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Detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and its first variants in fourplex real-time quantitative reverse transcription-PCR assays.
Durand, Mathieu; Thibault, Philippe; Lévesque, Simon; Brault, Ariane; Carignan, Alex; Valiquette, Louis; Martin, Philippe; Labbé, Simon.
  • Durand M; Plateforme RNomique et de Génomique Fonctionnelle, Université de Sherbrooke, Sherbrooke, QC, Canada.
  • Thibault P; Plateforme RNomique et de Génomique Fonctionnelle, Université de Sherbrooke, Sherbrooke, QC, Canada.
  • Lévesque S; Département de Microbiologie et d'Infectiologie, Faculté de médecine et des sciences de la santé, Université de Sherbrooke, Sherbrooke, QC, Canada.
  • Brault A; Laboratoire de Microbiologie, Centre Intégré Universitaire de Santé et de Services Sociaux (CIUSSS) de l'Estrie, Centre Hospitalier Universitaire de Sherbrooke (CHUS), Sherbrooke, QC, Canada.
  • Carignan A; Département de Biochimie et de Génomique Fonctionnelle, Faculté de médecine et des sciences de la santé, Université de Sherbrooke, Sherbrooke, QC, Canada.
  • Valiquette L; Département de Microbiologie et d'Infectiologie, Faculté de médecine et des sciences de la santé, Université de Sherbrooke, Sherbrooke, QC, Canada.
  • Martin P; Département de Microbiologie et d'Infectiologie, Faculté de médecine et des sciences de la santé, Université de Sherbrooke, Sherbrooke, QC, Canada.
  • Labbé S; Département de Microbiologie et d'Infectiologie, Faculté de médecine et des sciences de la santé, Université de Sherbrooke, Sherbrooke, QC, Canada.
Microb Cell ; 9(1): 1-20, 2022 Jan 03.
Article in English | MEDLINE | ID: covidwho-1622900
ABSTRACT
The early diagnosis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections is required to identify and isolate contagious patients to prevent further transmission of SARS-CoV-2. In this study, we present a multitarget real-time TaqMan reverse transcription PCR (rRT-PCR) assay for the quantitative detection of SARS-CoV-2 and some of its circulating variants harboring mutations that give the virus a selective advantage. Seven different primer-probe sets that included probes containing locked nucleic acid (LNA) nucleotides were designed to amplify specific wild-type and mutant sequences in Orf1ab, Envelope (E), Spike (S), and Nucleocapsid (N) genes. Furthermore, a newly developed primer-probe set targeted human ß2-microglobulin (B2M) as a highly sensitive internal control for RT efficacy. All singleplex and fourplex assays detected ≤ 14 copies/reaction of quantified synthetic RNA transcripts, with a linear amplification range of nine logarithmic orders. Primer-probe sets for detection of SARS-CoV-2 exhibited no false-positive amplifications with other common respiratory pathogens, including human coronaviruses NL63, 229E, OC43, and HKU-1. Fourplex assays were evaluated using 160 clinical samples positive for SARS-CoV-2. Results showed that SARS-CoV-2 viral RNA was detected in all samples, including viral strains harboring mutations in the Spike coding sequence that became dominant in the pandemic. Given the emergence of SARS-CoV-2 variants and their rapid spread in some populations, fourplex rRT-PCR assay containing four primer-probe sets represents a reliable approach to allow quicker detection of circulating relevant variants in a single reaction.
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Full text: Available Collection: International databases Database: MEDLINE Type of study: Diagnostic study / Experimental Studies / Prognostic study Topics: Variants Language: English Journal: Microb Cell Year: 2022 Document Type: Article Affiliation country: Mic2022.01.767

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Full text: Available Collection: International databases Database: MEDLINE Type of study: Diagnostic study / Experimental Studies / Prognostic study Topics: Variants Language: English Journal: Microb Cell Year: 2022 Document Type: Article Affiliation country: Mic2022.01.767