Journal Article

Pooled RNA sample reverse transcriptase real time PCR assay for SARS CoV-2 infection: A reliable, faster and economical method

Dubey, Shantanu, Kumar, Guresh, Padhi, Abhishek, Khodare, Arvind, Ramachandran, Krithiga, Kilikdar, Mousumi, Sarin, Shiv Kumar, Gupta, Ekta, Mehta, Vibha, Agarwal, Reshu

2020

Overview

Corona virus disease 2019 (COVID-19) which initially started as a cluster of pneumonia cases in the Wuhan city of China has now become a full-blown pandemic. Timely diagnosis of COVID-19 is the key in containing the pandemic and breaking the chain of transmission. In low- and middle-income countries availability of testing kits has become the major bottleneck in testing. Novel methods like pooling of samples are the need of the hour. We undertook this study to evaluate a novel protocol of pooling of RNA samples/elutes in performance of PCR for SARS CoV-2 virus. Extracted RNA samples were randomly placed in pools of 8 on a 96 well plate. Both individual RNA (ID) and pooled RNA RT-qPCR for the screening E gene were done in the same plate and the positivity for the E gene was seen. The present study demonstrated that pool testing with RNA samples can easily detect even up to a single positive sample with Ct value as high as 38. The present study also showed that the results of pool testing is not affected by number of positive samples in a pool. Pooling of RNA samples can reduce the time and expense, and can help expand diagnostic capabilities, especially during constrained supply of reagents and PCR kits for the diagnosis of SARS-CoV-2 infection.

• Publisher: Public Library of Science,Public Library of Science (PLoS)
• Subject: Betacoronavirus - genetics / Biology and Life Sciences / Corona / Coronavirus Envelope Proteins / Coronavirus Infections - diagnosis / Coronavirus Infections - epidemiology / Coronavirus Infections - virology / Coronaviruses / COVID-19 / Diagnosis / Diagnostic systems / Diagnostic Tests, Routine - economics / Diagnostic Tests, Routine - methods / E protein / Ethics / Humans / India - epidemiology / Infections / Laboratories / Liver / Mass Screening - economics / Mass Screening - methods / Medicine and Health Sciences / Pandemics / Pneumonia, Viral - diagnosis / Pneumonia, Viral - epidemiology / Pneumonia, Viral - virology / Prospective Studies / Reagents / Real time / Real-Time Polymerase Chain Reaction - economics / Real-Time Polymerase Chain Reaction - methods / Reproducibility of Results / Research and Analysis Methods / Reverse Transcriptase Polymerase Chain Reaction - economics / Reverse Transcriptase Polymerase Chain Reaction - methods / Ribonucleic acid / RNA / RNA viruses / RNA, Viral - genetics / RNA, Viral - isolation & purification / RNA-directed DNA polymerase / SARS-CoV-2 / Severe acute respiratory syndrome / Severe acute respiratory syndrome coronavirus 2 / Viral diseases / Viral Envelope Proteins - genetics / Virology / Viruses