Amity Institute of Biotechnology, Amity University Haryana, Gurugram, India.
Department of Forensic Sciences, Amity School of Applied Sciences, Amity University Haryana, Gurugram, India.
Pathog Glob Health. 2022 Oct;116(7):410-420. doi: 10.1080/20477724.2022.2035625. Epub 2022 Feb 10.
To address the challenges associated with COVID-19 diagnosis, we need a faster, direct, and more versatile detection method for efficient epidemiological management of the COVID-19 pandemic. RT-qPCR (reverse transcription quantitative real-time Polymerase Chain Reaction) although the most popular diagnostic method suffers from a major drawback of equipment dependency and trained molecular biologists that limits rapid and large-scale screening, particularly in low resource regions. Reverse transcription loop-mediated isothermal amplification (RT-LAMP) is a feasible alternative for RT-qPCR; however, it also suffers from the drawback of false-positive issues. Recently, RT-LAMP has been integrated with the CRISPR-Cas technique to take care of the problems associated with RT-LAMP for COVID-19 diagnosis. In this study, a meta-analysis was conducted using three scientific databases considering the PRISMA guidelines to assess the diagnostic efficiency of RT-LAMP integrated CRISPR-Cas technology. Out of a total of 1286 studies on COVID-19, we identified 15 articles that met our eligibility criteria of using simultaneous RT-LAMP and CRISPR-Cas technique. Our meta-analysis of the included studies revealed that most of the studies were conducted in the USA with the N gene as the most common target and fluorescence-based detection method. The meta-analysis results of all included studies have further revealed a pooled sensitivity value of higher than 85% and a pooled specificity value of 80% with the confidence interval of 95%, respectively, as revealed from the forest plot and SROC curve. The accuracy rate of included studies was also calculated which varied from 77.4% to 100%. Furthermore, the precision of included studies varied from 75% to 100%. Lastly, a quality assessment of bias and applicability was performed based on QUADAS-2. Taken together, combined RT-LAMP and CRISPR-Cas technique could be a potential alternative to RT-qPCR particularly in low resource regions having a high demand for rapid testing.
为了解决与 COVID-19 诊断相关的挑战，我们需要一种更快、更直接、更通用的检测方法，以便对 COVID-19 大流行进行有效的流行病学管理。尽管 RT-qPCR（逆转录定量实时聚合酶链反应）是最流行的诊断方法，但它存在设备依赖性和需要经过训练的分子生物学家的主要缺点，这限制了快速和大规模筛查，特别是在资源匮乏的地区。逆转录环介导等温扩增（RT-LAMP）是 RT-qPCR 的可行替代方法；然而，它也存在假阳性问题的缺点。最近，RT-LAMP 已与 CRISPR-Cas 技术相结合，以解决与 COVID-19 诊断相关的 RT-LAMP 问题。在这项研究中，我们使用三个科学数据库进行了荟萃分析，考虑到 PRISMA 指南，以评估 RT-LAMP 与 CRISPR-Cas 技术相结合的诊断效率。在总共 1286 项关于 COVID-19 的研究中，我们确定了 15 篇符合我们使用同时进行 RT-LAMP 和 CRISPR-Cas 技术的资格标准的文章。我们对纳入研究的荟萃分析表明，大多数研究是在美国进行的，最常见的靶标是 N 基因，检测方法是基于荧光的。从森林图和 SROC 曲线可以看出，所有纳入研究的荟萃分析结果进一步显示，综合敏感性值高于 85%，综合特异性值为 80%，置信区间为 95%。纳入研究的准确性也有所计算，从 77.4%到 100%不等。此外，纳入研究的精度从 75%到 100%不等。最后，根据 QUADAS-2 对偏倚和适用性进行了质量评估。综上所述，联合 RT-LAMP 和 CRISPR-Cas 技术可能是 RT-qPCR 的一种潜在替代方法，特别是在对快速检测有高需求的资源匮乏地区。
