Fig. 3.

The pgtE promoter T^D23580 SNP mediates increased resistance to human serum killing, decreases complement component 3 deposition, enhances cleavage of human complement factor B, and promotes virulence in the chicken infection model. (A) Sensitivity to pooled healthy human serum was assayed in a ΔwaaG background (truncated LPS), to observe only the effect of outer membrane proteases. D23580 ΔwaaG showed significantly greater serum resistance than D23580 P_pgtE^4/74 ΔwaaG (P = 0.048). D23580 (B) or 4/74 (C) strains were incubated with pooled human sera and C3 deposition was evaluated by flow cytometry. Data shown is the average of five (B) or three (C) biological replicates, with error bars representing SD. (B) D23580 WT (orange) displayed decreased geometric mean fluorescence intensity (C3 deposition) compared with D23580 P_pgtE^4/74 (light blue) after 5-min incubation with healthy human sera. (C) 4/74 WT (red) showed an increase of C3 deposition compared with 4/74 P_pgtE^D23580 (dark blue) after 10-min incubation with healthy human sera. (D) PgtE-dependent cleavage of complement factor B detected by Western blotting. Polyclonal antibody against factor B was used. (E) Viable counts of S. Typhimurium D23580-derived strains as log CFU/g of spleen at 3-d postoral infection (10⁸ CFU) of 7-d-old Lohmann Brown Layers. Data based on 19 individually sampled birds for each group; combined data for two separately repeated experiments. Each symbol represents the value for an individual chicken and the bars represent the median value for each group.