Molecular (real-time reverse transcription polymerase chain reaction) diagnosis of SARS-CoV-2 infections: Complexity and challenges

Abstract

The outbreak of coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was first recorded in Wuhan, China. The World Health Organization initially classified COVID-19 as a public health emergency and subsequently declared the disease a global pandemic. COVID-19 can take at least three distinct forms: severe acute distress syndrome with a potentially fatal outcome, mild respiratory illness (pneumonia with eventual recovery) and asymptomatic infection. All three disease forms have the potential to transmit the infection to healthy contacts. At present, real-time reverse transcription polymerase chain reaction (RT-PCR) is the only available laboratory tool to confirm the presence of viral RNA in patient specimens. These assays are designed to detect one or more (at least 2) SARS-CoV-2 RNA gene targets allowing the detection of the virus. Commercially available RT-PCR assays employ various gene targets of the viral genome in their assay systems. Additionally, there are differences in primer selection for the same gene region of SARS-CoV-2. At present, it is unclear whether the results from different RT-PCR assays are comparable in detecting the spectrum of COVID-19 manifestations. The purpose of the present article is twofold: first, to briefly focus on the findings of these reports; and second, to emphasize the various challenges and flaws that can potentially impact the diagnostic accuracy of RT-PCR testing for SARS-CoV-2.