Distinct Features and Functions of Systemic and Mucosal Humoral Immunity Among SARS-CoV-2 Convalescent Individuals

  1. Savannah E. Butler
  2. Andrew R. Crowley
  3. Harini Natarajan
  4. Shiwei Xu
  5. Joshua A. Weiner
  6. Carly A. Bobak
  7. Daniel E. Mattox
  8. Jiwon Lee
  9. Wendy Wieland-Alter
  10. Ruth I. Connor
  11. Peter F. Wright
  12. Margaret E. Ackerman

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Abstract

Understanding humoral immune responses to SARS-CoV-2 infection will play a critical role in the development of vaccines and antibody-based interventions. We report systemic and mucosal antibody responses in convalescent individuals who experienced varying severity of disease. Whereas assessment of neutralization and antibody-mediated effector functions revealed polyfunctional antibody responses in serum, only robust neutralization and phagocytosis were apparent in nasal wash samples. Serum neutralization and effector functions correlated with systemic SARS-CoV-2-specific IgG response magnitude, while mucosal neutralization was associated with nasal SARS-CoV-2-specific IgA. Antibody depletion experiments support the mechanistic relevance of these correlations. Associations between nasal IgA responses, virus neutralization at the mucosa, and less severe disease suggest the importance of assessing mucosal immunity in larger natural infection cohorts. Further characterization of antibody responses at the portal of entry may define their ability to contribute to protection from infection or reduced risk of hospitalization, informing public health assessment strategies and vaccine development efforts.

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  1. ScreenIT

    SciScore for 10.1101/2020.08.05.20168971: (What is this?)

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

    Table 1: Rigor

    Institutional Review Board StatementIRB: All research involving human subjects was approved by the Dartmouth College and Dartmouth-Hitchcock Medical Center Committee for the Protection of Human Subjects (Institutional Review Board) and written informed consent was obtained from all participants.
    Consent: All research involving human subjects was approved by the Dartmouth College and Dartmouth-Hitchcock Medical Center Committee for the Protection of Human Subjects (Institutional Review Board) and written informed consent was obtained from all participants.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variableStudy subjects included both males (17) and females (18).
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Experimental Models: Cell Lines
    SentencesResources
    Antigen and Fc Receptor expression and purification: Prefusion-stabilized, trimer-forming spike protomers (S-2P) of SARS-CoV-2, closely related and/or epidemic strains (SARS-CoV-1, WIV1, and MERS), and endemic coronaviruses (229E, OC43, NL63, and HKU1), and the receptor-binding domain of SARS-CoV-2 fused to a monomeric form of the human IgG4 Fc region were transiently expressed in either Expi 293 or Freestyle 293-F cells, and purified via affinity chromatography according to the manufacturers’ protocols (Supplemental Tables 1 and 2).
    293-F
    suggested: RRID:CVCL_6642)
    Briefly, 1 ^m yellow-green fluorescent microspheres (Thermo, F8813) were covalently conjugated with recombinant RBD and incubated for 3 hrs with dilute serum or nasal wash specimens and either the human monocytic THP-1 cell line (ATCC, TIB-202), or with freshly-isolated primary neutrophils.
    THP-1
    suggested: None
    CD16 reporter assay: The ADCC potential of the specimens was measured using a Jurkat Lucia NFAT cell line (Invivogen, jktl-nfat-cd16), cultured according to the manufacturer’s recommendations, in which engagement of FcyR3a (CD16) on the cell surface leads to the secretion of luciferase.
    Jurkat
    suggested: None
    Software and Algorithms
    SentencesResources
    Data analysis and visualization: Basic analysis and visualization were performed using GraphPad Prism.
    GraphPad Prism
    suggested: (GraphPad Prism, RRID:SCR_002798)
    Heatmaps, correlation plots, and boxplots were made in R (supported by R packages pheatmap, corrplot, and ggplot2).
    ggplot2
    suggested: (ggplot2, RRID:SCR_014601)

    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:
    Chief among the limitations of this work is the small sample size, linked to low local disease prevalence. Other limitations include reliance on self-reported disease severity, and cross-sectional analysis of responses at a somewhat variable period after diagnosis and following varying degree and duration of symptoms among subjects with varying co-morbidities. Additionally, as with most mucosal sampling techniques, there was variability in the volume of nasal wash collected from each subject. This variability appeared to be largely due to accidental ingestion of the fluid during collection and affected volume to a greater degree than concentration. Experimental limitations include the use of lab-adapted cell lines rather than autologous or primary cells for evaluating some Ab functions. Surrogate endpoints of anti-viral activities, such as the substitution of FcγR3a activation and complement C3b deposition were employed as alternatives to assessing infected cell death or viral lysis, and compromises were made in antigenic fidelity by use of recombinant spike proteins. However, assays simplified in these ways may represent approaches with a broad ability to be deployed in global efforts to understand responses to infection and define protective immunity. These compromises notwithstanding, these data have important implications for our understanding of the protection afforded by vaccination or prior infection. When considering vaccine development, an ideal candidate would not o...

    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.

    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 10.3389/fimmu.2020.618685
  3. Version published to 10.1101/2020.08.05.20168971v1 on medRxiv