A Photoactivable Natural Product with Broad Antiviral Activity against Enveloped Viruses, Including Highly Pathogenic Coronaviruses

  1. Thomas Meunier
  2. Lowiese Desmarets
  3. Simon Bordage
  4. Moussa Bamba
  5. Kévin Hervouet
  6. Yves Rouillé
  7. Nathan François
  8. Marion Decossas
  9. Valentin Sencio
  10. François Trottein
  11. Fézan Honora Tra Bi
  12. Olivier Lambert
  13. Jean Dubuisson
  14. Sandrine Belouzard
  15. Sevser Sahpaz
  16. Karin Séron

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Abstract

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) outbreak has highlighted the need for broad-spectrum antivirals against coronaviruses (CoVs). Here, pheophorbide a (Pba) was identified as a highly active antiviral molecule against human CoV-229E after bioguided fractionation of plant extracts.

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  1. Version published to 10.1128/aac.01581-21
  2. ScreenIT

    SciScore for 10.1101/2021.07.09.451770: (What is this?)

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

    Table 1: Rigor

    EthicsField Sample Permit: Plant collection and extraction: The fifteen plants were collected in the Bafing region (North-West Côte d’Ivoire
    Sex as a biological variablenot detected.
    RandomizationFor each coverslip, a series of six 8-bit images of randomly picked areas were recorded.
    Blindingnot detected.
    Power Analysisnot detected.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    Polyclonal rabbit anti-SARS-CoV-2 nucleocapsid antibodies were from Novus.
    anti-SARS-CoV-2 nucleocapsid antibodies were from Novus .
    suggested: None
    Cyanine 3-conjugated goat anti-mouse IgG and HRP-labeled goat-anti rabbit IgG antibodies were from Jackson Immunoresearch.
    Cyanine 3-conjugated goat anti-mouse IgG
    suggested: None
    anti-mouse IgG
    suggested: None
    Membranes were incubated overnight at 4°C with polyclonal rabbit anti-SARS-CoV-2 nucleocapsid antibodies in 5% (w/v) non-fat dry milk in PBS with 0.1% (v/v) Tween-20.
    anti-SARS-CoV-2 nucleocapsid
    suggested: None
    After being washed 3 times with PBS with 0.1% (v/v) Tween-20, membranes were incubated for 1 h at RT with HRP-labeled goat-anti rabbit IgG antibodies, after which membranes were washed 3 times.
    rabbit IgG
    suggested: None
    They were then rinsed with PBS and processed for immunofluorescence as previously described (56) using primary mouse antibodies specific to HCV E1 (for both HCV and SINV), YFV E, or dsRNA (for CVB4), followed by a cyanine-3-conjugated goat anti-mouse IgG secondary antibody for the detection of infected cells.
    CVB4
    suggested: None
    Twenty-four hours p.t., cells were fixed with 3% paraformaldehyde in PBS for 20 min at RT and syncytia were visualized by immunofluorescence, by incubating the cells with a monoclonal anti-SARS-CoV-2-spike antibody in 10% normal goat serum, followed by incubation with Cyanine-3-conjugated goat anti-mouse IgG antibodies.
    anti-SARS-CoV-2-spike
    suggested: None
    Experimental Models: Cell Lines
    SentencesResources
    Cells and culture conditions: Huh-7, Vero-81 (ATCC number CCL-81) and Vero-E6 were grown in DMEM with glutaMAX-I and 10% FBS in an incubator at 37°C with 5% CO2.
    Huh-7
    suggested: None
    Vero-E6
    suggested: None
    Viruses: The following viral strains were used: HCoV-229E strain VR-740 (ATCC), and a recombinant HCoV-229E-Luc (kind gift of Pr. V. Thiel) (53); SARS-CoV-2 (isolate SARS-CoV-2/human/FRA/Lille_Vero-81-TMPRSS2/2020, NCBI MW575140) was propagated on Vero-81-TMPRSS2 cells.
    Vero-81-TMPRSS2
    suggested: None
    HCoV-229E titers: Huh-7 and Huh-7-TMPRSS2 cells seeded in 24-well plates were inoculated with HCoV-229E at a MOI of 0.5 in the presence of Pba at different concentrations for 1 h at 37°C.
    Huh-7-TMPRSS2
    suggested: None
    Cell-cell fusion assay by transient expression of the SARS-CoV-2 spike protein: Vero-81 cells were seeded on coverslips in 24-wells 16 h before transfection.
    Vero-81
    suggested: None
    Experimental Models: Organisms/Strains
    SentencesResources
    Viruses: The following viral strains were used: HCoV-229E strain VR-740 (ATCC), and a recombinant HCoV-229E-Luc (kind gift of Pr. V. Thiel) (53); SARS-CoV-2 (isolate SARS-CoV-2/human/FRA/Lille_Vero-81-TMPRSS2/2020, NCBI MW575140) was propagated on Vero-81-TMPRSS2 cells.
    HCoV-229E
    suggested: None
    Recombinant DNA
    SentencesResources
    Cells were transfected with 250 ng of a pCDNA3.1(+
    pCDNA3.1
    suggested: RRID:Addgene_79663)
    Software and Algorithms
    SentencesResources
    Images were acquired on an Evos M5000 imaging system (Thermo Fisher Scientic) equipped with light cubes for DAPI, and RFP, and a 10× objective.
    Thermo Fisher Scientic
    suggested: None
    Infected cells and nuclei were automatically counted using macros written in ImageJ.
    ImageJ
    suggested: (ImageJ, RRID:SCR_003070)
    Statistical analysis and IC50 and CC50 calculation: Values were graphed and IC50 calculated by non-linear regression curve fitting with variable slopes constraining the top to 100% and the bottom to 0%, using GraphPad PRISM software.
    GraphPad PRISM
    suggested: (GraphPad Prism, RRID:SCR_002798)

    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.

    Results from scite Reference Check: We found no unreliable references.

    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.

  3. Version published to 10.1101/2021.07.09.451770v1 on bioRxiv