Mobile based Nanoplasmonic Quantification of Mtb-derived Exosomes in Serum for Pediatric TB diagnosis

基于移动设备的纳米等离子体定量血清中结核分枝杆菌衍生的外泌体，用于儿童结核病诊断

• 批准号: 10019544
• 负责人: Christopher J Lyon
• 金额: $ 17.99万
• 依托单位: TULANE UNIVERSITY OF LOUISIANA
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-18 至 2021-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10019544
• 关键词: Algorithms; Area; Bacillus; Bacteria; Biological Assay; Biological Markers; Biopsy; Blood; Blood Circulation; Blood Tests; Blood specimen; Car Phone; Caregivers; Cells; Cessation of life; Child; Childhood; Clinical; Dark-Field Microscope; Data; Data Analyses; Detection; Devices; Diagnosis; Diagnostic; Diagnostic Procedure; Diagnostic Sensitivity; Disease; Drug Exposure; End Point Assay; Evaluation; Exhibits; Goals; Health Personnel; Healthcare; High Prevalence; Housing; Human Resources; Image; Imagery; Individual; Infrastructure; Label; Light; Logistic Regressions; Measures; Methods; Microscope; Modeling; Monitor; Mycobacterium tuberculosis; Mycobacterium tuberculosis antigens; Nature; Outcome; Parents; Patient-Focused Outcomes; Patients; Pediatric cohort; Performance; Procedures; Reading; Resistance; Resources; Risk; Sampling; Sensitivity and Specificity; Serum; Severities; Severity of illness; Slide; Sputum; Stress; Symptoms; System; Telephone; Temperature; Testing; Time; Training; Treatment Efficacy; Tuberculosis; Validation; antibody conjugate; base; clinical translation; communicable disease diagnosis; cost; cost effective; design; diagnostic accuracy; diagnostic assay; disease diagnosis; disorder control; exosome; extracellular vesicles; high reward; improved; innovation; interest; lens; lipoarabinomannan; low and middle-income countries; microscopic imaging; nanoparticle; nanoplasmonic; novel; novel strategies; pathogen; pediatric patients; portability; prototype; rapid diagnosis; rapid technique; real time monitoring; regression algorithm; sample collection; screening; transmission process; treatment response; tuberculosis diagnostics; tuberculosis drugs; user-friendly

Project Summary/Abstract There were an estimated 10.4 million tuberculosis (TB) cases, and 170,000 TB pediatric deaths in 2015, with >95% of TB deaths occur in low- and middle-income countries (LMIC). Rapid diagnosis is essential for improved TB outcomes, but diagnosis is more difficult in children, since children are less likely to exhibit typical TB symptoms and have paucibacillary (few bacteria) TB cases that are more difficult to detect by current methods, while parents are often resistant to stressful TB sample collection procedures. We therefore propose to explore a novel rapid, accurate, low-cost and easy-to-use mobile phone-based blood test for pediatric TB diagnosis in LMIC settings. Our novel detection method rapidly quantifies circulating extracellular vesicles (EVs) derived from host cells infected with the TB pathogen Mycobacterium tuberculosis (Mtb). Our preliminary data indicate that we can rapidly quantitate EVs carrying Mtb biomarkers, such as lipoarabinomannan and LpqH, in pediatric TB patient blood samples using a mobile-phone-based dark-field microscope (MDFM) platform. These findings support our long-term goal to develop a low-cost system to improve pediatric TB diagnosis in LMIC settings. We also anticipate that this system will allow rapid treatment monitoring to improve therapy and reduce exposure of pediatric patients to toxic anti-TB drugs. To achieve these outcomes we propose to pursue three goals. We collected our preliminary data with a MDFM that imaged transmitted light, but plan to design, develop, and validate the Aim 1 compact MDFM with a reflected light path to decrease the size of the device, simplify imagery, and increase the physical stability of the system. We will also develop a user-friendly application for image capture and data analysis on this platform. These features should markedly increase its feasibility for use in LMIC settings. Aim 2 will optimize assay performance for LMIC settings by redesigning assay materials for long-term storage under ambient conditions, and by determining optimal incubation times for different LMIC ambient temperature ranges. Aim 3 will analyze candidate Mtb-EV markers, including lipoarabinomannan and LpqH, build and a diagnostic model based on a multiple logistic regression algorithm, and validate diagnostic thresholds in separate validation cohort of pediatric TB patients and healthy controls with traceable clinical information. It will also compare the diagnostic performance of this multi-marker Mtb-EV assay to traditional diagnostic methods to evaluate its relative diagnostic performance. Our approach offers several innovative features valuable for pediatric TB control. It quantifies stable TB biomarkers on EVs in serum (1µL), rather than detecting Mtb in difficult to obtain, variable, non-quantitative and infectious biopsies. It employs a novel, compact MDFM and a nanoparticle-based end-point assay that is stable at ambient conditions. Finally, the ability of this approach to quantitate Mtb-EVs should allow rapid evaluation of disease severity for real-time monitoring of treatment efficacy and cures to minimize toxic drug exposure times.

项目总结/摘要 据估计，2015年有1040万结核病病例，17万结核病儿童死亡， 95%以上的结核病死亡发生在低收入和中等收入国家（LMIC）。快速诊断是必不可少的， 改善结核病的结果，但儿童的诊断更困难，因为儿童不太可能表现出典型的 结核病症状和少杆菌（细菌很少）结核病病例更难以通过目前的方法检测， 而父母通常对压力大的结核病样本采集程序有抵抗力。因此，我们建议探讨 一种新型的快速、准确、低成本和易于使用的基于移动的电话的儿科结核病血液检测 LMIC环境中的诊断。我们的新检测方法快速定量循环细胞外囊泡 (EVs)来源于感染TB病原体结核分枝杆菌（Mtb）的宿主细胞。我们的初步 数据表明，我们可以快速定量携带Mtb生物标志物的EV，例如脂阿拉伯甘露聚糖和LpqH， 使用基于移动电话的暗场显微镜（MDFM）平台， 这些发现支持了我们的长期目标，即开发一种低成本系统，以改善儿童结核病的诊断， LMIC设置。我们还预计，该系统将允许快速治疗监测，以改善治疗， 减少儿科患者接触有毒抗结核药物。为了实现这些目标，我们建议 三个目标。 我们用成像透射光的MDFM收集了我们的初步数据，但计划设计，开发， 并验证了Aim 1紧凑型MDFM与反射光路，以减少设备的尺寸，简化 图像，并增加系统的物理稳定性。我们亦会开发一个方便使用的应用程式， 图像采集和数据分析。这些特点应显着增加其使用的可行性 在LMIC设置中。目标2将通过重新设计测定材料，优化LMIC设置的测定性能， 在环境条件下长期储存，并通过确定不同LMIC的最佳孵育时间 环境温度范围。目标3将分析候选Mtb-EV标志物，包括脂阿拉伯甘露聚糖和 LpqH，建立了基于多元逻辑回归算法的诊断模型，并验证了诊断结果 具有可追溯临床特征的儿科TB患者和健康对照的单独验证队列中的阈值 信息.它还将比较这种多标志物Mtb-EV检测与传统Mtb-EV检测的诊断性能。 诊断方法，以评估其相对诊断性能。 我们的方法提供了几个创新的特点，对儿科结核病控制有价值。它量化了稳定的结核病 血清中EV的生物标志物（1μL），而不是检测难以获得、可变、非定量和 感染性活检它采用了一种新型的，紧凑的MDFM和基于纳米颗粒的终点测定， 在环境条件下。最后，这种方法定量Mtb-EV的能力应该允许快速评估Mtb-EV。 疾病的严重程度，以实时监测治疗效果和治愈，从而最大限度地减少有毒药物的暴露时间。