Conventional reverse transcription-polymerase chain reaction assay as an alternative, low-cost, and reliable method for the detection of COVID-19

Abstract

Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a highly contagious infection causing a large number of deaths in susceptible individuals throughout the world. Objectives: In this study, a low-cost, sensitive, and easy-to-perform conventional Polymerase chain reaction (PCR)-based RNA detection method was evaluated to diagnose the infection that was feasible at a laboratory with minimal molecular infrastructure. Methods: From 4 July to 31 August 2020, a total of 277 nasopharyngeal/oropharyngeal swab samples consisting of 72 samples from hospitalized patients with a severe respiratory infection and 205 suspected patients into four different age groups of under 20 years (n=8), 20-39 years (n=90), 40-59 years (n=93), and over 60 years (n=86) in Isfahan, Iran, were tested using the real-time reverse transcription polymerase chain reaction (rRT-PCR), and conventional PCR assays. Results: Overall, 40 cases out of 72 hospitalized patients were aged more than 60 years, and the majority of the study population were male (54.1%, n=150). Out of 160 clinical samples tested by RT-PCR with the E gene, the sensitivity and specificity of the conventional PCR method were determined at 100%. Furthermore, out of 117 clinical samples evaluated by the probe-based RT-PCR with the N gene, 74.4% of the samples were positive. Moreover, the duplex PCR method using the N gene and RNase P as an internal control reference gene showed that 68.4% of the samples were positive. Therefore, the tested PCRs could detect positive samples with a sensitivity of 92.55% and a specificity of 100%. Conclusion: According to the results, this method is a simple, inexpensive, and valuable alternative as well as a suitable procedure for the laboratory diagnosis of SARS-CoV-2 infection.