Tfh2 and a subset of Tfh1 cells associate with antibody-mediated immunity to malaria

High-affinity antibody production depends on CD4+ T follicular helper (Tfh) cells. In humans, peripheral blood Tfh cells are heterogenous, as evidenced by differential expression of the chemokine receptors CXCR3 and CCR6, which to date have served to classify 3 subsets, pTfh1, pTfh2, and pTfh17. Although pTfh1 responses dominate during blood-stage Plasmodium infections, a clear association with protective antibody responses remains to be described. We hypothesized that pTfh cells exhibit greater phenotypic and functional heterogeneity than described by CXCR3/CCR6 and that more nuanced pTfh subsets play distinct roles during Plasmodium infection. We mapped pTfh cell heterogeneity in healthy individuals prior to and during controlled human malaria infection (CHMI) using parallel single-cell RNA-Seq and VDJ-Seq. We uncovered 2 pTfh1 subsets or differential phenotypic states, distinguishable by CCR7 expression. Prior to infection, Tfh1-CCR7- cells exhibited higher baseline expression of inflammatory cytokines and genes associated with cytotoxicity. Tfh1-CCR7+ cells had higher germinal center signatures. Indeed, during CHMI, Tfh1-CCR7+, Tfh1-CCR7-, and Tfh2 cells all clonally expanded and became activated. However, only Tfh1-CCR7+ and Tfh2 cells positively associated with protective antibody production. Hence, our data reveal further complexity among human Tfh cells and highlight 2 distinct subsets associated with antibody-mediated immunity to malaria.