Real-time RT-PCR for COVID-19 diagnosis: challenges and prospects

COVID-19 impacts global public health, economy, education, tourism/hospitality and sports; rapid and accurate testing of clinical samples dictate effective response. So far, the real-time reverse transcriptase-polymerase chain reaction (RT-qPCR) is the assay of choice for COVID-19 diagnosis considering its rapidity and accuracy in informing on active coronavirus (CoV) infection. Presently, several RT-qPCR protocols with differing sensitivity/specificity are used for performing this assay; some of them are known to have generated debatable test results to constitute challenges worthy of consideration. This review provides a critical assessment of various published works on RT-qPCR assays used for COVID-19 diagnosis with their different indicators of positivity i.e., cycle threshold (Ct) cut-off values. Knowledge of diagnostic tests for COVID-19 is still evolving and, as a prospect, underscores the need for local validation of positive-negative Ct cut-off values when establishing RT-qPCR assays for SARS-CoV-2 detection.