Massive and rapid COVID-19 testing is feasible by extraction-free SARS-CoV-2 RT-PCR

Nat Commun. 2020 Sep 23;11(1):4812. doi: 10.1038/s41467-020-18611-5.

Abstract

Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is commonly diagnosed by reverse transcription polymerase chain reaction (RT-PCR) to detect viral RNA in patient samples, but RNA extraction constitutes a major bottleneck in current testing. Methodological simplification could increase diagnostic availability and efficiency, benefitting patient care and infection control. Here, we describe methods circumventing RNA extraction in COVID-19 testing by performing RT-PCR directly on heat-inactivated or lysed samples. Our data, including benchmarking using 597 clinical patient samples and a standardised diagnostic system, demonstrate that direct RT-PCR is viable option to extraction-based tests. Using controlled amounts of active SARS-CoV-2, we confirm effectiveness of heat inactivation by plaque assay and evaluate various generic buffers as transport medium for direct RT-PCR. Significant savings in time and cost are achieved through RNA-extraction-free protocols that are directly compatible with established PCR-based testing pipelines. This could aid expansion of COVID-19 testing.

Publication types

  • Comparative Study
  • Evaluation Study
  • Research Support, Non-U.S. Gov't
  • Validation Study

MeSH terms

  • Benchmarking
  • Betacoronavirus / genetics*
  • Betacoronavirus / isolation & purification*
  • COVID-19
  • COVID-19 Testing
  • Clinical Laboratory Techniques / methods*
  • Clinical Laboratory Techniques / standards
  • Clinical Laboratory Techniques / statistics & numerical data
  • Coronavirus Infections / diagnosis*
  • Coronavirus Infections / epidemiology
  • Coronavirus Infections / virology*
  • DNA Primers / genetics
  • Hot Temperature
  • Humans
  • Pandemics
  • Pneumonia, Viral / diagnosis*
  • Pneumonia, Viral / epidemiology
  • Pneumonia, Viral / virology*
  • RNA, Viral / genetics
  • RNA, Viral / isolation & purification
  • Reverse Transcriptase Polymerase Chain Reaction / methods*
  • Reverse Transcriptase Polymerase Chain Reaction / standards
  • Reverse Transcriptase Polymerase Chain Reaction / statistics & numerical data
  • SARS-CoV-2
  • Sensitivity and Specificity
  • Sweden / epidemiology
  • Viral Plaque Assay / methods

Substances

  • DNA Primers
  • RNA, Viral