A highly carbapenem-resistant Pseudomonas aeruginosa isolate with a novel blaVIM-4/blaP1b integron overexpresses two efflux pumps and lacks OprD.
OBJECTIVE
A Pseudomonas aeruginosa clinical isolate that exhibited high-level carbapenem resistance and produced metallo-beta-lactamase (MBL) was recovered from a Greek patient. This study was conducted to determine the underlying mechanisms that conferred the carbapenem resistance phenotype.


METHODS
MICs were determined by Etest and Etest MBL. PCR assays were performed for identification of bla(VIM-type), other antibiotic resistance and efflux pump genes and mapping of class 1 integrons. Expression of efflux pump genes was quantified by real-time PCR. Nucleotide sequencing was used to determine the bla(VIM) allele. The location of the MBL allele was investigated by mating experiments, plasmid analysis and hybridization studies.


RESULTS
The isolate was highly carbapenem-resistant (MICs of imipenem and meropenem were 512 and 128 mg/L, respectively) and multidrug-resistant. It harboured the beta-lactamase genes bla(VIM-4) and bla(P1b) in a novel class 1 integron named InV4P1, and a second integron with aac(6')-Ib and bla(OXA-35) gene cassettes. The isolate was deficient in porin OprD and overexpressed efflux pumps MexAB-OprM and MexXY-OprM. Conjugation experiments failed to detect transferable MBL determinants, plasmids were not visualized and bla(VIM) was detected by PCR in the chromosomal band.


CONCLUSIONS
Multiple carbapenem resistance mechanisms are demonstrated to coexist in a single P. aeruginosa isolate and might confer the high-level carbapenem resistance.
Maniati
Maria
M
Department of Microbiology, Medical School, University of Thessaly, Larissa, Greece.
Ikonomidis
Alexandros
A
Mantzana
Paraskevi
P
Daponte
Alexandros
A
Maniatis
Antonios N
AN
Pournaras
Spyros
S
D000368:Aged; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D015780:Carbapenems / Q000494:pharmacology*; D024901:Drug Resistance, Multiple, Bacterial / Q000235:genetics*; D005260:Female; D006115:Greece; D006801:Humans; D032023:Integrons / Q000235:genetics*; D008826:Microbial Sensitivity Tests; D018272:Porins / Q000378:metabolism; D011552:Pseudomonas Infections / Q000382:microbiology; D011550:Pseudomonas aeruginosa / Q000187:drug effects* / Q000201:enzymology / Q000235:genetics / Q000302:isolation & purification; D018440:beta-Lactam Resistance; D001618:beta-Lactamases / Q000235:genetics* / Q000378:metabolism
D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D015780:Carbapenems; D018272:Porins; C077157:OprD protein, Pseudomonas aeruginosa; D001618:beta-Lactamases; C477893:beta-lactamase bla(VIM-4), Pseudomonas aeruginosa