Persistence of SARS-CoV-2 specific B- and T-cell responses in convalescent COVID-19 patients 6-8 months after the infection

  1. Natalia Sherina
  2. Antonio Piralla
  3. Likun Du
  4. Hui Wan
  5. Makiko Kumagai-Braesh
  6. Juni Andréll
  7. Sten Braesch-Andersen
  8. Irene Cassaniti
  9. Elena Percivalle
  10. Antonella Sarasini
  11. Federica Bergami
  12. Raffaella Di Martino
  13. Marta Colaneri
  14. Marco Vecchia
  15. Margherita Sambo
  16. Valentina Zuccaro
  17. Raffaele Bruno
  18. Tiberio Oggionni
  19. Federica Meloni
  20. Hassan Abolhassani
  21. Federico Bertoglio
  22. Maren Schubert
  23. Miranda Byrne-Steele
  24. Jian Han
  25. Michael Hust
  26. Yintong Xue
  27. Lennart Hammarström
  28. Fausto Baldanti
  29. Harold Marcotte
  30. Qiang Pan-Hammarström

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Abstract

Background

The longevity of the immune response against SARS-CoV-2 is currently debated. We thus profiled the serum anti-SARS-CoV-2 antibody levels and virus specific memory B- and T-cell responses over time in convalescent COVID-19 patients.

Methods

A cohort of COVID-19 patients from the Lombardy region in Italy who experienced mild to critical disease and Swedish volunteers with mild symptoms, were tested for the presence of elevated anti-spike and anti-receptor binding domain antibody levels over a period of eight months. In addition, specific memory B- and T-cell responses were tested in selected patient samples.

Results

Anti-SARS-CoV-2 antibodies were present in 85% samples collected within 4 weeks after onset of symptoms in COVID-19 patients. Levels of specific IgM or IgA antibodies declined after 1 month while levels of specific IgG antibodies remained stable up to 6 months after diagnosis. Anti-SARS-CoV-2 IgG antibodies were still present, though at a significantly lower level, in 80% samples collected at 6-8 months after symptom onset. SARS-CoV-2-specific memory B- and T-cell responses were developed in vast majority of the patients tested, regardless of disease severity, and remained detectable up to 6-8 months after infection.

Conclusions

Although the serum levels of anti-SARS-CoV-2 IgG antibodies started to decline, virus-specific T and/or memory B cell responses increased with time and maintained during the study period (6-8 months after infection).

Funding

European Union’s Horizon 2020 research and innovation programme (ATAC), the Italian Ministry of Health, CIMED, the Swedish Research Council and the China Scholarship Council.

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  1. ScreenIT

    SciScore for 10.1101/2020.11.06.371617: (What is this?)

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

    Table 1: Rigor

    Institutional Review Board StatementConsent: Study inclusion criteria included subjects over 18 years of age, who were willing and able to provide informed consent, confirmed positivity of SARS-CoV-2 by real-time RT-PCR targeting the E and RdRp genes according to Corman et al. protocols 12 and monitored until two subsequent samples with negative results.
    IRB: The study was performed under the approval of the Institutional Review Board of Policlinico San Matteo (protocol number P_20200029440).
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variableThe patients had a median age of 63 years (range 32-89) with 45 (58%) males and 33 (42%) females.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    Enumeration of B cells secreting IgG antibodies specific for SARS-CoV-2 RBD and T cells secreting IFN-γ and IL-2 in response to SARS-CoV-2 peptides: PBMCs were incubated for four days in RPMI-1640 medium with 10% FCS, supplemented with the TLR7 and TLR8 agonist imidazoquinoline resiquimod (R848, 1 µg/ml; Mabtech AB, Nacka, Sweden), and recombinant human IL-2 (10 ng/ml) for stimulation of memory B cells 18.
    TLR7
    suggested: None
    The ELISpot plates pre-coated with capturing monoclonal anti-human IgG antibodies were incubated with a total of 300 000 or 30 000 pre-stimulated cells per well for detection of RBD-specific IgG and total IgG secreting cells, respectively.
    anti-human IgG
    suggested: None
    total IgG
    suggested: None
    The number of B cells secreting IgG antibodies specific for SARS-CoV-2 RBD and cells secreting IgG (total IgG) were measured using the Human IgG SARS-CoV-2 RBD ELISpotPLUS (ALP) kit according to the manufacturer’s protocol (Mabtech AB).
    ALP
    suggested: None
    Statistical analysis: Mann-Whitney U test was used for comparisons between groups in anti-SARS-CoV-2 antibody levels and numbers of specific memory B- and T-cells.
    anti-SARS-CoV-2
    suggested: None
    Experimental Models: Cell Lines
    SentencesResources
    Detection of antibodies specific to SARS-CoV-2: RBD-His protein was expressed in Expi293 cells and purified on Ni-NTA resin (#88221, Thermofisher) followed by size-exclusion chromatography on a Superdex 200 gel filtration column in PBS 13.
    Expi293
    suggested: RRID:CVCL_D615)
    Experimental Models: Organisms/Strains
    SentencesResources
    S1-S2-His (referred as S) protein was expressed baculovirus-free in High Five insect cells 14 and purified on HisTrap excel column (Cytiva) followed by preparative size exclusion chromatography on 16/600 Superdex 200 kDa pg column (Cytiva) 15.
    S1-S2-His
    suggested: None
    Software and Algorithms
    SentencesResources
    High-binding Corning Half area plates (Corning #3690) were coated over night at 4°C with S or RBD protein (1.7 μg/ml for IgG and IgM; 2.0 μg/ml for IgA) in PBS; washed three times in PBS-Tween (0.05%) and blocked with 2% BSA in PBS for 1h at room temperature.
    IgM
    suggested: None
    Results of ELISpot and Fluorospot assays were evaluated using an IRIS-reader and analyzed by IRIS software version 1.1.9 (Mabtech AB).
    IRIS
    suggested: None
    All analyses and data plotting were performed using GraphPad or R version 3.6.1.
    GraphPad
    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: 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.

    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 10.1101/2020.11.06.371617v1 on bioRxiv