Evaluation of alternative RNA extraction methods for detection of SARS-CoV-2 in nasopharyngeal samples using the recommended CDC primer-probe set

Abstract

Background: The efficiency of isolation and purification of the viral genome is a critical step to the accuracy and reliability of RT-qPCR to detect SARS-CoV-2. However, COVID-19 testing laboratories were overwhelmed by a surge in diagnostic demand that affected supply chains especially in low and middle-income facilities. Objectives: Thus, this study compares the performance of alternative methods to extraction and purification of viral RNA in samples of patients diagnosed with COVID-19. Study design: Nasopharyngeal swabs were submitted to three in-house protocols and three commercial methods; viral genome was detected using the primer-probe (N1 and N2) described by CDC and viral load of samples were determined. Results: The in-house protocols resulted in detection of virus in 82.4 to 86.3% of samples and commercial methods in 94.1 to 98%. The disagreement results were observed in samples with low viral load or below the estimated limit of detection of RT-qPCR. Conclusion: The simplified methods proposed might be less reliable for patients with low viral load and alternative commercial methods showed comparable performance.

Keywords

• RNA extraction
• Direct RT-qPCR
• Viral diagnostic
• COVID-19