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A RT-PCR assay for the detection of coronaviruses from four genera

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Date
2020-07
Author
Xiu, Leshan
Binder, Raquel A.
Alarja, Natalie A.
Kochek, Kara
Coleman, Kristen K.
Than, Son T.
Bailey, Emily S.
Bui, Vuong N.
Toh, Teck Hock
Erdman, Dean D.
Gray, Gregory C.
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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 × 102 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.
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http://hdl.handle.net/20.500.12010/11181
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https://www.sciencedirect.com/science/article/pii/S1386653220301335?via%3Dihub
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Carrera 4 # 22-61 Teléfono: (+57 1) 242 7030 - 018000111022 Fax: (+57 1) 561 2107 Bogotá D.C., Colombia

Fundación Universitaria de Bogotá Jorge Tadeo Lozano | Vigilada Mineducación

Institución de educación superior privada, de utilidad común, sin ánimo de lucro y su carácter académico es el de Universidad.

Reconocimiento personería jurídica: Resolución 2613 del 14 de agosto de 1959 Minjusticia.

Institución de Educación Superior sujeta a inspección y vigilancia por el Ministerio de Educación Nacional.

 

Términos y condiciones | Políticas