Comparative evaluation of six nucleic acid amplification kits for SARS-CoV-2 RNA detection

• Main Authors: Shuang Wu (Author), Xiaolu Shi (Author), Qiongcheng Chen (Author), Yixiang Jiang (Author), Le Zuo (Author), Lei Wang (Author), Min Jiang (Author), Yiman Lin (Author), Shisong Fang (Author), Bo Peng (Author), Weihua Wu (Author), Hui Liu (Author), Renli Zhang (Author), Patrick S. L. Kwan (Author), Qinghua Hu (Author)
• Format: Book
• Published: BMC, 2021-05-01T00:00:00Z.
• Subjects: article
• Online Access: Connect to this object online.
• Notes: 10.1186/s12941-021-00443-w; 1476-0711
• Keywords: SARS-CoV-2; COVID-19; Nucleic acid detection; Real-time reverse transcriptase PCR (RT-qPCR); Cross-priming isothermal amplification (CPA); Therapeutics. Pharmacology; RM1-950; Infectious and parasitic diseases; RC109-216; Microbiology; QR1-502
• Source: Annals of Clinical Microbiology and Antimicrobials, Vol 20, Iss 1, Pp 1-6 (2021)

Abstract Background SARS-CoV-2 is a newly emerged coronavirus, causing the coronavirus disease 2019 (COVID-19) outbreak in December, 2019. As drugs and vaccines of COVID-19 remain in development, accurate virus detection plays a crucial role in the current public health crisis. Quantitative real-time reverse transcriptase-polymerase chain reaction (RT-qPCR) kits have been reliably used for detection of SARS-CoV-2 RNA since the beginning of the COVID-19 outbreak, whereas isothermal nucleic acid amplification-based point-of-care automated kits have also been considered as a simpler and rapid alternative. However, as these kits have only been developed and applied clinically within a short timeframe, their clinical performance has not been adequately evaluated to date. We describe a comparative study between a newly developed cross-priming isothermal amplification (CPA) kit (Kit A) and five RT-qPCR kits (Kits B-F) to evaluate their sensitivity, specificity, predictive values and accuracy. Methods Fifty-two clinical samples were used including throat swabs (n = 30), nasal swabs (n = 7), nasopharyngeal swabs (n = 7) and sputum specimens (n = 8), comprising confirmed (n = 26) and negative cases (n = 26). SARS-CoV-2 detection was simultaneously performed on each sample using six nucleic acid amplification kits. The sensitivity, specificity, positive/negative predictive values (PPV/NPV) and the accuracy for each kit were assessed using clinical manifestation and molecular diagnoses as the reference standard. Reproducibility for RT-qPCR kits was evaluated in triplicate by three different operators using a SARS-CoV-2 RNA-positive sample. On the basis of the six kits' evaluation results, CPA kit (Kit A) and two RT-qPCR Kits (Kit B and F) were applied to the SARS-CoV-2 detection in close-contacts of COVID-19 patients. Results For Kit A, the sensitivity, specificity, PPV/NPV and accuracy were 100%. Among the five RT-qPCR kits, Kits B, C and F had good agreement with the clinical diagnostic reports (Kappa ≥ 0.75); Kits D and E were less congruent (0.4 ≤ Kappa < 0.75). Differences between all kits were statistically significant (P < 0.001). The reproducibility of RT-qPCR kits was determined using a coefficients of variation (CV) between 0.95% and 2.57%, indicating good reproducibility. Conclusions This is the first comparative study to evaluate CPA and RT-qPCR kits' specificity and sensitivity for SARS-CoV-2 detection, and could serve as a reference for clinical laboratories, thus informing testing protocols amid the rapidly progressing COVID-19 pandemic.