Immune Memory in Mild COVID-19 Patients and Unexposed Donors Reveals Persistent T Cell Responses After SARS-CoV-2 Infection

  1. Asgar Ansari
  2. Rakesh Arya
  3. Shilpa Sachan
  4. Someshwar Nath Jha
  5. Anurag Kalia
  6. Anupam Lall
  7. Alessandro Sette
  8. Alba Grifoni
  9. Daniela Weiskopf
  10. Poonam Coshic
  11. Ashok Sharma
  12. Nimesh Gupta

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Abstract

Understanding the causes of the diverse outcome of COVID-19 pandemic in different geographical locations is important for the worldwide vaccine implementation and pandemic control responses. We analyzed 42 unexposed healthy donors and 28 mild COVID-19 subjects up to 5 months from the recovery for SARS-CoV-2 specific immunological memory. Using HLA class II predicted peptide megapools, we identified SARS-CoV-2 cross-reactive CD4 + T cells in around 66% of the unexposed individuals. Moreover, we found detectable immune memory in mild COVID-19 patients several months after recovery in the crucial arms of protective adaptive immunity; CD4 + T cells and B cells, with a minimal contribution from CD8 + T cells. Interestingly, the persistent immune memory in COVID-19 patients is predominantly targeted towards the Spike glycoprotein of the SARS-CoV-2. This study provides the evidence of both high magnitude pre-existing and persistent immune memory in Indian population. By providing the knowledge on cellular immune responses to SARS-CoV-2, our work has implication for the development and implementation of vaccines against COVID-19.

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  1. Version published to 10.3389/fimmu.2021.636768
  2. ScreenIT

    SciScore for 10.1101/2020.11.16.20232967: (What is this?)

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

    Table 1: Rigor

    Institutional Review Board StatementIRB: Ethics Statement: This study was approved by the Institutional review boards of the National Institute of Immunology and All India Institute of Medical Sciences, New Delhi, India.
    Consent: Informed consent was obtained from all subjects during the enrolment.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    The sample absorbance was inversely proportional to the titre of the anti-SARS-CoV-2 neutralizing antibodies.
    anti-SARS-CoV-2 neutralizing antibodies .
    suggested: None
    After 24 hours, cells were washed with 1 mL of PBS with 2% FBS (FACS buffer) and surface stained with antibody cocktail for 1 hour at 4° C in the dark; CD20, CD14, CD16, CD8a and fixable-viability dye coupled with APC eflour 780 in the dump channel, CD4-AlexaFluor 700 (RPA-T4), OX40-FITC (Ber-ACT35)
    CD20
    suggested: (BioLegend Cat# 348805, RRID:AB_2889063)
    CD16
    suggested: None
    After stimulation, cells were washed and surface stained with antibody cocktail for 1 hour at 4°C in the dark; CD20, CD14, CD16, CD4 and fixable-viability dye coupled with APC eflour 780 in the dump channel, CD8-APC (RPA-T8), CD69-BV510 (FN50) and CD137 PE Dazzle (4B4-1).
    CD14
    suggested: (BioLegend Cat# 348805, RRID:AB_2889063)
    CD4
    suggested: None
    CD69-BV510
    suggested: None
    As a control, for total memory B cell analysis, plates were coated with anti-human IgG, IgM, IgA coating antibodies (Mabtech) at concentration of 15µg/mL.
    anti-human IgG
    suggested: None
    For detection of antibody secreting cell (ASC) spots, anti-human IgG-550, IgM-640 and IgA-490 detection antibodies (Mabtech) were added and plate was incubated for 2 hours at room temperature in dark.
    anti-human IgG-550
    suggested: None
    IgA-490 detection antibodies ( Mabtech )
    suggested: None
    Software and Algorithms
    SentencesResources
    Data were analysed using FlowJo 10.5.3.
    FlowJo
    suggested: (FlowJo, RRID:SCR_008520)
    Statistical analyses were performed with the GraphPad Prism software version v8.
    GraphPad Prism
    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:
    The limitations in our study include the sample size and the longitudinal sampling to probe the stability in immunological memory. As this study was of exploratory nature, the 28 long-term recovery samples represent the recruitment in a reasonable timeframe. In fact, the sample size was sufficient to determine the existence of cross-reactive CD4+ T cells and to reveal the persistence of memory CD4+ T cells in several months after recovery from COVID-19. Besides, the predicted epitopes utilized in this study to examine the CD4+ T cells may not cover the responses to all the epitopes in viral genome. However, these predicted peptide pools cover most of the immunodominant epitopes and provide an opportunity to detect the virus-specific CD4+ T cells in limitedly available patient blood sample. Certainly, further studies in long-term after recovery in a larger longitudinal cohort will be helpful in defining the breadth and durability of SARS-CoV-2 reactive memory CD4+ T cells. Moreover, it will be very important to determine if similar characteristics of memory CD4+ T cells exist in the recovery from different outcomes of disease from the asymptomatic to the severe COVID-19. While our work was in review, the knowledge on multiple virus variants emerged in the literature. Because the peptide pools used in our study originates from the reference strain of SARS-CoV-2 (GenBank: MN908947), the current analyses do not provide the information on reactivity of T cells to the mutated epito...

    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.

  3. Version published to 10.1101/2020.11.16.20232967 on medRxiv