Genetic and Phenotypic Variations of a ResistantPseudomonas aeruginosaEpidemic Clone

Abstract

Although constitutively overproduced in these bacteria, the MexXY efflux system did not appear to contribute significantly to aminoglycoside resistance. ␤-Lactam resistance was associated with derepression of intrinsic AmpC ␤-lactamase (with isolate-to-isolate variations of up to 58-fold) and sporadic deficiency in a 46-kDa protein identified as the carbapenem-selective porin OprD. Of the 18 isolates, 14 were also found to overproduce the efflux system MexAB-OprM as a result of alteration of the repressor protein MexR (His1073Pro). However, complementation experiments with the cloned mexR gene demonstrated that MexAB-OprM contributed only marginally to ␤-lactam and fluoroquinolone resistance. Of the four isolates exhibiting wild-type MexAB-OprM expression despite the MexR alteration, two appeared to harbor secondary mutations in the mexA-mexR intergenic region and one harbored secondary mutations in the putative ribosome binding site located upstream of the mexAB oprM operon. In conclusion, this study shows that many mechanisms were involved in the multiresistance phenotype of this highly epidemic strain of P. aeruginosa. Our results also demonstrate that the clone sporadically underwent substantial genetic and phenotypic variations during the course of the outbreak, perhaps in relation to local or individual selective drug pressures.

Molecular Characterization of an Epidemic Clone of Panantibiotic-Resistant Pseudomonas aeruginosa

“…This mechanism contributed to resistance to ␤-lactams, as no acquired extended-spectrum beta-lactamase or oxacillinase was detected in these multiresistant P. aeruginosa isolates. However, differences in resistance to ␤-lactams between isolates were observed (for example, strain P4 was susceptible to ticarcillin/clavulanic acid and strain P5 was susceptible to ceftazidime) and these were probably due to various levels of AmpC cephalosporinase expression (8).…”

“…In contrast, an outbreak reported in Greece as being caused by multiresistant P. aeruginosa serotype O:11 showed distinct PFGE subtypes, which were also recovered from different units of the hospital (24). A recent study from France described the wide dissemination of an epidemic clone of multiresistant P. aeruginosa serotype O:6 inside and outside the hospital (8). Nosocomial P. aeruginosa strains are often multidrug resistant and acquire resistance determinants during the course of antimicrobial therapy in heavily colonized patients.…”

“…Isolates P8 and P17, belonging to PFGE types A1 and K1, respectively, were randomly selected for further analyses. Production of the chromosomally encoded ampC cephalosporinase and oprD porin was quantified as previously described (8). The expression of the MexAB-OprM, MexCD-OprJ, MexEF-OprN, and MexXY efflux pump systems was determined by quantitative real-time reverse transcription-PCR as previously described (8,10).…”

“…The expression of the MexAB-OprM, MexCD-OprJ, MexEF-OprN, and MexXY efflux pump systems was determined by quantitative real-time reverse transcription-PCR as previously described (8,10). DNA sequencing of the putative repressor gene mexZ adjacent to the mexXY operon was done as described by Hocquet et al (8).…”

“…aeruginosa exhibits intrinsic resistance to several ␤-lactams and may acquire additional resistance mechanisms, including decreased outer membrane permeability, penicillin binding protein modifications, production of extended-spectrum ␤-lactamases, acquisition of metallo-␤-lactamases or other enzymes, increased expression of efflux pump systems, and decreased porin expression (5,8,(16)(17)(18)(19). These multipleantibiotic-resistant P. aeruginosa strains caused sporadic nosocomial outbreaks (1,8,24), but several drug-resistant P. aeruginosa clones are now spreading among vulnerable patients, such as those with cystic fibrosis, as recently described (20).…”