Utility of Nucleic Acid Extraction Free COVID-19 Real Time PCR Protocol in Resource Limited Setting: A Pilot Study DC01-DC03
Professor and Head, Department of Microbiology, AFMC, Pune, Maharashtra, India.
Introduction: Coronavirus Disease-19 (COVID-19), a respiratory infection, caused by Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2), was first identified in Wuhan, Hubei province, China in December 2019. Alarming increase in the number of cases has put tremendous pressure on existing health resources. Real Time Reverse Transcriptase Polymerase Chain Reaction (rRT-PCR), a molecular diagnostic method, is considered gold standard for diagnosis of SARSCoV-2 infection. It involves Ribonucleic Acid (RNA) extraction as the preliminary step. Innovations to cut down cost and time involved in SARS-CoV-2 testing are need of hour.
Aim: To assess the feasibility of Nucleic Acid Extraction Free (NEF) protocol for COVID-19 diagnosis in resource limited settings.
Materials and Methods: In this pilot study, a panel of 148 Nasopharyngeal (NP) samples was subjected to the novel NEF RT-PCR protocol and results were compared to gold standard RT-PCR on RNA extracted from NP specimen. The cycle threshold value (Ct value) for each target was tabulated in MS Excel Spreadsheet and data analysis was performed using Statistical Package for Social Sciences (SPSS) software version 15.0.
Results: Out of 148 collected samples, 120 showed amplification of E and RNA polymerase gene (RdRp) targets by RNA extractionbased RT-PCR. Overall sensitivity and specificity observed for NEF protocol was 43.94% and 96.42%, respectively.
Conclusion: Further refinement in the protocol would be required to improve the sensitivity of NEF protocol and widespread use in laboratories.