True or false: what are the factors that influence COVID-19 diagnosis by RT-qPCR?

Expert Rev Mol Diagn. 2022 Feb;22(2):157-167. doi: 10.1080/14737159.2022.2037425. Epub 2022 Feb 21.

Abstract

Introduction: The Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) disease has had a catastrophic impact on the world resulting in several deaths. Since World Health Organization declared the pandemic status of the disease, several molecular diagnostic kits have been developed to help the tracking of viruses spread.

Areas covered: This review aims to describe and evaluate the currently reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR) diagnosis kit. Several processes used in COVID-19 diagnostic procedures are detailed in further depth to demonstrate optimal practices. Therefore, we debate the main factors that influence the viral detection of SARS-COV-2 and how they can affect the diagnosis of patients.

Expert opinion: Here is highlighted and discussed several factors that can interfere in the RT-PCR analysis, such as the viral load of the sample, collection site, collection methodology, sample storage, transport, primer, and probe mismatch/dimerization in different brand kits. This is a pioneer study to discuss the factor that could lead to the wrong interpretation of RT-qPCR diagnosis of SARS-CoV-2. This study aimed to help the readers to understand what very likely is behind a bad result of SARS-CoV-2 detection by RT-PCR and what could be done to reach a reliable diagnosis.

Keywords: COVID-19; Coronavirus; RT-PCR; diagnosis; sensitivity.

Publication types

  • Research Support, Non-U.S. Gov't
  • Review

MeSH terms

  • COVID-19 Testing
  • COVID-19* / diagnosis
  • Humans
  • RNA, Viral / analysis
  • RNA, Viral / genetics
  • Reverse Transcriptase Polymerase Chain Reaction
  • SARS-CoV-2 / genetics
  • Sensitivity and Specificity

Substances

  • RNA, Viral

Grants and funding

This work was supported by the Fundação Cearense de Apoio ao Desenvolvimento Científico e Tecnológico (number: 03195011/2020) and Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (number: 88881.505364/2020-01 - edital N° 9/2020).