Distinct time-dependent S. Typhi-specific CD8⁺ T cell programs are associated with clinical outcome following Ty21a...

CD8⁺ T cells are essential components of the adaptive immune system that play a critical role in determining the clinical severity of typhoid disease by directly killing Salmonella enterica serovar Typhi (S. Typhi)-infected cells, producing cytokines, chemokines, and establishing memory responses that have been linked to clinical outcome in a controlled human infection model (CHIM) of typhoid. Despite decades of extensive research, the role of memory CD8⁺ T (T_M) cells and its subsets as correlates of protection against S. Typhi infection and typhoid disease remains incompletely understood.

In this study, we used specimens from a typhoid CHIM study in which participants who were immunized with live attenuated S. Typhi vaccine, Ty21a, were subsequently challenged with virulent S. Typhi. Some participants developed typhoid disease (TD) and some did not (NoTD), allowing us to assess the immune program of CD8⁺ T_M subsets involved in the prevention of typhoid disease using mass cytometry, CyTOF.

Our high-dimensional profiling data showed that TD and NoTD participants followed divergent memory CD8⁺ T cell trajectories. We observed that CD8⁺ T clusters from NoTD participants are enriched for early multifunctional, protection-associated programs while TD participants exhibited a distinct clustering associated with susceptibility states. Specifically, using unsupervised dimensionality reduction and clustering analysis, T-REX, and feature-ranking analyses, we identified memory CD8⁺ T cell subsets including effector/memory (T_EM), T central/memory (T_CM) and CD45RA+ T_EM (T_EMRA) associated with clinical outcomes. CD8⁺ T_EM from NoTD participants were generally characterized by stronger effector-associated signatures such as CXCR3, IL-17A, TNFα, and MIP1b. In contrast, CD8⁺ T_EM from TD participants showed CCR6- and integrin α4β7-biased immune profiles. Detailed analysis revealed that CD8⁺ T_CM and T_EMRA subsets also contained distinct outcome-associated states. These patterns were confirmed by conventional supervised gating that validated both protection-associated and susceptibility-associated marker combinations across CD8⁺ T_EM, T_CM and T_EMRA subsets. These data suggest that: 1. Protection and susceptibility are not restricted to one CD8⁺ T cell subset but rather are distributed across the broader CD8⁺ T memory landscape and, 2. Typhoid clinical outcome is associated with qualitatively distinct CD8⁺ T memory programs rather than differences only in the magnitude of the overall anti-S. Typhi immune response.