National Medical Research Center of Tuberculosis and Infectious Diseases, Ministry of Health, Moscow, Russia.
National Medical Research Center of Tuberculosis and Infectious Diseases, Ministry of Health, Moscow, Russia; Institute of Immunology, Moscow, Russia.
Methods. 2022 Jul;203:431-446. doi: 10.1016/j.ymeth.2021.04.007. Epub 2021 Apr 9.
Infectious diseases are a global health problem affecting billions of people. Developing rapid and sensitive diagnostic tools is key for successful patient management and curbing disease spread. Currently available diagnostics are very specific and sensitive but time-consuming and require expensive laboratory settings and well-trained personnel; thus, they are not available in resource-limited areas, for the purposes of large-scale screenings and in case of outbreaks and epidemics. Developing new, rapid, and affordable point-of-care diagnostic assays is urgently needed. This review focuses on CRISPR-based technologies and their perspectives to become platforms for point-of-care nucleic acid detection methods and as deployable diagnostic platforms that could help to identify and curb outbreaks and emerging epidemics. We describe the mechanisms and function of different classes and types of CRISPR-Cas systems, including pros and cons for developing molecular diagnostic tests and applications of each type to detect a wide range of infectious agents. Many Cas proteins (Cas3, Cas9, Cas12, Cas13, Cas14 etc.) have been leveraged to create highly accurate and sensitive diagnostic tools combined with technologies of signal amplification and fluorescent, potentiometric, colorimetric, lateral flow assay detection and other. In particular, the most advanced platforms -- SHERLOCK/v2, DETECTR, CARMEN or CRISPR-Chip -- enable detection of attomolar amounts of pathogenic nucleic acids with specificity comparable to that of PCR but with minimal technical settings. Further developing CRISPR-based diagnostic tools promises to dramatically transform molecular diagnostics, making them easily affordable and accessible virtually anywhere in the world. The burden of socially significant diseases, frequent outbreaks, recent epidemics (MERS, SARS and the ongoing COVID-19) and outbreaks of zoonotic viruses (African Swine Fever Virus etc.) urgently need the developing and distribution of express-diagnostic tools. Recently devised CRISPR-technologies represent the unprecedented opportunity to reshape epidemiological surveillance and molecular diagnostics.
传染病是影响数十亿人的全球健康问题。开发快速和敏感的诊断工具是成功管理患者和遏制疾病传播的关键。目前可用的诊断方法非常特异和敏感，但耗时且需要昂贵的实验室设置和经过良好培训的人员；因此，它们在资源有限的地区不可用，无法用于大规模筛查以及在爆发和流行时使用。迫切需要开发新的、快速的和负担得起的即时护理诊断检测方法。本综述重点介绍了基于 CRISPR 的技术及其成为即时护理核酸检测方法平台的前景，以及作为可部署的诊断平台，有助于识别和遏制爆发和新兴的流行疾病。我们描述了不同类别和类型的 CRISPR-Cas 系统的机制和功能，包括开发分子诊断测试的优缺点以及每种类型在检测广泛的传染病原中的应用。许多 Cas 蛋白（Cas3、Cas9、Cas12、Cas13、Cas14 等）已被用于创建高度准确和敏感的诊断工具，结合信号放大和荧光、电位、比色、侧向流动检测等技术。特别是最先进的平台——SHERLOCK/v2、DETECTR、CARMEN 或 CRISPR-Chip——能够以与 PCR 相当的特异性检测纳摩尔级别的致病核酸，但技术设置最少。进一步开发基于 CRISPR 的诊断工具有望极大地改变分子诊断，使其在世界任何地方都能轻松负担得起和使用。社会意义重大的疾病负担、频繁的爆发、最近的流行（MERS、SARS 和持续的 COVID-19）以及人畜共患病毒的爆发（非洲猪瘟病毒等）迫切需要开发和分发快速诊断工具。最近设计的 CRISPR 技术代表了重塑流行病学监测和分子诊断的前所未有的机会。
