Fig. 6

Co-infection induces early apoptosis of CD4 + T cells in the popliteal lymph node (pLN). Joint inflammation (a), viraemia (b) and viral load in the whole body (c) and footpad (d) of WT + CHIKV (n = 5), WT + CHIKV + PbA (n = 5), LTa^-/- + CHIKV (n = 4) and LTa^-/- + CHIKV + PbA (n = 4) groups. The orange box denotes suppression of joint inflammation (5–8 dpi) in co-infected WT mice compared to WT + CHIKV, which is lost in the co-infected LTa^-/- mice. Data from co-infected mice was compared with their respective CHIKV infected controls on the same genetic background using Mann-Whitney two-tailed analysis. Comparisons between WT + CHIKV and WT + CHIKV + PbA are represented by *P < 0.05 and **P < 0.01; comparisons between LTa^-/- + CHIKV and LTa^-/- + CHIKV + PbA are represented by ⁺P < 0.05. Quantification of CHIKV-specific CD4 + T cells (ELISPOT) (e), total CD4 + T cells (flow cytometry) (f) and late apoptotic CD4 + T cells (g) in the pLN of mock, CHIKV and CHIKV + PbA (n ≥ 5 per group) at 2, 4, 5 and 6 dpi. e Representative ELISPOT images showing spots generated by virus-stimulated CD4 + T cells isolated from 100,000 pLN cells (4 and 5 dpi) and 200,000 pLN cells (6 dpi) in CHIKV-infected and co-infected mice. g Representative dot plots displaying late apoptotic CD4 + T cells gating on 6 dpi. Late apoptotic CD4 + T cells are defined as CD45 + CD3 + CD4 + FVD + AnnexinV + cells. Data shown are pooled from four independent experiments. Data (e–g) from the co-infected group were compared against CHIKV-infected controls at the respective time-point using Mann-Whitney two-tailed test (*P < 0.05 and **P < 0.01). Data represent the means ± SD. Abbreviations: CHIKV, Chikungunya virus; dpi, days post infection; PbA, Plasmodium berghei-ANKA