Multiplexed point-of-care molecular detection for multiple infections in co-endemic settings

项目摘要：影响热带贫困地区的传染病有 对人类健康和经济发展的破坏性影响，数十亿人处于危险之中。这些 疾病包括艾滋病毒/艾滋病、疟疾和结核病，但也包括被忽视的热带病(例如， 血吸虫病、丝虫病和其他蠕虫、微生物和原虫感染)。导致这些疾病的 这些疾病通常是共同流行的。伴随感染可以改变宿主的反应，疾病的预后， 传播动态和治疗结果，以及对准确解释 数据集中在单一感染上。一种单一、简单的诊断测试，可以特定和同时进行 以高灵敏度检测每个个体中存在的感染将是成本方面的一个突破， 时间和准确的诊断。不幸的是，目前对疾病传播和监测的诊断和监测 治疗的效果在很大程度上取决于通常不准确、劳动密集型或不可靠的方法。这些 限制在大规模药物管理项目中变得更加重要，在这些项目中，有效性的衡量标准 需要准确监测感染(和混合感染)状态、治疗成功、疾病传播 以及出现抗药性。宿主体液中病原体核酸的分子检测 组织提供可靠性、敏感性和特异性，但目前的方法需要基础设施的支持，高度- 训练有素的员工和昂贵而精致的设备。在这个项目中，我们将在以下方面取得实质性进展： 开发能够实现现场廉价分子的微流控、护理点/接触(POC)设备 由受过最低限度培训的人员进行感染诊断和疾病传播监测。具体来说，我们 将：目标1)开发、优化和验证新的两阶段扩增病原体检测方案 (坡道)使用台式测试；以及目标2)转让和调整开发的台式坡道测试 在目标1中提供一种新的微流控、多路复用芯片，并验证该芯片是否允许同时POC 多种病原体核酸的检测。这些目标的实现将作为概念的证明 并为在未来的工作中使用感染和动物模型进行更广泛的改进和测试奠定了基础 混合感染、临床样本和实地研究。这种方法将很容易转化为开创性的新 热带疾病和混合感染的现场就绪、POC分子诊断和监测技术。

Project summary: Infectious diseases that disproportionately impact impoverished regions in the tropics have devastating impacts on human health and economic development, with billions of people at risk. These diseases include HIV/AIDS, malaria, and tuberculosis, but also the neglected tropical diseases (eg, schistosomiasis, filariasis, and other helminth, microbial, and protozoan infections). The agents that cause these diseases are often co-endemic. Concomitant infections can alter host responses, disease prognosis, transmission dynamics, and treatment outcomes, as well as posing difficulties for accurate interpretation of data focused on single infections. A single, simple diagnostic test that could specifically and concurrently detect those infections present in each individual with high sensitivity would be a breakthrough in terms of cost, time, and accurate diagnosis. Unfortunately, current diagnosis and monitoring of disease transmission and efficacy of treatment depend largely on methods that are often inaccurate, labor-intensive, or unreliable. These limitations acquire added significance in mass drug administration programs, where measures of effectiveness require accurate monitoring of infection (and coinfection) status, treatment success, disease transmission rates, and emergence of drug resistance. Molecular detection of pathogen nucleic acids in host fluids or tissues offers reliability, sensitivity, and specificity, but current methods require infrastructural support, highly- trained staff, and expensive, delicate equipment. In this project, we will build upon our substantial advances in development of microfluidic, point-of-care/contact (POC) devices that enable on-site, inexpensive molecular diagnosis of infection and monitoring of disease transmission by minimally-trained personnel. Specifically, we will: Aim 1) develop, optimize, and validate a new 2-stage amplification pathogen detection protocol (RAMP) using benchtop assays; and Aim 2) transfer and adapt the benchtop RAMP assays developed in Aim 1 to a new microfluidic, multiplexing chip, and verify that the chip allows for simultaneous POC detection of multiple pathogen nucleic acids. Accomplishment of these aims will serve as proof of concept and set the stage for more extensive refinement and testing in future work using animal models of infection and coinfection, clinical samples, and field studies. The approach will readily translate into groundbreaking new technology for field-ready, POC molecular diagnosis and monitoring of tropical diseases and coinfections.