Determinants of Spike infectivity, processing, and neutralization in SARS-CoV-2 Omicron subvariants BA.1 and BA.2

  1. Chiara Pastorio
  2. Fabian Zech
  3. Sabrina Noettger
  4. Christoph Jung
  5. Timo Jacob
  6. Theo Sanderson
  7. Konstantin M.J. Sparrer
  8. Frank Kirchhoff

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Abstract

No abstract available

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  1. Version published to 10.1016/j.chom.2022.07.006
  2. ScreenIT

    SciScore for 10.1101/2022.04.13.488221: (What is this?)

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

    Table 1: Rigor

    EthicsConsent: Sera from vaccinated individuals: Blood samples of fully BNT162b2 vaccinated individuals were obtained after the participants information and written consent.
    IRB: Ethics approval was provided by the Ethic Committee of Ulm University (vote 99/21–FSt/Sta).
    Sex as a biological variablenot detected.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    Proteins were stained using primary antibodies directed against rabbit anti-V5 (Cell Signaling #13202; 1:1000), VSV-M (Absolute Antibody, 23H12, #Ab01404-2.0; 1:2000), actin (Anti-beta Actin antibody Abcam, ab8227, 1:5000,) and Infrared Dye labeled secondary antibodies (LI-COR IRDye
    anti-V5
    suggested: (Cell Signaling Technology Cat# 13202, RRID:AB_2687461)
    actin
    suggested: (Abcam Cat# ab8227, RRID:AB_2305186)
    Anti-beta Actin
    suggested: (Abcam Cat# ab8227, RRID:AB_2305186)
    ab8227
    suggested: None
    Thereafter, samples were washed with PBS and incubated for 2 h at 4°C with primary antibody (anti-V5(MS) (1:1,000
    MS
    suggested: None
    The samples were washed with PBS/0.1% Tween 20 and incubated in the dark for 2 h at 4°C with the secondary antibody (Alexa Fluor-647-conjugated anti-mouse antibody, 1:1000, Thermo Fisher Scientific) and 500 ng/ml DAPI.
    anti-mouse
    suggested: None
    Experimental Models: Cell Lines
    SentencesResources
    Caco-2 (human epithelial colorectal adenocarcinoma) cells were cultivated in DMEM containing 10% FCS, 2mM glutamine, 100 μg/ml streptomycin and 100 U/ml penicillin, 1mM NEAA supplement.
    Caco-2
    suggested: None
    ACE2 interaction assay: HEK293T cells expressing Spike were collected 48 h after the transfection, washed with phosphate-buffered saline (PBS), lysed in a non-denaturizing lysis buffer.
    HEK293T
    suggested: None
    Recombinant DNA
    SentencesResources
    Expression Constructs: pCG_SARS-CoV-2-Spike-IRES_eGFP encoding the spike protein of SARS-CoV-2 isolate Wuhan-Hu-1(NCBI reference Sequence YP_009724390.1), pCG_SARS-CoV-2-S (1.617), pCG1_SARS-2-S (B.1.1.529) and pCG1_SARS-2-SΔ18 (BA.2) were kindly provided by Stefan Pöhlmann (DPZ Göppingen, Germany).
    pCG_SARS-CoV-2-Spike-IRES_eGFP
    suggested: None
    pCG_SARS-CoV-2-S
    suggested: None
    pCG1_SARS-2-S
    suggested: None
    pCG1_SARS-2-SΔ18
    suggested: None
    pCG_SARS-CoV-2-Spike C-V5-IRES_eGFP was PCR amplified and subcloned into a pCG-IRES_eGFP expression construct using the restriction enzymes XbaI+MluI.
    pCG_SARS-CoV-2-Spike C-V5-IRES_eGFP
    suggested: None
    ACE2 was synthezised by Twist bioscience, PCR amplified, and subcloned into a pCG-IRES_eGFP expression construct using the restriction enzymes XbaI+MluI.
    pCG-IRES_eGFP
    suggested: None
    Software and Algorithms
    SentencesResources
    Twenty-four hours post-transfection, fluorescence microscopy images were acquired using the Cytation 3 microplate reader (BioTek Instruments) and the GFP area was quantified using ImageJ.
    ImageJ
    suggested: (ImageJ, RRID:SCR_003070)
    Statistical analysis: Statistical analyses were performed using GraphPad PRISM 9.2 (GraphPad Software).
    GraphPad PRISM
    suggested: (GraphPad Prism, RRID:SCR_002798)
    GraphPad
    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: We detected the following sentences addressing limitations in the study:
    Limitations of the study: In the present study, we used pseudotyped viral particles instead of replication-competent recombinant SARS-CoV-2 variants, which serves as a proxy to assess infectivity, fusion activity and incorporation. In addition, the impact of many changes might be context-dependent and this might explain why some individual changes had disruptive effects on Hu-1 S function although they are found in Omicron S proteins. It is difficult to predict which of the numerous mutations in the Omicron S might compensate for disruptive mutations. For example, we found that the BA.1 S was less effective in mediating infection than the BA.2 S protein (Figure 2). This agrees, with accumulating evidence that the BA.2 VOC might be more infectious and more virulent than the BA.1 VOC (Suzuki et al., 2022). In addition, we analysed only a limited number of sera from individuals who received a single vaccine regimen (BNT/BNT) and just a few therapeutic antibodies. While further studies are required to fully understand the full consequences of all the complex changes in the Omicron Spike on viral infectivity, tropism, transmission and pathogenesis our results provide first important insights into the functional impact of mutations characteristic for the Omicron VOC Spike that currently dominates the pandemic.

    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 10.1101/2022.04.13.488221 on bioRxiv