Differences in neuroinflammation in the olfactory bulb between D614G, Delta and Omicron BA.1 SARS-CoV-2 variants in the hamster model

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Abstract

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is associated with various neurological complications. SARS-CoV-2 infection induces neuroinflammation in the central nervous system (CNS), whereat the olfactory bulb seems to be involved most frequently. Here we show differences in the neuroinvasiveness and neurovirulence among SARS-CoV-2 variants in the hamster model five days post inoculation. Replication in the olfactory mucosa was observed in all hamsters, but most prominent in D614 inoculated hamsters. We observed neuroinvasion into the CNS via the olfactory nerve in D614G-, but not Delta (B.1.617.2)- or Omicron BA.1 (B.1.1.529) inoculated hamsters. Neuroinvasion was associated with neuroinflammation in the olfactory bulb of hamsters inoculated with D614G but hardly in Delta or Omicron BA.1. Altogether, this indicates that there are differences in the neuroinvasive and neurovirulent potential among SARS-CoV-2 variants in the acute phase of the infection in the hamster model.

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  1. SciScore for 10.1101/2022.03.24.485596: (What is this?)

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

    Table 1: Rigor

    EthicsField Sample Permit: Research was conducted under a project license from the Dutch competent authority and the study protocol (#17-4312) was approved by the institutional Animal Welfare Body.
    Euthanasia Agents: Hamsters were euthanized by cardiac puncture under isoflurane anesthesia and cervical dislocation.
    Sex as a biological variableAnimals and experimental setup: Female Syrian golden hamsters (Mesocricetus auratus; 6 weeks old; Janvier, France) were handled in an ABSL-3 biocontainment laboratory.
    RandomizationFor unbiased experiments, all animals were randomly assigned to experimental groups.
    BlindingCounting of IBA-1+ cells was determined by manual counting by two independent blinded observes.
    Power Analysisnot detected.
    Cell Line AuthenticationContamination: Both cells were grown at 37°C in a humidified CO2◻incubator and routinely tested for mycoplasma.

    Table 2: Resources

    Antibodies
    SentencesResources
    Cells were incubated in 3% BSA (bovine serum albumin; Sigma) in PBS and stained with a rabbit anti-nucleocapsid antibody (Sino biological; 1:2000) in PBS containing 0.1% BSA, washed thrice in PBS, and stained with donkey anti-rabbit Alexa Fluor 488 (Invitrogen; 1:4000) in PBS containing 0.1% BSA.
    anti-nucleocapsid
    suggested: None
    anti-rabbit
    suggested: None
    Slides were then incubated with a rabbit polyclonal antibody against SARS-CoV/SARS-CoV-2-nucleoprotein (40143-T62, Sino Biological, Pennsylvania, USA) (1:1000), mouse CD3 (ab16669, Abcam, Cambridge, UK) (1:10, 20μg/ml), human Iba-1 (019-19741
    SARS-CoV/SARS-CoV-2-nucleoprotein ( 40143-T62 , Sino Biological , Pennsylvania , USA )
    suggested: None
    CD3
    suggested: (Abcam Cat# ab16669, RRID:AB_443425)
    Experimental Models: Cell Lines
    SentencesResources
    Drosten) was propagated to passage three on Vero E6 cells in Opti-MEM I (1X) + GlutaMAX (Gibco), supplemented with penicillin (10,000 IU/mL) and streptomycin (10,000 IU/mL).
    Vero E6
    suggested: None
    Virus Titrations: Ten-fold serial diluted samples were added to monolayers of Calu-3 cells.
    Calu-3
    suggested: KCLB Cat# 30055, RRID:CVCL_0609)
    Software and Algorithms
    SentencesResources
    Plaque assay analysis was performed using ImageQuant TL 8.2 software (GE Healthcare).
    ImageQuant
    suggested: (ImageQuant, RRID:SCR_014246)
    Images were taken with a 200x magnification (20x air objective; Olympus) with CellSens software.
    CellSens
    suggested: None
    Graphs and statistical tests were made with GraphPad Prism version 9.
    GraphPad Prism
    suggested: (GraphPad Prism, RRID:SCR_002798)
    Figures were prepared with Adobe Illustrator CC2019
    Adobe Illustrator
    suggested: (Adobe Illustrator, RRID:SCR_010279)
    , Adobe Photoshop CC2019 and Biorender.
    Adobe Photoshop
    suggested: (Adobe Photoshop, RRID:SCR_014199)

    Results from OddPub: Thank you for sharing your data.


    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

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