RESUMO
Although mRNA vaccine efficacy against severe COVID-19 remains high, variant emergence and breakthrough infections have changed vaccine policy to include booster immunizations. However, the effect of diverse and repeated antigen exposures on SARS-CoV-2 memory T cells is poorly understood. Here, we utilize DNA-barcoded MHC-multimers combined with scRNAseq and scTCRseq to capture the ex vivo profile of SARS-CoV-2-responsive T cells within a cohort of individuals with one, two, or three antigen exposures, including vaccination, primary infection, and breakthrough infection. We found that the order of exposure determined the relative distribution between spike- and non-spike-specific responses, with vaccination after infection leading to further expansion of spike-specific T cells and differentiation to a CCR7-CD45RA+ effector phenotype. In contrast, individuals experiencing a breakthrough infection mount vigorous non-spike-specific responses. In-depth analysis of over 4,000 epitope-specific T cell receptor sequences demonstrates that all types of exposures elicit diverse repertoires characterized by shared, dominant TCR motifs, with no evidence for repertoire narrowing from repeated exposure. Our findings suggest that breakthrough infections diversify the T cell memory repertoire and that current vaccination protocols continue to expand and differentiate spike-specific memory responses.
RESUMO
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged in late 2019 and has since caused a global pandemic resulting in millions of cases and deaths1-4. Diagnostic tools and serological assays are critical for controlling the outbreak5-7, especially assays designed to quantitate neutralizing antibody levels, considered the best correlate of protection8-11. As vaccines become increasingly available12, it is important to identify reliable methods for measuring neutralizing antibody responses that correlate with authentic virus neutralization but can be performed outside of biosafety level 3 (BSL3) laboratories. While many neutralizing assays using pseudotyped virus have been developed, there have been few studies comparing the different assays to each other as surrogates for authentic virus neutralization9,10,13,14. Here we characterized three enzyme-linked immunosorbent assays (ELISAs) and three pseudotyped VSV virus neutralization assays and assessed their concordance with authentic virus neutralization. The most accurate assays for predicting authentic virus neutralization were luciferase and secreted embryonic alkaline phosphatase (SEAP) expressing pseudotyped virus neutralizations, followed by GFP expressing pseudotyped virus neutralization, and then the ELISAs.
