Abstract
SARS-CoV-2 real-time reverse-transcription PCR (rRT-PCR) is the most effective testing system currently available to counter COVID-19 epidemics when potent treatments and vaccines are unavailable. Therefore, four SARS-CoV-2 rRT-PCR kits have been approved by the emergency-use-authorization (EUA) without clinical validation in Korea until March 15, 2020. This study evaluated the analytical and clinical performance of these kits. Allplex 2019-nCoV Real-time PCR (Seegene, Seoul, Korea), PowerChek 2019-nCoV (KogeneBiotech, Seoul), Real-Q 2019-nCoV Real-Time Detection (BioSewoom, Seoul), and StandardM nCoV Detection (SD BIOSENSOR, Osong, Korea) were evaluated. The limit of detection (LODs) of Allplex, PowerChek, and Real-Q was determined by testing the transcribed RNA of SARS-CoV-2 E and the RNA of SARS-CoV Frankfurt1. A total of 27 consecutive samples comprising 13 sputum, 12 nasopharyngeal swab (NPS), 1 urine and 1 stool sample were collected from 2 COVID-19 patients for sensitivity analysis. Precision was assessed via daily tests of positive and negative controls in each kit for 5 d. Reproducibility was examined by repeating 21 samples and 10-fold dilutions of 14 samples in pairs using Allplex. Specificity was evaluated with 24 other respiratory virus-positive samples. LOD of Allplex, PowerChek, and Real-Q were 153.9, 84.1, and 80.6 copies/mL, respectively. The degrees of association between Cts and log viral concentrations by Allplex and PowerChek was expressed as y = −3.319 log (x) + 42.039 (R = 0.96) and y = −3.392 log(x) + 43.113 (R = 0.98), respectively. One or more of the 4 kits detected 20 out of 27 clinical samples positive. Of the 20 positive samples, the detection rates of positives for Allplex, PowerChek, Real-Q, and StandardM were 90.0, 82.3, 75.0, and 100.0%, respectively, but those of PowerChek and Real-Q would be 100% if out-of-cutoff Cts were counted as positives. Precision was 100%. Interpretation of Allplex results was reproducible when Ct of E ≤33. All 4 kits showed no cross-reactivity with other respiratory viruses. Performance of the 4 kits indicated the suitability of these for diagnosis and follow-up testing of COVID-19. Laboratory doctors who initially implement these EUA kits must be able to interpret quality control parameters.
Highlights
Coronavirus disease 2019 (COVID-19) is an infectious disease caused by a novel coronavirus which was first detected in Wuhan city, China, in December 2019 [1,2,3]
limit of detection (LOD) values, the calculation of which were based on the power of detecting E-tRNA, were 153.94 copies/reaction for Allplex and 84.12 copies/reaction for PowerChek, respectively
LOD value for RealQ, the calculation of which was based on the power of detecting SARS-RNA was 80.60 copies/reaction, respectively (Figure 2)
Summary
Coronavirus disease 2019 (COVID-19) is an infectious disease caused by a novel coronavirus which was first detected in Wuhan city, China, in December 2019 [1,2,3]. 369,530 people were tested for SARS-CoV2 via rRT-PCR and 9,645 laboratory-confirmed COVID-19 cases were diagnosed nationwide until March 28 [21]. Commercial SARS-CoV-2 rRT-PCR kits have been approved by the EUA without clinical validation [6], because measures needed to counteract the COVID-19 pandemic require urgent deployment. All EUA kits should be prepared for clinical validation to obtain Ministry of Food and Drug Safety approval for the in vitro diagnostic kit before the end of the COVID-19 pandemic. It is, thereby, essential to evaluate their performance timely when the COVID-19 is less likely to end. The purpose of this study was to analytically and clinically validate 4 commercial kits approved by EUA in Korea
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