Comprehensive longitudinal profiling of SARS-CoV-2-specific CD8+ T-cells reveal strong functional impairment and recognition bias as markers for disease severity

CD8+ T-cells are essential for controlling and resolving SARS-CoV-2 infection, yet their antigen-specific resolution in relation to disease severity, functional dynamics during acute infection, and long-term memory formation remain incompletely understood. Using comprehensive longitudinal profiling of 553 SARS-CoV-2 immunogenic antigens across globally prevalent HLAs, we identified antigen-specific CD8+ T-cell responses that were either critical for early viral clearance or associated with severe disease outcomes. During acute infection, patients with severe COVID-19 exhibited a broader and more robust CD8+ T-cell response than those with mild disease. Notably, we identified HLA-A1-restricted immunodominant antigen-specific T-cells strongly associated with severe disease. These T-cells were present at extremely high frequencies but showed significantly reduced expression of cytotoxic molecules at both the transcriptomic (PRF1, GZMB, GZMH, GNLY) and protein levels (IFN-{gamma}, TNF-, IL-2), as revealed by multidimensional single-cell and cytokine profiling. In contrast, patients with mild disease had T-cells that recognized a more restricted set of antigens, showed only partial overlap with those in severe cases, and showed enhanced cytotoxicity, along with enrichment in gene sets associated with cytotoxic function, hypoxia, and glycolysis. Furthermore, the long-term memory CD8+ T-cells were maintained for a limited subset of immunodominant antigens, with their persistence correlating with their initial frequency during infection. Importantly, SARS-CoV-2 vaccination following infection expanded the long-term T-cell repertoire by enhancing pre-existing responses and generating de novo responses, regardless of prior disease severity. These findings resolve the antigen-specific kinetics and durability of CD8+ T-cells in SARS-CoV-2 infection and provide key insights into their functional landscape. This knowledge could inform future vaccine strategies and therapeutic interventions to enhance protective immunity against emerging viral threats.