Pooling of samples to optimize SARS-CoV-2 diagnosis by RT-qPCR: comparative analysis of two protocols
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BRIEF REPORT
Pooling of samples to optimize SARS-CoV-2 diagnosis by RT-qPCR: comparative analysis of two protocols Fabiana Volpato 1,2,3 & Daiana Lima-Morales 1,3 & Priscila Lamb Wink 1,3,4 & Julia Willig 3,5 & Fernanda de-Paris 3 & Patricia Ashton-Prolla 6,7 & Afonso Luís Barth 1,2,3,4 Received: 1 September 2020 / Accepted: 9 October 2020 # Springer-Verlag GmbH Germany, part of Springer Nature 2020
Abstract This study has aimed to evaluate the use pool of samples as a strategy to optimize the diagnostic of SARS-CoV-2 by RT-qPCR. A total of 220 naso/orofaryngeal swab samples were collected and tested using two different protocols of sample pooling. Results from protocol A were identical with the individual results. However, for results from protocol B, reduced agreement (91%) was observed in relation to individual testing. Inconsistencies observed were related to RT-qPCR results with higher cycle thresholds. These results suggest that pooling of samples before RNA extraction is preferable in terms of diagnostic for SARS-CoV-2. Keywords Molecular diagnostic . Pooling sample . SARS-CoV-2 . RT-qPCR . Massive testing
Introduction Due to the exponential increase of respiratory syndromes by SARS-CoV-2, the World Health Organization (WHO) declared COVID-19 a pandemic of worldwide importance [1]. The rapid propagation of the virus significantly increases the demand on health care system. In response to the challenge of
* Afonso Luís Barth [email protected] 1
LABRESIS- Laboratório de Pesquisa em Resistência Bacteriana, Centro de Pesquisa Experimental, Hospital de Clínicas de Porto Alegre (HCPA), Porto Alegre, RS, Brazil
2
Programa de Pós-Graduação em Ciências Médicas (PPGCM), Faculdade de Medicina, Universidade Federal do Rio Grande do Sul (UFRGS), Porto Alegre, RS, Brazil
3
Laboratório de Diagnóstico de SARS-CoV-2 (Lab Covid), Hospital de Clínicas de Porto Alegre (HCPA), Porto Alegre, RS, Brazil
4
Programa de Pós-Graduação em Ciências Farmacêuticas (PPGCF), Faculdade de Farmácia, UFRGS, Porto Alegre, RS, Brazil
5
Programa de Residência Multiprofissional do Hospital de Clínicas de Porto Alegre (HCPA), Porto Alegre, RS, Brazil
6
Departamento de Genética, UFRGS, Porto Alegre, RS, Brazil
7
Grupo de Pesquisa e Pós-Graduação e Pós-Graduação, HCPA, Porto Alegre, RS, Brazil
identifying and isolating infected patients, two different types of tests have been used: real-time reverse transcriptase polymerase chain reaction (RT-qPCR) and serum IgM and/or IgG antibody detection assays. In order to avoid diagnostic cross-reactivity with other coronaviruses, most RT-qPCR assays include primers for at least two different molecular targets, such as nucleocapsid proteins 1 (N1) and 2 (N2) [2]. Also, this procedure could minimize false-negatives results associated with difference in sensitivity of primers. A major advantage of real time assays is that amplification and analysis of the final product are performed simultaneously in a closed system. Several countries have used RT-qPCR in large-scale efforts as a mainstream in
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