Genetic determinants involved in the susceptibility of Pseudomonas aeruginosa to beta-lactam antibiotics

Abstract

The resistome of P. aeruginosa for three β-lactam antibiotics, namely, ceftazidime, imipenem, and meropenem, was deciphered by screening a comprehensive PA14 mutant library for mutants with increased or reduced susceptibility to these antimicrobials. Confirmation of the phenotypes of all selected mutants was performed by Etest. Of the total of 78 confirmed mutants, 41 demonstrated a reduced susceptibility phenotype and 37 a supersusceptibility (i.e., altered intrinsic resistance) phenotype, with 6 mutants demonstrating a mixed phenotype, depending on the antibiotic. Only three mutants demonstrated reduced (PA0908) or increased (glnK and ftsK) susceptibility to all three antibiotics. Overall, the mutant profiles of susceptibility suggested distinct mechanisms of action and resistance for the three antibiotics despite their similar structures. More detailed analysis indicated important roles for novel and known β-lactamase regulatory genes, for genes with likely involvement in barrier function, and for a range of regulators of alginate biosynthesis.

FIG. 1.

Global analysis of the mutants that exhibited a decrease or an increase in susceptibility to at least one β-lactam. Abbreviations for antibiotics are as follows: IPM, imipenem; CAZ, ceftazidime; and MEM, meropenem. (A and B) Venn diagrams resulting from a Boolean analysis of the phenotypes of the mutants. (A) Venn diagram created with mutants showing a decrease in susceptibility to at least one β-lactam. R stands for “resistant” mutants. (B) Venn diagram created with mutants showing an increase in susceptibility to at least one β-lactam. S stands for “sensitive” mutants. (C) Analysis of the association of the phenotypes of the different mutants. The analysis was performed by hierarchical clustering of the MIC fold changes shown in Tables 1 and 2. The green bars stand for mutants with an increase in susceptibility; the red bars stand for mutants with a decrease in susceptibility. The heat map was constructed by using Cluster and TreeView software (14).