Figure 3

Frequencies of splenic IFN-γ-producing CD4⁻CD8β⁻ cells following stimulation with PMA/ionomycin or H. meleagridis. A, B Representative pseudocolor plots show IFN-γ versus CD4 expression in CD4⁻CD8β⁻ pre-gated (not depicted) splenocytes isolated from birds 2 weeks pi and 5 weeks pi. Approximately A 200 000 and B 180 000 CD4⁻CD8β⁻ cells are shown in each plot and numbers indicate frequencies of IFN-γ⁺CD4⁻CD8β⁻ cells within total CD4⁻CD8β⁻ cells. Graphs on the right display frequencies of IFN-γ-producing CD4⁻CD8β⁻ cells from all birds. Each symbol represents one bird, black and red colored symbols represent birds sacrificed 2 weeks pi and 5 weeks pi, respectively. A Scatter plots show percent of IFN-γ⁺ cells within the CD4⁻CD8β⁻ subset after PMA/ionomycin stimulation compared to medium in control and infected birds (left panel). Right panel: comparison of IFN-γ-producing CD4⁻CD8β⁻ cell frequencies after stimulation with PMA/ionomycin between infected and control birds. B Scatter plots as in A but after H. meleagridis/E. coli stimulation and E. coli-only stimulation. Right panel shows in addition percent of IFN-γ⁺ cells after E. coli correction for infected and control birds. Asterisks indicate different p-values: *p ≤ 0.05 and **p ≤ 0.01.