Naturally enhanced neutralizing breadth against SARS-CoV-2 one year after infection

  1. Zijun Wang
  2. Frauke Muecksch
  3. Dennis Schaefer-Babajew
  4. Shlomo Finkin
  5. Charlotte Viant
  6. Christian Gaebler
  7. Hans- Heinrich Hoffmann
  8. Christopher O. Barnes
  9. Melissa Cipolla
  10. Victor Ramos
  11. Thiago Y. Oliveira
  12. Alice Cho
  13. Fabian Schmidt
  14. Justin Da Silva
  15. Eva Bednarski
  16. Lauren Aguado
  17. Jim Yee
  18. Mridushi Daga
  19. Martina Turroja
  20. Katrina G. Millard
  21. Mila Jankovic
  22. Anna Gazumyan
  23. Zhen Zhao
  24. Charles M. Rice
  25. Paul D. Bieniasz
  26. Marina Caskey
  27. Theodora Hatziioannou
  28. Michel C. Nussenzweig

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Abstract

More than one year after its inception, the coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) remains difficult to control despite the availability of several working vaccines. Progress in controlling the pandemic is slowed by the emergence of variants that appear to be more transmissible and more resistant to antibodies 1,2 . Here we report on a cohort of 63 individuals who have recovered from COVID-19 assessed at 1.3, 6.2 and 12 months after SARS-CoV-2 infection, 41% of whom also received mRNA vaccines 3,4 . In the absence of vaccination, antibody reactivity to the receptor binding domain (RBD) of SARS-CoV-2, neutralizing activity and the number of RBD-specific memory B cells remain relatively stable between 6 and 12 months after infection. Vaccination increases all components of the humoral response and, as expected, results in serum neutralizing activities against variants of concern similar to or greater than the neutralizing activity against the original Wuhan Hu-1 strain achieved by vaccination of naive individuals 2,5–8 . The mechanism underlying these broad-based responses involves ongoing antibody somatic mutation, memory B cell clonal turnover and development of monoclonal antibodies that are exceptionally resistant to SARS-CoV-2 RBD mutations, including those found in the variants of concern 4,9 . In addition, B cell clones expressing broad and potent antibodies are selectively retained in the repertoire over time and expand markedly after vaccination. The data suggest that immunity in convalescent individuals will be very long lasting and that convalescent individuals who receive available mRNA vaccines will produce antibodies and memory B cells that should be protective against circulating SARS-CoV-2 variants.

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  1. Version published to 10.1038/s41586-021-03696-9
  2. ScreenIT

    SciScore for 10.1101/2021.05.07.443175: (What is this?)

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

    Table 1: Rigor

    EthicsConsent: All participants at Rockefeller University provided written informed consent before participation in the study and the study was conducted in accordance with Good Clinical Practice.
    Sex as a biological variablenot detected.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    Plates were washed 6 times with washing buffer and then incubated with anti-human IgG, IgM or IgA secondary antibody conjugated to horseradish peroxidase (HRP) (Jackson Immuno Research
    anti-human IgG
    suggested: None
    IgA
    suggested: None
    The half-maximal neutralization titers for plasma (NT50) or half-maximal and 90% inhibitory concentrations for monoclonal antibodies (IC50 and IC90) were determined using four-parameter nonlinear regression (least squares regression method without weighting; constraints: top=1, bottom=0) (GraphPad Prism).
    IC90
    suggested: None
    The enriched B cells were incubated in FACS buffer (1× PBS, 2% FCS, 1 mM EDTA) with the following anti-human antibodies (all at 1:200 dilution): anti-CD20-PECy7 (BD Biosciences, 335793), anti-CD3-APC-eFluro 780 (Invitrogen, 47-0037-41)
    anti-human
    suggested: (GenWay Biotech Inc. Cat# 18-202-335793-0.1 mg, RRID:AB_1981874)
    anti-CD20-PECy7
    suggested: None
    anti-CD3-APC-eFluro 780
    suggested: None
    For B cell phenotype analysis, in addition to above antibodies, B cells were also stained with following anti-human antibodies: anti-IgD-BV421 (Biolegend, 348226), anti-CD27-FITC (BD biosciences, 555440), anti-CD19-BV605 (Biolegend, 302244), anti-CD71-PerCP-Cy5.5 (Biolegend, 334114), anti-IgG-PECF594 (BD biosciences, 562538), anti-IgM-AF700 (Biolegend, 314538), anti-IgA-Viogreen (Miltenyi Biotec, 130-113-481)
    anti-human antibodies: anti-IgD-BV421 ( Biolegend , 348226
    suggested: None
    anti-CD27-FITC
    suggested: (Millipore Cat# FCMAB191F, RRID:AB_10807274)
    anti-CD19-BV605
    suggested: None
    anti-CD71-PerCP-Cy5.5
    suggested: None
    anti-IgG-PECF594
    suggested: None
    anti-IgM-AF700 ( Biolegend , 314538) , anti-IgA-Viogreen ( Miltenyi Biotec , 130-113-481)
    suggested: None
    The frequency distributions of human V genes in anti-SARS-CoV-2 antibodies from this study was compared to 131,284,220 IgH and IgL sequences generated by 45 and downloaded from cAb-Rep46, a database of human shared BCR clonotypes available at https://cab-rep.c2b2.columbia.edu/.
    anti-SARS-CoV-2
    suggested: None
    Experimental Models: Cell Lines
    SentencesResources
    Briefly, 293T cells were transfected with pNL4-3ΔEnv-nanoluc and pSARS-CoV-2-SΔ19, particles were harvested 48 hpt, filtered and stored at -80°C.
    293T
    suggested: RRID:CVCL_H376)
    Recombinant DNA
    SentencesResources
    Briefly, 293T cells were transfected with pNL4-3ΔEnv-nanoluc and pSARS-CoV-2-SΔ19, particles were harvested 48 hpt, filtered and stored at -80°C.
    pNL4-3ΔEnv-nanoluc
    suggested: None
    pSARS-CoV-2-SΔ19
    suggested: None
    Software and Algorithms
    SentencesResources
    The average of its signal was used for normalization of all of the other values on the same plate with Excel software before calculating the area under the curve using Prism V9.1(GraphPad).
    Excel
    suggested: None
    Prism
    suggested: (PRISM, RRID:SCR_005375)
    For monoclonal antibodies, the EC50 was determined using four-parameter nonlinear regression (GraphPad Prism V9.1).
    GraphPad Prism
    suggested: (GraphPad Prism, RRID:SCR_002798)
    Proteins: Mammalian expression vectors encoding the RBDs of SARS-CoV-2 (GenBank MN985325.1; S protein residues 319-539) or K417N, E484K, N501Y RBD mutants with an N-terminal human IL-2 or Mu phosphatase signal peptide were previously described 43.
    Proteins
    suggested: None
    The half-maximal neutralization titers for plasma (NT50) or half-maximal and 90% inhibitory concentrations for monoclonal antibodies (IC50 and IC90) were determined using four-parameter nonlinear regression (least squares regression method without weighting; constraints: top=1, bottom=0) (GraphPad Prism).
    GraphPad
    suggested: (GraphPad Prism, RRID:SCR_002798)
    Single CD3-CD8-CD14-CD16-CD20+Ova-RBD-PE+RBD-AF647+ B cells were sorted into individual wells of 96-well plates containing 4 μl of lysis buffer (0.5× PBS, 10 mM DTT, 3,000 units/ml RNasin Ribonuclease Inhibitors (Promega, N2615) per well using a FACS Aria III and FACSDiva software (Becton Dickinson) for acquisition and FlowJo for analysis.
    FACSDiva
    suggested: (BD FACSDiva Software, RRID:SCR_001456)
    FlowJo
    suggested: (FlowJo, RRID:SCR_008520)
    For B cell phenotype analysis, in addition to above antibodies, B cells were also stained with following anti-human antibodies: anti-IgD-BV421 (Biolegend, 348226), anti-CD27-FITC (BD biosciences, 555440), anti-CD19-BV605 (Biolegend, 302244), anti-CD71-PerCP-Cy5.5 (Biolegend, 334114), anti-IgG-PECF594 (BD biosciences, 562538), anti-IgM-AF700 (Biolegend, 314538), anti-IgA-Viogreen (Miltenyi Biotec, 130-113-481)
    Biolegend , 314538) , anti-IgA-Viogreen ( Miltenyi Biotec
    suggested: None
    Miltenyi
    suggested: (Miltenyi Biotec, RRID:SCR_008984)
    Sequence analysis was performed using MacVector.
    MacVector
    suggested: (MacVector, RRID:SCR_015700)

    Results from OddPub: Thank you for sharing your code and 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

    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 10.1101/2021.05.07.443175 on bioRxiv