tetano
Editor, Senior Moderator
J Clin Virol. 2020 Apr 30;128:104391. doi: 10.1016/j.jcv.2020.104391. [Epub ahead of print]
A RT-PCR assay for the detection of coronaviruses from four genera.
Xiu L[SUP]1[/SUP], Binder RA[SUP]2[/SUP], Alarja NA[SUP]2[/SUP], Kochek K[SUP]2[/SUP], Coleman KK[SUP]3[/SUP], Than ST[SUP]3[/SUP], Bailey ES[SUP]4[/SUP], Bui VN[SUP]5[/SUP], Toh TH[SUP]6[/SUP], Erdman DD[SUP]7[/SUP], Gray GC[SUP]8[/SUP].
Author information
Abstract
BACKGROUND:
During the past two decades, three novel coronaviruses (CoVs) have emerged to cause international human epidemics with severe morbidity. CoVs have also emerged to cause severe epidemics in animals. A better understanding of the natural hosts and genetic diversity of CoVs are needed to help mitigate these threats.
OBJECTIVE:
To design and evaluate a molecular diagnostic tool for detection and identification of all currently recognized and potentially future emergent CoVs from the Orthocoronavirinae subfamily.
STUDY DESIGN AND RESULTS:
We designed a semi-nested, reverse transcription RT-PCR assay based upon 38 published genome sequences of human and animal CoVs. We evaluated this assay with 14 human and animal CoVs and 11 other non-CoV respiratory viruses. Through sequencing the assay's target amplicon, the assay correctly identified each of the CoVs; no cross-reactivity with 11 common respiratory viruses was observed. The limits of detection ranged from 4 to 4 ? 10[SUP]2[/SUP] copies/reaction, depending on the CoV species tested. To assess the assay's clinical performance, we tested a large panel of previously studied specimens: 192 human respiratory specimens from pneumonia patients, 5 clinical specimens from COVID-19 patients, 81 poultry oral secretion specimens, 109 pig slurry specimens, and 31 aerosol samples from a live bird market. The amplicons of all RT-PCR-positive samples were confirmed by Sanger sequencing. Our assay performed well with all tested specimens across all sample types.
CONCLUSIONS:
This assay can be used for detection and identification of all previously recognized CoVs, including SARS-CoV-2, and potentially any emergent CoVs in the Orthocoronavirinae subfamily.
Copyright ? 2020 Elsevier B.V. All rights reserved.
KEYWORDS:
COVID-19; Coronavirus; Emerging; Infectious diseases; SARS-CoV-2
PMID:32403008DOI:10.1016/j.jcv.2020.104391
A RT-PCR assay for the detection of coronaviruses from four genera.
Xiu L[SUP]1[/SUP], Binder RA[SUP]2[/SUP], Alarja NA[SUP]2[/SUP], Kochek K[SUP]2[/SUP], Coleman KK[SUP]3[/SUP], Than ST[SUP]3[/SUP], Bailey ES[SUP]4[/SUP], Bui VN[SUP]5[/SUP], Toh TH[SUP]6[/SUP], Erdman DD[SUP]7[/SUP], Gray GC[SUP]8[/SUP].
Author information
Abstract
BACKGROUND:
During the past two decades, three novel coronaviruses (CoVs) have emerged to cause international human epidemics with severe morbidity. CoVs have also emerged to cause severe epidemics in animals. A better understanding of the natural hosts and genetic diversity of CoVs are needed to help mitigate these threats.
OBJECTIVE:
To design and evaluate a molecular diagnostic tool for detection and identification of all currently recognized and potentially future emergent CoVs from the Orthocoronavirinae subfamily.
STUDY DESIGN AND RESULTS:
We designed a semi-nested, reverse transcription RT-PCR assay based upon 38 published genome sequences of human and animal CoVs. We evaluated this assay with 14 human and animal CoVs and 11 other non-CoV respiratory viruses. Through sequencing the assay's target amplicon, the assay correctly identified each of the CoVs; no cross-reactivity with 11 common respiratory viruses was observed. The limits of detection ranged from 4 to 4 ? 10[SUP]2[/SUP] copies/reaction, depending on the CoV species tested. To assess the assay's clinical performance, we tested a large panel of previously studied specimens: 192 human respiratory specimens from pneumonia patients, 5 clinical specimens from COVID-19 patients, 81 poultry oral secretion specimens, 109 pig slurry specimens, and 31 aerosol samples from a live bird market. The amplicons of all RT-PCR-positive samples were confirmed by Sanger sequencing. Our assay performed well with all tested specimens across all sample types.
CONCLUSIONS:
This assay can be used for detection and identification of all previously recognized CoVs, including SARS-CoV-2, and potentially any emergent CoVs in the Orthocoronavirinae subfamily.
Copyright ? 2020 Elsevier B.V. All rights reserved.
KEYWORDS:
COVID-19; Coronavirus; Emerging; Infectious diseases; SARS-CoV-2
PMID:32403008DOI:10.1016/j.jcv.2020.104391