Direct diagnostic testing of SARS-CoV-2 without the need for prior RNA extraction

Sci Rep. 2021 Jan 28;11(1):2402. doi: 10.1038/s41598-021-81487-y.

Abstract

The COVID-19 pandemic has resulted in an urgent need for a rapid, point of care diagnostic testing that could be rapidly scaled on a worldwide level. We developed and tested a highly sensitive and robust assay based on reverse transcription loop mediated isothermal amplification (RT-LAMP) that uses readily available reagents and a simple heat block using contrived spike-in and actual clinical samples. RT-LAMP testing on RNA-spiked samples showed a limit of detection (LoD) of 2.5 copies/μl of viral transport media. RT-LAMP testing directly on clinical nasopharyngeal swab samples in viral transport media had an 85% positive percentage agreement (PPA) (17/20), and 100% negative percentage agreement (NPV) and delivered results in 30 min. Our optimized RT-LAMP based testing method is a scalable system that is sufficiently sensitive and robust to test for SARS-CoV-2 directly on clinical nasopharyngeal swab samples in viral transport media in 30 min at the point of care without the need for specialized or proprietary equipment or reagents. This cost-effective and efficient one-step testing method can be readily available for COVID-19 testing world-wide, especially in resource poor settings.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • COVID-19 / diagnosis*
  • COVID-19 Testing / methods*
  • Clinical Laboratory Techniques / methods
  • Diagnostic Techniques and Procedures
  • Diagnostic Tests, Routine
  • Humans
  • Limit of Detection
  • Molecular Diagnostic Techniques / methods*
  • Nucleic Acid Amplification Techniques / methods*
  • Point-of-Care Testing
  • RNA, Viral / genetics
  • RNA, Viral / isolation & purification*
  • Reverse Transcription
  • SARS-CoV-2 / chemistry
  • SARS-CoV-2 / genetics*
  • SARS-CoV-2 / metabolism
  • Sensitivity and Specificity

Substances

  • RNA, Viral

Supplementary concepts

  • LAMP assay