End-point RT-PCR: A potential alternative for diagnosing coronavirus disease 2019 (COVID-19).

Autor: Silva Júnior JVJ; Setor de Virologia, Laboratório de Imunopatologia Keizo Asami, Universidade Federal de Pernambuco, Pernambuco, Brazil; Departamento de Microbiologia e Parasitologia, Universidade Federal de Santa Maria, Rio Grande do Sul, Brazil; Setor de Virologia, Departamento de Medicina Veterinária Preventiva, Universidade Federal de Santa Maria, Rio Grande do Sul, Brazil., Merchioratto I; Setor de Virologia, Departamento de Medicina Veterinária Preventiva, Universidade Federal de Santa Maria, Rio Grande do Sul, Brazil., de Oliveira PSB; Setor de Virologia, Departamento de Medicina Veterinária Preventiva, Universidade Federal de Santa Maria, Rio Grande do Sul, Brazil., Rocha Lopes TR; Setor de Virologia, Laboratório de Imunopatologia Keizo Asami, Universidade Federal de Pernambuco, Pernambuco, Brazil; Setor de Virologia, Departamento de Medicina Veterinária Preventiva, Universidade Federal de Santa Maria, Rio Grande do Sul, Brazil., Brites PC; Laboratório de Análises Clínicas, Setor de Biologia Molecular, Hospital Universitário de Santa Maria, Rio Grande do Sul, Brazil., de Oliveira EM; Laboratório de Análises Clínicas, Setor de Biologia Molecular, Hospital Universitário de Santa Maria, Rio Grande do Sul, Brazil., Weiblen R; Setor de Virologia, Departamento de Medicina Veterinária Preventiva, Universidade Federal de Santa Maria, Rio Grande do Sul, Brazil., Flores EF; Setor de Virologia, Departamento de Medicina Veterinária Preventiva, Universidade Federal de Santa Maria, Rio Grande do Sul, Brazil. Electronic address: eduardofurtadoflores@gmail.com.
Jazyk: angličtina
Zdroj: Journal of virological methods [J Virol Methods] 2021 Feb; Vol. 288, pp. 114007. Date of Electronic Publication: 2020 Oct 29.
DOI: 10.1016/j.jviromet.2020.114007
Abstrakt: Real-time reverse transcription-polymerase chain reaction (RT-qPCR) is considered the "gold standard" for the direct diagnosis of SARS-CoV-2 infections. However, routine diagnosis by RT-qPCR is a limitation for many laboratories, mainly due to the infrastructure and/or disproportionate relationship between demand and supply of inputs. In this context, and to increase the diagnostic coverage of SARS-CoV-2 infections, we describe an alternative, sensitive and specific one-step end-point RT-PCR for the detection of the SARS-CoV-2 E gene. The performance of the RT-PCR was evaluated in 43 clinical samples, of which 10 and 33 were previously identified as negative and positive, respectively, by RT-qPCR. Among the positive samples, 15 and 18 were from asymptomatic and symptomatic individuals, respectively. Here, 32/33 of the positive samples in the RT-qPCR, including from asymptomatic individuals, were found positive in the RT-PCR (Ct 15.94-34.92). The analytical sensitivity of the assay was about 7.15-9 copies of vRNA/μL, and nonspecific amplifications were not observed in SARS-CoV-2 negative samples. Importantly, the RT-PCR reactions were performed in a 10 μL final volume. Finally, considering specificity, analytical sensitivity and cost reduction, we believe that the RT-PCR platform described here may be a viable option for the diagnostic of SARS-CoV-2 infections in laboratories in which RT-qPCR is not available.
(Copyright © 2020 Elsevier B.V. All rights reserved.)
Databáze: MEDLINE
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