Synergistic effect of CRAMP and antibiotic against Pseudomonas aeruginosa biofilm

Abstract:

[Objective] To investigate the effect of modified mouse cathelicidin related antimicrobial peptide (CRAMP) combined with antibiotics against the mature biofilm of Pseudomonas aeruginosa PAO1 and thereby lay a theoretical basis for the combined application of anti-biofilm drugs. [Methods] Broth microdilution was employed for the determination of minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), and minimum biofilm eradication concentration (MBEC) of CRAMP and antibiotics on PAO1. The time-kill curve (TKC) method was used to determine the bactericidal activities of CRAMP alone, antibiotics alone, and the combination of CRAMP and antibiotics against the mature biofilm of PAO1. Colony counting method and confocal laser scanning microscope (CLSM) were applied to evaluate the dispersal effect of CRAMP combined with antibiotics on the biofilm. [Results] Compared with the antibiotics alone, the combination of antibiotics (except for two carbapenems and four β-lactam antibiotics) with CRAMP recorded the decrease in MBEC, particularly vancomycin, roxithromycin, and azithromycin (1/4 that of antibiotics alone). According to the TKC, the combination of vancomycin, roxithromycin, and azithromycin with CRAMP showed faster and stronger bactericidal activity than antibiotics alone, especially the combination of vancomycin which killed all (100%) biofilm bacteria in 3 h. Under CLSM, the number, volume, area, and fluorescence intensity per unit area of biofilm were significantly changed. [Conclusion] CRAMP can disperse the mature biofilm of PAO1 and has synergistic effect with vancomycin, and the synergistic effect is the most significant at 1 h.

Key words:Pseudomonas aeruginosa;biofilm;antimicrobial peptide;vancomycin;dispersion