Contribution of PilR to K. kingae PilA1 expression and adherence. (A) Western blot for PilA1 in K. kingae strains KK01, 269-492, KK03, 269-492 pilA1 (pilA1), 269-492 pilR, KK01 pilR, and KK03 pilR. NS/NC, nonspreading/noncorroding; S/C, spreading/corroding; NP, nonpiliated. (B) Adherence to Chang respiratory epithelial cells by K. kingae strains 269-492, 269-492 pilR, KK01 pilR, KK03 pilR, and 269-492 pilA1. (C) Levels of pilA1 transcript measured by quantitative real-time PCR in K. kingae strains KK01, KK03, 269-492 pilR, KK01 pilR, and KK03 pilR relative to those of strain 269-492 (*, P ≤ 0.05 using the unpaired t test compared to results for parental strain). (D) Gel shifts with MBP-PilR. Lanes 1 to 4 contain increasing quantities of MBP-PilR (0, 0.3, 3, and 30 μg) incubated with 50 ng of labeled probe. Lanes 5 and 6 show 3 μg of MBP-PilR incubated with 50 ng of labeled probe and either 400× specific competitor (specific) or 400× nonspecific competitor (nonspecific). Lane 7 shows 30 μg of MPB with 50 ng of labeled probe. The arrow highlights the shifted DNA band at the top of the gel, indicating interaction with PilR.