Diagnostic efficiency of RT-LAMP integrated CRISPR-Cas technique for COVID-19: A systematic review and meta-analysis

Abstract

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.

Reverse Transcription-Loop-Mediated Isothermal Amplification-CRISPR-Cas13a Technology as a Promising Diagnostic Tool for SARS-CoV-2

“…We obtained an output of more than 7,000 articles as a result of a search using the keywords “RT-PCR diagnosis COVID-19”, of which almost 4,000 were published in 2021 alone. This result was compared with the findings of Bhatt et al ( 20 ) concerning papers related to RT-LAMP and CRISPR for SARS-CoV-2 diagnosis. Of these, we analyzed 10 articles on the RT-LAMP technique and 10 articles related to RT-LAMP-CRISPR-Cas technology, finding that only 1 applied the endonuclease Cas13 for SARS-CoV-2 diagnosis, but always on samples treated with an RNA extraction kit ( 21 ) ( Table 1 ).…”

“…Study of the state of the art revealed that greater efforts must be made to innovate in diagnostic methods; Bhatt et al ( 20 ) found 1,286 papers related to RT-LAMP and CRISPR for SARS-CoV-2 diagnosis (surprisingly, only 98 of these applied RT-LAMP integrated with CRISPR-Cas technology), in contrast with the 7,000 studies involving RT-PCR. This indicates that efforts should also be focused on developing more efficient RT-LAMP-CRISPR-Cas protocols without RNA purification, which would reduce the cost of the testing and also produce results faster.…”

RT-LAMP-CRISPR-Cas13a technology as a promising diagnostic tool for the SARS-CoV-2 virus

“…We obtained an output of more than 7000 articles as a result of "qPCR diagnosis COVID19" search, of almost 4000 in 2021 alone. This result was compared with findings of Bhatt A. et al [15] concerning papers related to RT-LAMP and CRISPR for SARS-CoV-2 diagnosis. Of these, we analyzed 10 articles on the RT-LAMP technique and 10 articles related to RT-LAMP-CRISPR-Cas technology, finding that only 1 applied the endonuclease Cas13 for SARS-CoV-2 diagnosis always on samples treated with an RNA extraction kit [35] (Table 3).…”

“…Study of the state of the art revealed that greater efforts must be made to innovate in diagnostic methods, as Bhatt A. et al [15] found 1286 papers related to RT-LAMP and CRISPR for SARS-CoV-2 diagnosis, in contrast to the 7000 studies involving qPCR ( surprisingly only 98 of these applied RT-LAMP integrated with CRISPR-Cas technology). This indicates that efforts should also be focused on developing more efficient RT-LAMP-CRISPR-Cas protocols without purification of RNA, which would reduce the cost of the testing and also produce results faster.…”

“…A study of the state of the art was conducted with the aim of comparing the use of different novel diagnostic techniques. First, we conducted a search in PubMed with "qPCR diagnosis COVID19" as keywords and compared the output with the number of publications on RT-LAMP and RT-LAMP-CRISPR strategies for COVID-19 diagnosis [15]. Finally, we collected data on the different sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) from 10 papers related to RT-LAMP and 10 papers on the RT-LAMP-CRISPR-Cas COVID19 diagnostic technique [16][17][18][19][20][21][22][23][24][25][26][27][28][29][30][31][32][33][34][35].…”