Lower cost alternatives for molecular diagnosis of COVID-19: conventional RT-PCR and SYBR Green-based RT-qPCR
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CLINICAL MICROBIOLOGY - RESEARCH PAPER
Lower cost alternatives for molecular diagnosis of COVID-19: conventional RT-PCR and SYBR Green-based RT-qPCR Erick Gustavo Dorlass 1 & Cairo Oliveira Monteiro 1 & Amanda Oliveira Viana 1 & Camila Pereira Soares 1 & Rafael Rahal Guaragna Machado 1 & Luciano Matsumiya Thomazelli 1 & Danielle Bastos Araujo 1 & Fabyano Bruno Leal 1,11 & Erika Donizette Candido 1 & Bruna Larotonda Telezynski 1 & Camila Araujo Valério 1 & Vanessa Nascimento Chalup 1 & Ralyria Mello 1 & Flavia Jaqueline Almeida 2 & Andressa Simões Aguiar 3,5 & Anna Carlotta Mott Barrientos 4 & Carolina Sucupira 5 & Milena De Paulis 6 & Marco Aurélio Palazzi Sáfadi 2 & Daniella Gregorio Bonfim Prado Silva 3 & Janaina Joice Martins Sodré 4 & Mariana Pereira Soledade 5 & Samantha Faria Matos 2 & Sabrina Rodrigues Ferreira 4 & Célia Miranda Nunez Pinez 8 & Carolina Palamin Buonafine 2 & Leticia Nery Ferreira Pieroni 4 & Fernanda Mello Malta 9 & Rubia Anita Ferraz Santana 9 & Eloisa Corrêa Souza 6 & Ricardo Ambrosio Fock 7,8 & João Renato Rebelo Pinho 9,10 & Luís Carlos Souza Ferreira 1 & Viviane Fongaro Botosso 11 & Edison Luiz Durigon 1,12 & Danielle Bruna Leal Oliveira 1,11,13 Received: 18 May 2020 / Accepted: 22 July 2020 # Sociedade Brasileira de Microbiologia 2020
Abstract In March 2020, WHO declared a pandemic state due to SARS-CoV-2 having spread. TaqMan-based real-time RT-qPCR is currently the gold standard for COVID-19 diagnosis. However, it is a high-cost assay, inaccessible for the majority of laboratories around the world, making it difficult to diagnose on a large scale. The objective of this study was to standardize lower cost molecular methods for SARS-CoV-2 identification. E gene primers previously determined for TaqMan assays by Colman et al. (2020) were adapted in SYBR Green assay and RT-PCR conventional. The cross-reactivity test was performed with 17 positive samples for other respiratory viruses, and the sensibility test was performed with 8 dilutions (10 based) of SARS-CoV-2 isolated and 63 SARS-CoV-2-positive samples. The SYBR Green assays and conventional RT-PCR have not shown amplification of the 17 respiratory samples positives for other viruses. The SYBR Green-based assay was able to detect all 8 dilutions of the isolate. Erick Gustavo Dorlass, Cairo Oliveira Monteiro, Amanda Oliveira Viana and Camila Pereira Soares contributed equally to this work. Responsible Editor: Marina Baquerizo Martinez. Electronic supplementary material The online version of this article (https://doi.org/10.1007/s42770-020-00347-5) contains supplementary material, which is available to authorized users. * Danielle Bruna Leal Oliveira [email protected]
7
Department of Clinical and Toxicological Analysis, School of Pharmaceutical Sciences, University of São Paulo, São Paulo, Brazil
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Clinical Laboratory Division, Pharmacy and Clinical Laboratory Department, University Hospital, University of São Paulo, São Paulo, Brazil
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Department of Microbiology, Institute of Biomedical Sciences, University of São Paulo, Sã
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