A. PBMCs were stained with antibodies against CD4 and CD25 or Foxp3 and then analyzed on a flow cytometer. Representative dot plots show the identification of CD25++ T cells (upper panels) or Foxp3+ T cells (lower panels) within CD4+ T cells. Numbers in the plots indicate the frequencies (%) of the gated cells in total PBMCs. B. The frequency of CD4+CD25++ or CD4+Foxp3+ T cells were compared between healthy controls (HC, n = 7, open circle) and scrub typhus patients at acute phase (AP, n = 10, gray circle) or convalescent phase (CP, n = 12, black circle). C. PBMCs were stained with antibodies against CD4 and Fas or CCR4, followed by intracellular staining of Foxp3 and/or CTLA-4 after fixation and permeablization. Mean fluorescent Intensities (MFI) representing CTLA-4, Fas, or CCR4 expression in CD4+Foxp3+ regulatory cells from healthy controls (HC, n = 7–8, open circle) or the patients (AP, n = 7–10, gray circle and CP, n = 9–12, black circle) were compared. Red bars indicate the mean value and p values were obtained using the Mann-Whitney U test or Wilcoxon signed-rank test. Statistically significant p values (<0.05) are shown in bold.