Clinical validation of a multiplex PCR-based detection assay using saliva or nasopharyngeal samples for SARS-Cov-2, influenza A and B

• Published in: Scientific reports Vol. 12; no. 1; p. 3480
• Main Authors: Sahajpal, Nikhil S, Mondal, Ashis K, Ananth, Sudha, Njau, Allan, Jones, Kimya, Ahluwalia, Pankaj, Oza, Eesha, Ross, Ted M, Kota, Vamsi, Kothandaraman, Arvind, Fulzele, Sadanand, Hegde, Madhuri, Chaubey, Alka, Rojiani, Amyn M, Kolhe, Ravindra
• Format: Journal Article
• Language: English
• Published: England Nature Publishing Group 03.03.2022; Nature Publishing Group UK; Nature Portfolio
• Subjects: Coronaviruses; COVID-19; Humans; Influenza; Influenza A; Influenza A virus - isolation & purification; Influenza B virus - isolation & purification; Limit of Detection; Multiplex Polymerase Chain Reaction - methods; Nasopharynx - virology; Orthomyxoviridae; Pandemics; Polymerase chain reaction; Public health; Reproducibility of Results; Saliva; Saliva - virology; SARS-CoV-2 - isolation & purification; Severe acute respiratory syndrome coronavirus 2; Viruses
• ISSN: 2045-2322; 2045-2322
• DOI: 10.1038/s41598-022-07152-0

The COVID-19 pandemic has resulted in significant diversion of human and material resources to COVID-19 diagnostics, to the extent that influenza viruses and co-infection in COVID-19 patients remains undocumented and pose serious public-health consequences. We optimized and validated a highly sensitive RT-PCR based multiplex-assay for the detection of SARS-CoV-2, influenza A and B viruses in a single-test. This study evaluated clinical specimens (n = 1411), 1019 saliva and 392 nasopharyngeal swab (NPS), tested using two-assays: FDA-EUA approved SARS-CoV-2 assay that targets N and ORF1ab gene, and the PKamp-RT-PCR based assay that targets SARS-CoV-2, influenza viruses A and B. Of the 1019 saliva samples, 17.0% (174/1019) tested positive for SARS-CoV-2 using either assay. The detection rate for SARS-CoV-2 was higher with the multiplex assay compared to SARS-specific assay [91.9% (160/174) vs. 87.9% (153/174)], respectively. Of the 392 NPS samples, 10.4% (41/392) tested positive for SARS-CoV-2 using either assay. The detection rate for SARS-CoV-2 was higher with the multiplex assay compared to SARS-specific assay [97.5% (40/41) vs. 92.1% (39/41)], respectively. This study presents clinical validation of a multiplex-PCR assay for testing SARS-CoV-2, influenza A and B viruses, using NPS and saliva samples, and demonstrates the feasibility of implementing the assay without disrupting the existing laboratory workflow.