Figure 2.

PfbA contributes to pneumococcal survival after incubation with neutrophils. (A) Growth of TIGR4 strains incubated with human fresh neutrophils. (B) Growth of TIGR4 strains incubated with neutrophil-like differentiated HL-60 cells. Bacterial cells were incubated with human neutrophils or differentiated HL-60 cells in the presence or absence of rPfbA for 1, 2, and 3 h at 37°C in a 5% CO₂ atmosphere. Next, the mixture was serially diluted and plated on TS blood agar. Following incubation, the number of CFUs was determined. Growth index was calculated by dividing CFUs after incubation by CFUs of the original inoculum. (C) Growth of R6 strains incubated with human fresh neutrophils. S. pneumoniae strains were added to human neutrophils without serum and gently mixed for 1, 2, or 3 h at 37°C. Next, the mixtures were serially diluted and plated on TS blood agar. After incubation, the number of CFUs was determined. (D) Growth of R6 strains incubated with human fresh neutrophils in the presence of inhibitors. S. pneumoniae strains were added to human neutrophils with or without cytochalasin D, or protease inhibitor cocktail in the absence of serum, then gently mixed for 1 h at 37°C. The percent bacterial survival was calculated based on viable counts relative to the wild-type strain. These data are presented as the mean values of six samples, with S.E. values represented by vertical lines. Differences between several groups were analyzed using a Kruskal-Wallis test followed by Dunn's multiple comparisons test (A,B). The Mann-Whitney's U-test was used to compare differences between two independent groups (C,D). Three experiments were performed, with data from a representative experiment shown.