限制突显了 独立于仪器的分析，可以区分病毒性脑膜炎和细菌性脑膜炎，具有错误检查功能 病毒基因组中冗余位点的性质和确证检测。 GeneTAG Technology，Inc.开发了错误检查DNA检测开关(IDDS)探针，该探针 使用荧光标记的探针和稍有不匹配的猝灭剂标记的反探针。在没有的情况下 作为预定的靶点，反探针与探针杂交，猝灭它们的荧光并防止关闭- 目标检测。最近，我们开发了检测EV和PV的方法，每种方法都使用两个IDDS探针 目标，提供同步双色信号和对阳性或阴性测试结果的自动确认。 我们之前开发了用于GN和GP检测的IDDS探针。最终，我们有兴趣开发一种 FDA批准的基于IDDS探针的脑膜炎检测样品-ReadyTM，冻干平板格式，将 由我们的合作者BioGX Inc.开发的平行类似测试，其中所有试剂、引发剂和探针 包括在内，测试只需要添加样品和分子级水。目标1的研究将集中在 优化双IDDS探针EV和PV检测，以在与商业EV和 将多重试验转换为Sample-ReadyTM格式，并重新进行 用原始样品测试Sample-ReadyTM分析。目标2将采用类似的策略来 制备样品-ReadyTM GN/GP细菌测定法。提供高保真、经济高效的脑膜炎检测 应对脑膜炎诊断产生积极影响，并直接惠及公众健康。