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

Vinícius Pietta Perez; Wallace Felipe Blohem Pessoa; Bruno Henrique Andrade Galvão; Eduardo Sergio Soares Sousa; Naiara Naiana Dejani; Eloiza Helena Campana; Marilia Gabriela Dos Santos Cavalcanti; Vlademir Vicente Cantarelli

doi:10.1016/j.jcvp.2021.100032

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

J Clin Virol Plus. 2021 Sep;1(3):100032. doi: 10.1016/j.jcvp.2021.100032. Epub 2021 Jul 3.

Authors

Vinícius Pietta Perez¹, Wallace Felipe Blohem Pessoa¹, Bruno Henrique Andrade Galvão¹, Eduardo Sergio Soares Sousa², Naiara Naiana Dejani^{1

2}, Eloiza Helena Campana^{2

3}, Marilia Gabriela Dos Santos Cavalcanti¹, Vlademir Vicente Cantarelli^{4

5}

Affiliations

¹ Departamento de Fisiologia e Patologia, Centro de Ciências da Saúde, Universidade Federal da Paraíba, Campus I - Cidade Universitária s/n, João Pessoa, PB 58051-900, Brazil.
² LaBiMol, Centro de Ciências Médicas, Universidade Federal da Paraíba, João Pessoa, Brazil.
³ Departamento de Ciências Farmacêuticas, Centro de Ciências da Saúde, Universidade Federal da Paraíba, João Pessoa, Brazil.
⁴ Universidade Feevale, Rio Grande de Sul, Brazil.
⁵ Universidade Federal de Ciências de Saúde de Porto Alegre (UFCSPA), Porto Alegre, Brazil.

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: COVID-19; Direct RT-qPCR; RNA extraction; Viral diagnostic.