Journal Article

Analytical comparisons of SARS-COV-2 detection by qRT-PCR and ddPCR with multiple primer/probe sets

Suo, Tao, Hu, Wenjia, Lan, Ke, Wang, Xin, Huang, Zhixiang, Guo, Dong, Feng, Jiangpeng, Chen, Yu, Liu, Xinjin, Zhang, Lu, Zhang, Qiuhan, Xiong, Yong, Chen, Guozhong, Guo, Ming

2020

Overview

Different primers/probes sets have been developed all over the world for the nucleic acid detection of SARS-CoV-2 by quantitative real time polymerase chain reaction (qRT-PCR) as a standard method. In our recent study, we explored the feasibility of droplet digital PCR (ddPCR) for clinical SARS-CoV-2 nucleic acid detection compared with qRT-PCR using the same primer/probe sets issued by Chinese Center for Disease Control and Prevention (CDC) targeting viral ORF1ab or N gene, which showed that ddPCR could largely minimize the false negatives reports resulted by qRT-PCR [Suo T, Liu X, Feng J, et al. ddPCR: a more sensitive and accurate tool for SARS-CoV-2 detection in low viral load specimens. medRxiv [Internet]. 2020;2020.02.29.20029439. Available from: https://medrxiv.org/content/early/2020/03/06/2020.02.29.20029439.abstract]. Here, we further stringently compared the performance of qRT-PCR and ddPCR for 8 primer/probe sets with the same clinical samples and conditions. Results showed that none of 8 primer/probe sets used in qRT-PCR could significantly distinguish true negatives and positives with low viral load (10 −4 dilution). Moreover, false positive reports of qRT-PCR with UCDC-N1, N2 and CCDC-N primers/probes sets were observed. In contrast, ddPCR showed significantly better performance in general for low viral load samples compared to qRT-PCR. Remarkably, the background readouts of ddPCR are relatively lower, which could efficiently reduce the production of false positive reports.

• Publisher: Taylor & Francis,Taylor & Francis Ltd,Taylor & Francis Group
• Subject: Betacoronavirus - genetics / Coronavirus Infections - diagnosis / Coronavirus Infections - virology / COVID-19 / diagnosis / digital PCR / DNA Primers / DNA Probes / false negative / false positive / Humans / Letter / Multiplex Polymerase Chain Reaction - methods / Pandemics / Pneumonia, Viral - diagnosis / Pneumonia, Viral - virology / real time PCR / Real-Time Polymerase Chain Reaction - methods / Real-Time Polymerase Chain Reaction - standards / SARS-CoV-2 / Sensitivity and Specificity / Severe acute respiratory syndrome coronavirus 2 / Viral Load