CHARM: COVID-19 Health Action Response for Marines–Association of antigen-specific interferon-gamma and IL2 responses with asymptomatic and symptomatic infections after a positive qPCR SARS-CoV-2 test

  1. Martha Sedegah
  2. Chad Porter
  3. Michael R. Hollingdale
  4. Harini Ganeshan
  5. Jun Huang
  6. Carl W. Goforth
  7. Maria Belmonte
  8. Arnel Belmonte
  9. Dawn L. Weir
  10. Rhonda A. Lizewski
  11. Stephen E. Lizewski
  12. Stuart C. Sealfon
  13. Vihasi Jani
  14. Ying Cheng
  15. Sandra Inoue
  16. Rachael Velasco
  17. Eileen Villasante
  18. Peifang Sun
  19. Andrew G. Letizia

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Abstract

SARS-CoV-2 T cell responses are associated with COVID-19 recovery, and Class I- and Class II-restricted epitopes have been identified in the spike (S), nucleocapsid (N) and membrane (M) proteins and others. This prospective COVID-19 Health Action Response for Marines (CHARM) study enabled assessment of T cell responses against S, N and M proteins in symptomatic and asymptomatic SARS-CoV-2 infected participants. At enrollment all participants were negative by qPCR; follow-up occurred biweekly and bimonthly for the next 6 weeks. Study participants who tested positive by qPCR SARS-CoV-2 test were enrolled in an immune response sub-study. FluoroSpot interferon-gamma (IFN-γ) and IL2 responses following qPCR-confirmed infection at enrollment (day 0), day 7 and 14 and more than 28 days later were measured using pools of 17mer peptides covering S, N, and M proteins, or CD4+CD8 peptide pools containing predicted epitopes from multiple SARS-CoV-2 antigens. Among 124 asymptomatic and 105 symptomatic participants, SARS-CoV-2 infection generated IFN-γ responses to the S, N and M proteins that persisted longer in asymptomatic cases. IFN-γ responses were significantly (p = 0.001) more frequent to the N pool (51.4%) than the M pool (18.9%) among asymptomatic but not symptomatic subjects. Asymptomatic IFN-γ responders to the CD4+CD8 pool responded more frequently to the S pool (55.6%) and N pool (57.1%), than the M pool (7.1%), but not symptomatic participants. The frequencies of IFN-γ responses to the S and N+M pools peaked 7 days after the positive qPCR test among asymptomatic (S pool: 22.2%; N+M pool: 28.7%) and symptomatic (S pool: 15.3%; N+M pool 21.9%) participants and dropped by >28 days. Magnitudes of post-infection IFN-γ and IL2 responses to the N+M pool were significantly correlated with IFN-γ and IL2 responses to the N and M pools. These data further support the central role of Th 1 -biased cell mediated immunity IFN-γ and IL2 responses, particularly to the N protein, in controlling COVID-19 symptoms, and justify T cell-based COVID-19 vaccines that include the N and S proteins.

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  1. Version published to 10.1371/journal.pone.0266691
  2. ScreenIT

    SciScore for 10.1101/2021.12.13.472352: (What is this?)

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

    Table 1: Rigor

    EthicsField Sample Permit: Ethics: The CHARM study was conducted at the Marine Corps Recruitment Depot, Parris Island, South Carolina and has been previously described [21].
    IRB: Institutional Review Board approval was obtained from the Naval Medical Research Center (protocol number NMRC.2020.0006) in compliance with all applicable US federal regulations governing the protection of human participants.
    Consent: All participants provided written informed consent.
    Sex as a biological variablenot detected.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.

    Table 2: Resources

    Software and Algorithms
    SentencesResources
    All statistical analyses were interpreted using a two-tailed alpha=0.05 and were made using SAS v9.4 or JMP v 12 (SAS Institute; Cary, NC).
    SAS
    suggested: (SASqPCR, RRID:SCR_003056)

    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: There are several limitations to this study. Firstly, the study population is young, healthy adults with few comorbidities and may not be reflective of the general population limiting the generalizability of the findings. Secondly, all illnesses were mild, limiting the ability to assess a range of clinical outcomes. Thirdly, the timing of FluoroSpot responses may not have been optimal and further post-infection analyses are warranted. Fourthly, we used cryopreserved cells, and it is possible the recovery of viable cells after thawing may have varied and affected assay readouts. Finally, although we tested immune responses to the S, N and M proteins, it is likely that responses to other proteins may also have a significant role in disease modulation, and responses may partially reflect the relevant sizes of each protein and the numbers of peptides in each pool that were higher for the S pool. We only measured IFN-γ and IL2 producing cells and it is also possible that different cytokines would help to better understand disease severity [30], and phenotypic analysis would also better identify the roles of CD4+ and CD8+ T cell responses.

    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.

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  3. Version published to 10.1101/2021.12.13.472352v1 on bioRxiv