Fig 2: Th2-cTfh Are Activated Early in P. falciparum InfectioncTfh (PD1+CXCR5+ CD4) cells were differentiated into subsets based on CXCR3 and CCR6 expression into Th1 (CXCR3+CCR6-), Th2 (CXCR3-CCR6-), and Th17-like (CXCR3-CCR6+) cell subsets.(A) Subsets as a frequency of cTfh cells after infection.(B) Activated ICOS+ subsets as a frequency of cTfh cells after infection.(C) Activated CD38+ subsets as a frequency of cTfh cells after infection.(D) Activated ICOS+ subsets as a frequency of cTfh cells for each individual study participant following experimental malaria infection. Participants are ranked based on the frequency of ICOS+ cTfh cells before infection. For participants 6,507, 6,505, and 6,506, no sample was available at EOS time point (NA). Flow cytometry-based assays were performed once on whole-blood samples tested in singles.Data are from 40 individuals at days 0, 8, and 14/15 and 37 individuals at EOS. For (A)–(C), Wilcoxon signed-rank tests compared to day 0 are indicated. Boxplots are Tukey style, with median and IQR. Whiskers are to data point no further than 1.5 × IQR; dots are outlying points. Paired data are joined by lines. Data are from 40 individuals at days 0, 8, and 14/15 and 37 individuals at EOS.

Fig 3: cTfh Are Activated in P. falciparum Infection(A) cTfh (PD1+CXCR5+CD4) T cells were assessed by flow cytometry at days 0, 8, and 14/15, and EOS.(B) cTfh cells as a frequency of total CD4 T cells after infection.(C) Representative flow cytometry gating of activated (ICOS+) cTfh cells following infection.(D) Activated cTfh (ICOS+) cells as a frequency of CD4 T cells.(E) Representative histograms of CD38 expression on activated (ICOS+) and non-activated (ICOS-) cTfh cells.(F) CD38 positivity as a frequency of activated ICOS+ cTfh cells. Flow cytometry-based assays were performed once on whole-blood samples tested in singles.Data are from 40 individuals at days 0, 8, and 14/15 and 37 individuals at EOS. (A), (C), and (E) are representative data from a single individual. For (B), (D), and (F), paired Wilcoxon signed-rank tests compared to day 0 are indicated. Boxplots are Tukey style, with median and interquartile range (IQR). Whiskers are to data point no further than 1.5 × IQR; dots are outlying points. Paired data are joined by lines. See also Figures S1 and S2.

Fig 4: Impact of age and malaria on Tfh cell activation and proliferation.Activation and proliferation of Tfh cells was measured by ICOS and Ki67 expression in 212 Ugandan children. A, B The relationship between ICOS+ and Ki67+ on Tfh cells with age. Line is LOESS fit curves with error bands of 95% confidence interval. Spearman's rho and p indicated. C, D The relationship between ICOS and Ki67 and current asymptomatic infection. Mann–Whitney U test indicated. E, F The relationship between ICOS and Ki67 on Tfh cells and household mosquito exposure. Kruskal–Wallis indicated. Box and whisker plots, box indicates first and third quartiles for hingers, median line, and whiskers are lowest and highest values no further than 1.5 interquartile range from hinges. Data beyond whisker lines are indicated with points and are treated as outliers. G Linear regression model coefficients for the relationship between ICOS and Ki67 on each Tfh subset, and age, Pf infection, and household mosquito exposure (HME). All statistical tests are two-sided, with no adjustment for multiple comparisons. Source data are provided as a Source Data file.

Fig 5: Th2-Tfh Cells Are Associated with Antibody Induction, and Th1-Tfh Cells Are Associated with Plasma Cell Expansion(A) To assess total breadth and magnitude of antibody response, responses were categorized as negative (0) or low (1) and high (2) responses and combined to calculate a total antibody score.(B) Association between antibody score and the frequency change of ICOS+ cTfh cells compared to day 0 at each time point.(C) Association between antibody score and the frequency change of each subset of ICOS+ cTfh cells at day 8.(D) Association between plasma cell induction and ICOS+ cTfh cells at day 14/15 following infection.(B–D) Red line is loess. Spearman correlation and ? are indicated. For (B) and (C), p-adj indicates adjusted p values using the Bonferroni-Holm method for the 6 Tfh comparisons with antibody.See also Figures S4 and S5.