Validation of an extraction-free RT-PCR protocol for detection of SARS-CoV2 RNA

  1. Julianne R Brown
  2. Laura Atkinson
  3. Divya Shah
  4. Kathryn Harris

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Abstract

In light of supply chain failures for reagents and consumables needed for purification of nucleic acid for detection of SARS-CoV-2 RNA by RT-PCR, we aim to verify the performance and utility of a non-extraction protocol for RT-PCR ("direct RT-PCR"). We report improved sensitivity compared to earlier reports of direct RT-PCR testing of swab samples, in particular at the lower limit of detection (sensitivity 93% overall; 100% for specimens with high to moderate viral titre, Ct <34; 81% for specimens with a low viral titre, Ct ≥34). Sensitivity is improved (from 90 to 93%) by testing in duplicate. We recommend swabs are re-suspended in water to minimise PCR inhibition. A cellular target is necessary to control for PCR inhibition and specimen quality. Direct RT-PCR is best suited to population level screening where results are not clinically actionable, however in the event of a critical supply chain failure direct RT-PCR is fit for purpose for the detection of SARS-CoV-2 infection. The results from our study offer front-line laboratories additional reagent options for performing extraction-free RT-PCR protocols.

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  1. ScreenIT

    SciScore for 10.1101/2020.04.29.20085910: (What is this?)

    Please note, not all rigor criteria are appropriate for all manuscripts.

    Table 1: Rigor

    Institutional Review Board Statementnot detected.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.

    Table 2: Resources

    No key resources detected.

    Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).

    Results from LimitationRecognizer: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.

    Results from TrialIdentifier: No clinical trial numbers were referenced.

    Results from Barzooka: We did not find any issues relating to the usage of bar graphs.

    Results from JetFighter: We did not find any issues relating to colormaps.

    Results from rtransparent:
    • Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
    • Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
    • No protocol registration statement was detected.

    About SciScore

    SciScore is an automated tool that is designed to assist expert reviewers by finding and presenting formulaic information scattered throughout a paper in a standard, easy to digest format. SciScore checks for the presence and correctness of RRIDs (research resource identifiers), and for rigor criteria such as sex and investigator blinding. For details on the theoretical underpinning of rigor criteria and the tools shown here, including references cited, please follow this link.

  2. Version published to 10.1101/2020.04.29.20085910 on medRxiv