FIG 1.

Influence of the B. pertussis lipid A GlcN modification on CAMP susceptibility. (A) B. pertussis lipid A GlcN modification increased resistance to CAMPs. B. pertussis BP338 and GlcN mutant strains were incubated with a range of CAMPs and antibiotics, including bacterial polymyxin B, bacterial polymyxin E (colistin), human LL-37, insect CP28, synthetic HHC-10, bovine indolicidin, and gentamicin. The minimum bactericidal concentrations (in micrograms per milliliter as determined by the assay depicted in panel B) for bacterial polymyxin B, bacterial polymyxin E (colistin), human LL-37, insect CP28, synthetic HHC-10, bovine indolicidin, and gentamicin are as follows (value for BP338 and then the value for the GlcN mutant): 8.0 and 2.0, >16 and 4.0, 32.0 and 16.0, 8.0 and 8.0, 8.0 and 8.0, 32.0 and 32.0, and 16.0 and 16.0, respectively. Gentamicin was used as a control. n = 3 for each sample. The results of one representative experiment of three experiments are shown. Data were analyzed with GraphPad Prism 6. Statistical significance was determined by analysis of variance (ANOVA), with a Bonferroni posttest to compare the values for the groups. Values that were significantly different are indicated by asterisks as follows: ****, P < 0.0001; **, P < 0.01. Values that were not significantly different (ns) are indicated. (B) Complementation of the B. pertussis GlcN mutant rescues resistance to polymyxin B. B. pertussis BP338, the GlcN mutant, and the GlcN mutant complemented with lgmABCD (BP338lgmABCDKO plus pPtacLgmABCD) (3) were incubated with a range of polymyxin B (PmB) concentrations, followed by 1/10 dilution, and 2-μl portions of these dilutions were spotted onto BG agar and grown at 37°C for 72 h to observe bacterial survival. The results of one representative experiment of three experiments are shown. Images were converted to gray scale from color images with PowerPoint (Microsoft).