Improved molecular diagnosis of COVID-19 by the novel, highly sensitive and specific COVID-19-RdRp/Hel Real-time reverse transcription-PCR assay validated in vitro and with clinical specimens
Journal of Clinical Microbiology
; 58(5), 2020.
Article
in English
| GIM | ID: covidwho-1723516
ABSTRACT
On 31 December 2019, the World Health Organization was informed of a cluster of cases of pneumonia of unknown etiology in Wuhan, China. Subsequent investigations identified a novel coronavirus, now named severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), from the affected patients. Highly sensitive and specific laboratory diagnostics are important for controlling the rapidly evolving SARS-CoV-2-associated coronavirus disease 2019 (COVID-19) epidemic. In this study, we developed and compared the performance of three novel real-time reverse transcription-PCR (RT-PCR) assays targeting the RNA-dependent RNA polymerase (RdRp)/helicase (Hel), spike (S), and nucleocapsid (N) genes of SARS-CoV-2 with that of the reported RdRp-P2 assay, which is used in >30 European laboratories. Among the three novel assays, the COVID-19-RdRp/Hel assay had the lowest limit of detection in vitro (1.8 50% tissue culture infective doses [TCID50]/ml with genomic RNA and 11.2 RNA copies/reaction with in vitro RNA transcripts). Among 273 specimens from 15 patients with laboratory-confirmed COVID-19 in Hong Kong, 77 (28.2%) were positive by both the COVID-19-RdRp/Hel and RdRp-P2 assays. The COVID-19- RdRp/Hel assay was positive for an additional 42 RdRp-P2-negative specimens (119/273 [43.6%] versus 77/273 [28.2%];P 0.001), including 29/120 (24.2%) respiratory tract specimens and 13/153 (8.5%) non-respiratory tract specimens. The mean viral load of these specimens was 3.21 x 104 RNA copies/ml (range, 2.21 x 102 to 4.71 x 105 RNA copies/ml). The COVID-19-RdRp/Hel assay did not cross-react with other human-pathogenic coronaviruses and respiratory pathogens in cell culture and clinical specimens, whereas the RdRp-P2 assay cross-reacted with SARS-CoV in cell culture. The highly sensitive and specific COVID-19-RdRp/Hel assay may help to improve the laboratory diagnosis of COVID-19.
63231-63-0; human diseases; Rna; assays; diagnosis; aetiology; cell culture; diagnostic techniques; epidemics; genes; in vitro; laboratory diagnosis; nucleocapsid proteins; pathogens; pneumonia; respiratory system; reverse transcriptase PCR; severe acute respiratory syndrome; tissue culture; viral load; man; Severe acute respiratory syndrome coronavirus; Central Southern China; China; Homo; Hominidae; primates; mammals; vertebrates; Chordata; animals; eukaryotes; APEC countries; Developing Countries; East Asia; Asia; Severe acute respiratory syndrome-related coronavirus; Betacoronavirus; Coronavirinae; Coronaviridae; Nidovirales; positive-sense ssRNA Viruses; ssRNA Viruses; RNA Viruses; viruses; Coronavirus; syndromes; severe acute respiratory syndrome coronavirus 2; Coronavirus disease; ribonucleic acid; causal agents; etiology; People's Republic of China; reverse transcriptase polymerase chain reaction; Rt-pcr; Sars
Full text:
Available
Collection:
Databases of international organizations
Database:
GIM
Type of study:
Diagnostic study
/
Prognostic study
Language:
English
Journal:
Journal of Clinical Microbiology
Year:
2020
Document Type:
Article
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