Longitudinal Assessment of SARS-CoV-2-Specific T Cell Cytokine-Producing Responses for 1 Year Reveals Persistence of Multicytokine Proliferative Responses, with Greater Immunity Associated with Disease Severity

J Virol. 2022 Jul 13;96(13):e0050922. doi: 10.1128/jvi.00509-22. Epub 2022 Jun 14.


Cell-mediated immunity is critical for long-term protection against most viral infections, including coronaviruses. We studied 23 severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-infected survivors over a 1-year post-symptom onset (PSO) interval by ex vivo cytokine enzyme-linked immunosorbent spot assay (ELISpot) assay. All subjects demonstrated SARS-CoV-2-specific gamma interferon (IFN-γ), interleukin 2 (IL-2), and granzyme B (GzmB) T cell responses at presentation, with greater frequencies in severe disease. Cytokines, mainly produced by CD4+ T cells, targeted all structural proteins (nucleocapsid, membrane, and spike) except envelope, with GzmB and IL-2 greater than IFN-γ. Mathematical modeling predicted that (i) cytokine responses peaked at 6 days for IFN-γ, 36 days for IL-2, and 7 days for GzmB, (ii) severe illness was associated with reduced IFN-γ and GzmB but increased IL-2 production rates, and (iii) males displayed greater production of IFN-γ, whereas females produced more GzmB. Ex vivo responses declined over time, with persistence of IL-2 in 86% and of IFN-γ and GzmB in 70% of subjects at a median of 336 days PSO. The average half-life of SARS-CoV-2-specific cytokine-producing cells was modeled to be 139 days (~4.6 months). Potent T cell proliferative responses persisted throughout observation, were CD4 dominant, and were capable of producing all 3 cytokines. Several immunodominant CD4 and CD8 epitopes identified in this study were shared by seasonal coronaviruses or SARS-CoV-1 in the nucleocapsid and membrane regions. Both SARS-CoV-2-specific CD4+ and CD8+ T cell clones were able to kill target cells, though CD8 tended to be more potent. IMPORTANCE Our findings highlight the relative importance of SARS-CoV-2-specific GzmB-producing T cell responses in SARS-CoV-2 control and shared CD4 and CD8 immunodominant epitopes in seasonal coronaviruses or SARS-CoV-1, and they indicate robust persistence of T cell memory at least 1 year after infection. Our findings should inform future strategies to induce T cell vaccines against SARS-CoV-2 and other coronaviruses.

Keywords: ELISpot assay; SARS-CoV-2; T cell immunity; cytokines; granzyme B; immune modeling.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • CD4-Positive T-Lymphocytes
  • CD8-Positive T-Lymphocytes
  • COVID-19 Vaccines
  • COVID-19* / immunology
  • Cytokines* / immunology
  • Female
  • Humans
  • Immunity*
  • Immunologic Memory
  • Interferon-gamma / metabolism
  • Interleukin-2 / immunology
  • Male
  • SARS-CoV-2*
  • Severity of Illness Index
  • Time Factors


  • COVID-19 Vaccines
  • Cytokines
  • Interleukin-2
  • spike protein, SARS-CoV-2
  • Interferon-gamma