Smartphone-linked system for diagnosis and epidemiological reporting of pathogens at the point of care

摘要 传染病仍然是世界上造成死亡和残疾的最大因素，而且由于其传播能力 迅速通过昆虫媒介或接触体液，迫切需要简单、敏感和 易于翻译的护理点测试，特别是对于难以根据以下条件进行区分的感染 仅仅是临床症状。该项目开发了一种新的护理点平台，用于定量诊断病毒 从全血样本中提取传染病(寨卡、登革热(1型和3型)和基孔肯雅病) 微流控平台，在微流控盒中执行样品前处理，然后实时 逆转录环介导等温扩增(RT-LAMP)在同一试剂盒中的应用 针对病原体靶标的特异引物。我们的手持式、廉价(约500美元)的护理点平台可与 它的测量要到智能手机上处理一系列放大反应的荧光图像。 动态扩增过程的图像分析被用来估计来自扩增的病原体数量 启动点使用了一种名为“空间”灯(S灯)的新方法。我们的初步数据显示 该方法提供了相当于传统聚合酶链的选择性和检测限 使用常规实验室仪器进行的反应(PCR)或LAMP反应，以及 从复杂介质中检测具有高特异性的靶标。S-LAMP方法展示了潜在的 每个反应检测1-5个病原体拷贝。该系统将使用严格的分层方法进行评估，使用 含有目标核酸序列的质粒，用于初步表征检测限，随后是 核酸病原体提取液化学裂解法检测全血病原体 以释放目标核酸。与巴西传染病研究所的合作者合作， 和Urbana的Carle诊所，该系统将用于患者样本库和测量 与金本位分析相比。重要的是，检测、图像处理和量化 使用智能手机微处理器执行检测，可以轻松地与基于云的数据库集成 便于与远程医生沟通和跟踪的报告系统 按卫生服务部门分列的流行病学数据。由此产生的平台将广泛适用于各种 对人类健康、动物健康、食品安全和 环境监测。

Abstract Infectious disease remains the world’s top contributor to death and disability, and, due to their ability to spread rapidly through insect vectors or contact with bodily fluids, there is an urgent need for simple, sensitive and easily translatable point-of-care tests, particularly for infections that are difficult to differentiate based upon clinical symptoms alone. This project develops a novel point-of-care platform to quantitatively diagnose viral infectious diseases (Zika, Dengue (types 1 and 3), and Chikungunya) from whole blood samples using a microfluidic platform that performs sample pre-processing in a microfluidic cartridge followed by real-time reverse-transcription loop-mediated isothermal amplification (RT-LAMP) in the same cartridge with pre-dried primers specific to pathogen targets. Our handheld, inexpensive (~$500), point-of-care platform communicates its measurements to a smartphone to process a sequence of fluorescence images of the amplification reaction. Image analysis of the dynamic amplification process is used to estimate the pathogen count from amplification initiation points using a novel approach called “spatial” LAMP (S-LAMP). Our preliminary data shows that the approach provides selectivity and detection limits that are equivalent to conventional Polymerase Chain Reaction (PCR) or LAMP reactions performed with conventional laboratory instruments, and the ability to detect the targets with high specificity from complex media. The S-LAMP approach demonstrates potential to detect 1-5 pathogen copies per reaction. The system will be evaluated using a rigorous tiered approach using plasmids containing the target nucleic acid sequence for initial characterization of detection limits, followed by nucleic acid pathogen extracts, culminating of detection of the pathogens in whole blood using chemical lysis to release target nucleic acids. Partnering with collaborators at the Institute for Infectious Diseases in Brazil, and Carle Clinic in Urbana, the system will be utilized on patient sample repositories and measurements compared to gold standard analysis. Importantly, as detection, image processing, and quantitation are performed with the smartphone microprocessor, detection is easily integrated with a cloud-based database reporting system that facilitates communication with remotely-located physicians and tracking of epidemiological data by health services. The resulting platform will be broadly applicable to a wide variety of multiplexed panels of pathogens that are of interest for human health, animal health, food safety, and environmental monitoring.