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

PLoS One. 2020 Jul 30;15(7):e0236859. doi: 10.1371/journal.pone.0236859. eCollection 2020.

Abstract

Background: 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.

Objective: We undertook this study to evaluate a novel protocol of pooling of RNA samples/elutes in performance of PCR for SARS CoV-2 virus.

Study design: 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.

Results: 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.

Conclusion: 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.

Publication types

  • Evaluation Study

MeSH terms

  • Betacoronavirus / genetics*
  • COVID-19
  • Coronavirus Envelope Proteins
  • Coronavirus Infections / diagnosis*
  • Coronavirus Infections / epidemiology*
  • Coronavirus Infections / virology
  • Diagnostic Tests, Routine / economics
  • Diagnostic Tests, Routine / methods
  • Humans
  • India / epidemiology
  • Mass Screening / economics
  • Mass Screening / methods
  • Pandemics
  • Pneumonia, Viral / diagnosis*
  • Pneumonia, Viral / epidemiology*
  • Pneumonia, Viral / virology
  • Prospective Studies
  • RNA, Viral / genetics*
  • RNA, Viral / isolation & purification
  • Real-Time Polymerase Chain Reaction / economics
  • Real-Time Polymerase Chain Reaction / methods*
  • Reproducibility of Results
  • Reverse Transcriptase Polymerase Chain Reaction / economics
  • Reverse Transcriptase Polymerase Chain Reaction / methods*
  • SARS-CoV-2
  • Viral Envelope Proteins / genetics

Substances

  • Coronavirus Envelope Proteins
  • RNA, Viral
  • Viral Envelope Proteins
  • envelope protein, SARS-CoV-2

Grants and funding

The author(s) received no specific funding for this work.