Objectives The SARS-CoV-2 pandemic has created an unprecedented need for rapid large-scale diagnostic testing to prompt clinical and public health interventions. Currently, several quantitative reverse-transcription polymerase chain reaction (RT-qPCR) assays recommended by the World Health Organization are being used by clinical and public health laboratories and typically target regions of the RNA-dependent RNA polymerase (RdRp), envelope (E) and nucleocapsid (N) coding region. However, it is currently unclear if results from different tests are comparable. This study aimed to clarify the clinical performances of the primer/probe sets designed by US CDC and Charité/Berlin to help clinical laboratories in assay selection for SARS-CoV-2 routine detection. Methods We compared the clinical performances of the recommended primer/probe sets using one hundred nasopharyngeal swab specimens from patients who were clinically diagnosed with COVID‐19. An additional 30 “pre-intervention screening” samples from patients who were not suspected of COVID-19 were also included in this study. We also performed sequence alignment between 31064 European SARS-CoV-2 and variants of concern genomes and the recommended primer/probe sets. Results The present study demonstrates substantial differences in SARS-CoV-2 RNA detection sensitivity among the primer/probe sets recommended by the World Health Organization especially for low-level viral loads. The alignment of thousands of SARS-CoV-2 sequences reveals that the genetic diversity remains relatively low at the primer/probe binding sites. However, multiple nucleotide mismatches might contribute to false negatives. Conclusion An understanding of the limitations depending on the targeted genes and primer/probe sets may influence the selection of molecular detection assays by clinical laboratories.
【저자키워드】 COVID-19, SARS-CoV-2, molecular detection, in silico analysis, Clinical performance, Real-time RT PCR, 【초록키워드】 public health, SARS-CoV-2 pandemic, Genome, diagnostic, variants of concern, Laboratory, COVID‐19, sensitivity, RT-qPCR, nucleocapsid, false negatives, Patient, Public health interventions, RdRP, RNA-dependent RNA polymerase, genetic diversity, molecular, Quantitative, binding, nucleotide, SARS-CoV-2 RNA detection, World Health Organization, viral loads, sequence, help, coding region, SARS-CoV-2 sequence, limitation, US CDC, nasopharyngeal swab specimen, objective, European, polymerase chain, Result, performed, clinically, diagnosed, contribute, comparable, reveal, target region, 【제목키워드】 real-time RT-PCR, primer set, the SARS-CoV-2 genome,