Figure 2

Enhanced Th1-biased immunity in chickens that received the co-administration of S. enterica serovar Typhimurium expressing chIFN-α and chIL-18. (A) AIV H9N2 antigen-specific proliferation of PBMCs. Groups of chickens were administered S. enterica serovar Typhimurium expressing chIFN-α and chIL-18 (10⁹ and 10¹¹ cfu/chicken) and vaccinated with inactivated AIV H9N2 three days later. The vaccination was performed by same protocol twice at 7-day intervals. PBMCs (responders) were prepared from chickens 14 days after booster vaccination, and subsequently stimulated with naïve PBMCs (stimulators) that had been pulsed with inactivated AIV H9N2 antigen. AIV H9N2 antigen-specific proliferation of PBMCs was assessed by measuring viable cell ATP bioluminescence following incubation for 72 h. (B) The expression of IFN-γ and IL-4 mRNA by PBMCs following stimulation with AIV H9N2 antigen. Total RNA was extracted from PBMCs stimulated with AIV H9N2 antigen for 72 h, and subjected to real-time qRT-PCR to determine the expression of IFN-γ and IL-4. Data show the average and SEM of IFN-γ and IL-4 mRNA expression normalized to GAPDH (n = 5). ***p < 0.001 compared to vehicle group treated with control bacteria. p < 0.001 compared to chIFN-α-treated chickens. ^†††p < 0.001 compared to chIL-18-treated chickens.