RNA-extraction-free diagnostic method to detect SARS-CoV-2: an assessment from two states, India

Abstract

With increasing demand for large numbers of testing during the coronavirus disease 2019 pandemic, alternative protocols were developed with shortened turn-around time. We evaluated the performance of such a protocol wherein 1138 consecutive clinic attendees were enrolled; 584 and 554 respectively from two independent study sites in the cities of Pune and Kolkata. Paired nasopharyngeal and oropharyngeal swabs were tested by using both reference and index methods in a blinded fashion. Prior to conducting real-time polymerase chain reaction, swabs collected in viral transport medium (VTM) were processed for RNA extraction (reference method) and swabs collected in a dry tube without VTM were incubated in Tris–EDTA–proteinase K buffer for 30 min and heat-inactivated at 98 °C for 6 min (index method). Overall sensitivity and specificity of the index method were 78.9% (95% confidence interval (CI) 71–86) and 99% (95% CI 98–99.6), respectively. Agreement between the index and reference method was 96.8% (k = 0.83, S.E. = 0.03). The reference method exhibited an enhanced detection of viral genes (E, N and RNA-dependent RNA polymerase) with lower Ct values compared to the index method. The index method can be used for detecting severe acute respiratory syndrome corona virus-2 infection with an appropriately chosen primer–probe set and heat treatment approach in pressing time; low sensitivity constrains its potential wider use.