Figure 6.

Cytotoxic T cells are silenced for a minimum of 8 mo. Whole blood was preserved within 30 min of blood draw at baseline, diagnosis, and T6 during VAC063C as well as 45 days after challenge (convalescence). Samples were stained with a T cell–focused antibody panel (see Table S3) and acquired on a Helios mass cytometer. After the exclusion of normalization beads and doublets, we gated on CD45^pos CD3^pos T cells for downstream steps. (A) Data from all volunteers and time points was concatenated for UMAP analysis and FlowSOM identified 49 discrete clusters of T cells across the dataset (each given a unique color). The major T cell subsets are labeled according to the expression of lineage, memory, and activation markers. (B) UMAP showing the T cell clusters that are differentially abundant at T6 (versus baseline) in the first and third infection (FDR < 0.05 and >2 fold-change). Clusters that are not significant are shown in black. (C) The mean frequency of each T cell cluster that is differentially abundant in first and/or third infection is shown as a proportion of all CD45^pos CD3^pos T cells at T6. CM, central memory; DN, double negative; EM, effector memory; MAIT, mucosal-associated invariant T cell; NK, natural killer; TEMRA, terminally differentiated effector memory cell re-expressing CD45RA; Treg, regulatory T cell. (D) Pies show the relative size of each differentially abundant cluster. (E) Heatmap showing the normalized median expression values of all markers used for clustering in each of the differentially abundant T cell clusters. The order of features was determined by unsupervised hierarchical clustering. Color codes to the left of the heatmap indicate cluster identity and show whether clusters were significant in the first infection or the first and third infections. Note that no cluster was unique to third infection. In A–E, n = 3 for the first infection and n = 6 for the third infection. The gap between VAC063B and C was 8 mo.