Germinal centre-driven maturation of B cell response to SARS-CoV-2 vaccination

  1. Wooseob Kim
  2. Julian Q. Zhou
  3. Alexandria J. Sturtz
  4. Stephen C. Horvath
  5. Aaron J. Schmitz
  6. Tingting Lei
  7. Elizaveta Kalaidina
  8. Mahima Thapa
  9. Wafaa B. Alsoussi
  10. Alem Haile
  11. Michael K. Klebert
  12. Teresa Suessen
  13. Luis Parra-Rodriguez
  14. Philip A. Mudd
  15. William D. Middleton
  16. Sharlene A. Teefey
  17. Iskra Pusic
  18. Jane A. O’Halloran
  19. Rachel M. Presti
  20. Jackson S. Turner
  21. Ali H. Ellebedy

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Abstract

Germinal centres (GC) are lymphoid structures where vaccine-responding B cells acquire affinity-enhancing somatic hypermutations (SHM), with surviving clones differentiating into memory B cells (MBCs) and long-lived bone marrow plasma cells (BMPCs) 1–4 . Induction of the latter is a hallmark of durable immunity after vaccination 5 . SARS-CoV-2 mRNA vaccination induces a robust GC response in humans 6–8 , but the maturation dynamics of GC B cells and propagation of their progeny throughout the B cell diaspora have not been elucidated. Here we show that anti-SARS-CoV-2 spike (S)-binding GC B cells were detectable in draining lymph nodes for at least six months in 10 out of 15 individuals who had received two doses of BNT162b2, a SARS-CoV-2 mRNA vaccine. Six months after vaccination, circulating S-binding MBCs were detected in all participants (n=42) and S-specific IgG-secreting BMPCs were detected in 9 out of 11 participants. Using a combined approach of single-cell RNA sequencing of responding blood and lymph node B cells from eight participants and expression of the corresponding monoclonal antibodies, we tracked the evolution of 1540 S-specific B cell clones. SHM accumulated along the B cell differentiation trajectory, with early blood plasmablasts showing the lowest frequencies, followed by MBCs and lymph node plasma cells whose SHM largely overlapped with GC B cells. By three months after vaccination, the frequency of SHM within GC B cells had doubled. Strikingly, S + BMPCs detected six months after vaccination accumulated the highest level of SHM, corresponding with significantly enhanced anti-S polyclonal antibody avidity in blood at that time point. This study documents the induction of affinity-matured BMPCs after two doses of SARS-CoV-2 mRNA vaccination in humans, providing a foundation for the sustained high efficacy observed with these vaccines.

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  1. ScreenIT

    SciScore for 10.1101/2021.10.31.466651: (What is this?)

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

    Table 1: Rigor

    EthicsField Sample Permit: Sample collection, preparation, and storage: All studies were approved by the Institutional Review Board of Washington University in St Louis.
    IRB: Sample collection, preparation, and storage: All studies were approved by the Institutional Review Board of Washington University in St Louis.
    Consent: Written consent was obtained from all participants.
    Sex as a biological variablenot detected.
    RandomizationFor selection, where a clone spanned both the GC and LNPC compartments, and/or multiple time points, a compartment and a timepoint were first randomly selected.
    Blindingnot detected.
    Power Analysisnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    HRP-conjugated goat anti-human IgG (H+L) antibody (Jackson ImmunoResearch, 109-035-088, 1:2500) was used to detect monoclonal antibodies.
    anti-human IgG
    suggested: (Jackson ImmunoResearch Labs Cat# 109-035-088, RRID:AB_2337584)
    HRP-conjugated goat anti-Human IgG Fcγ fragment (Jackson ImmunoResearch, 109-035-190, 1:1500), HRP-conjugated goat anti-human serum IgA α chain (Jackson ImmunoResearch, 109-035-011, 1:2500), and HRP-conjugated goat anti-human IgM (Caltag, H15007, 1:4000) were used to detect plasma antibodies.
    anti-human serum IgA α chain
    suggested: (Jackson ImmunoResearch Labs Cat# 109-035-011, RRID:AB_2337580)
    anti-human IgM (Caltag, H15007
    suggested: None
    Recombinant DNA
    SentencesResources
    In brief, a mammalian cell codon-optimized nucleotide sequences coding for the soluble version of S (GenBank: MN908947.3, amino acids 1-1,213) including a C-terminal thrombin cleavage site, T4 fold trimerization domain and hexahistidine tag was cloned into the mammalian expression vector pCAGGS.
    pCAGGS
    suggested: RRID:Addgene_18926)
    Software and Algorithms
    SentencesResources
    Flow cytometry data were analyzed using FlowJo v.
    FlowJo
    suggested: (FlowJo, RRID:SCR_008520)
    V(D)J gene annotation and genotyping: Initial germline V(D)J gene annotation was performed on the preprocessed BCRs using IgBLAST v.
    IgBLAST
    suggested: (IgBLAST, RRID:SCR_002873)
    1.17.138 with IMGT/GENE-DB release 202113-239.
    IMGT/GENE-DB
    suggested: (IMGT/GENE-DB, RRID:SCR_006964)
    BCR analysis: BCR analysis was performed in R v4.1.0 with visualization performed using base R, ggplot2 v3.3.544, and GraphPad Prism v9.
    ggplot2
    suggested: (ggplot2, RRID:SCR_014601)
    GraphPad Prism
    suggested: (GraphPad Prism, RRID:SCR_002798)
    Clonal overlap between B cell compartments was visualized using circlize v.0.4.1345.
    circlize
    suggested: (circlize, RRID:SCR_002141)
    Phylogenetic trees for S+ clones containing BMPCs were constructed on a by-participant basis using IgPhyML v1.1.346 with the HLP19 model47.
    IgPhyML
    suggested: None
    Gene annotation on human reference chromosomes and scaffolds in Gene Transfer Format (‘gencode.v32.primary_assembly.annotation.gtf’) was downloaded (2021-06-02) from GENCODE v3252, from which a biotype (‘gene_type’) was extracted for each feature.
    GENCODE
    suggested: (GENCODE, RRID:SCR_014966)
    Quality control was performed as follows on the aggregate gene expression matrix consisting of 360,803 cells and 36,601 features using SCANPY v1.7.253 and Python v3.8.8.
    Python
    suggested: (IPython, RRID:SCR_001658)

    Results from OddPub: Thank you for sharing your data.

    Results from LimitationRecognizer: We detected the following sentences addressing limitations in the study:
    A potential limitation to our analyses of S-binding clones is that our selection strategy may have excluded some low-abundance or low-affinity S-specific clones. Nonetheless, we were able to account for 45% and 67% of all GC B cell and LNPC clones, respectively identified by scRNA-seq. This is the first study to provide direct evidence for the induction of antigen-specific BMPCs by an mRNA-based vaccine in humans. Notably, none of the 11 participants from whom post-vaccination bone marrow specimens were examined had a history of SARS-CoV-2 infection. BMPCs that recognized contemporary seasonal influenza virus vaccine antigens and diphtheria/tetanus vaccine antigens were present at frequencies roughly 10- and 2-fold greater than those against SARS-CoV-2 S, respectively. This is likely due to both the greater number of antigenic targets contained in the former vaccines and the repeated exposures to influenza and tetanus/diphtheria vaccine antigens our study participants likely experienced in comparison to the initial exposure to the novel SARS-CoV-2 S antigen. There are some epitopes within the S protein that are conserved between human seasonal coronaviruses and SARS-CoV-228,29. Cross-reactive B cells targeting these epitopes participate in PB and GC B cell responses to SARS-CoV-2 vaccination6,30. It is unlikely, however, that a substantial proportion of the SARS-CoV-2 S+ BMPCs we observed six months after immunization were part of a pre-existing pool of BMPCs, as in a previou...

    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.

  2. Version published to 10.1101/2021.10.31.466651 on bioRxiv