Inactivation analysis of SARS-CoV-2 by specimen transport media, nucleic acid extraction reagents, detergents and fixatives

  1. Stephen R. Welch
  2. Katherine A. Davies
  3. Hubert Buczkowski
  4. Nipunadi Hettiarachchi
  5. Nicole Green
  6. Ulrike Arnold
  7. Matthew Jones
  8. Matthew J. Hannah
  9. Reah Evans
  10. Christopher Burton
  11. Jane E. Burton
  12. Malcolm Guiver
  13. Patricia A. Cane
  14. Neil Woodford
  15. Christine B. Bruce
  16. Allen D. G. Roberts
  17. Marian J. Killip

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Abstract

The COVID-19 pandemic has necessitated a rapid multi-faceted response by the scientific community, bringing researchers, health officials and industry together to address the ongoing public health emergency. To meet this challenge, participants need an informed approach for working safely with the etiological agent, the novel human coronavirus SARS-CoV-2. Work with infectious SARS-CoV-2 is currently restricted to high-containment laboratories, but material can be handled at a lower containment level after inactivation. Given the wide array of inactivation reagents that are being used in laboratories during this pandemic, it is vital that their effectiveness is thoroughly investigated. Here, we evaluated a total of 23 commercial reagents designed for clinical sample transportation, nucleic acid extraction and virus inactivation for their ability to inactivate SARS-CoV-2, as well as seven other common chemicals including detergents and fixatives. As part of this study, we have also tested five filtration matrices for their effectiveness at removing the cytotoxic elements of each reagent, permitting accurate determination of levels of infectious virus remaining following treatment. In addition to providing critical data informing inactivation methods and risk assessments for diagnostic and research laboratories working with SARS-CoV-2, these data provide a framework for other laboratories to validate their inactivation processes and to guide similar studies for other pathogens.

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  1. ScreenIT

    SciScore for 10.1101/2020.07.08.194613: (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 AnalysisThe fixative was removed, and monolayers washed three times with PBS before scraping cells into 1mL MEM/5% FBS and sonicated (3 × 10 second on,10 seconds off at 100% power and amplitude) using a UP200St with VialTweeter attachment (Hielscher Ultrasound Technology).
    Sex as a biological variablenot detected.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Experimental Models: Cell Lines
    SentencesResources
    Cells and virus: Vero E6 cells (Vero C1008; ATCC CRL-1586) were cultured in modified Eagle’s minimum essential medium (MEM) supplemented with 10% (v/v) fetal calf serum (FCS).
    Vero
    suggested: ATCC Cat# CRL-1586, RRID:CVCL_0574)
    For TCID50s, ten-fold dilutions of virus stock (25μL) were plated onto 96-well plates containing Vero E6 cell suspension (2.5 × 104 cells/well in 100μl
    Vero E6
    suggested: None
    Software and Algorithms
    SentencesResources
    Zymo Research); guanidine hydrochloride (GCHl) and guanidine thiocyanate (GITC) buffers containing Triton X-100 (both Oxoid/Thermo Fisher); Virus Transport and Preservation Medium Inactivated (BioComma)
    BioComma
    suggested: None
    All analyses were performed using GraphPad Prism 8 (v8.4.1, GraphPad Software). 2.4.
    GraphPad Prism
    suggested: (GraphPad Prism, RRID:SCR_002798)
    GraphPad
    suggested: (GraphPad Prism, RRID:SCR_002798)
    Nucleic acid was extracted from cell culture media manually using a QIAamp Viral RNA Mini Kit (QIAGEN) or using NucliSENS easyMAG or EMAG platforms (both BioMérieux).
    BioMérieux
    suggested: None

    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:
    • No conflict of interest statement was detected. If there are no conflicts, we encourage authors to explicit state so.
    • No funding statement was detected.
    • 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 10.1101/2020.07.08.194613 on bioRxiv