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Antimicrobial Agents and Chemotherapy, May 2009, p. 1987-1997, Vol. 53, No. 5
0066-4804/09/$08.00+0 doi:10.1128/AAC.01024-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.
Efflux Unbalance in Pseudomonas aeruginosa Isolates from Cystic Fibrosis Patients
Lucie Vettoretti,1
Patrick Plésiat,1*
Cédric Muller,1
Farid El Garch,2
Gilles Phan,3
Inna Attrée,4
Arnaud Ducruix,3 and
Catherine Llanes1
Department of Bacteriology, University of Franche-Comté, Faculty of Medicine, F-25030 Besançon, France,1 Unité de Pharmacologie Cellulaire et Moléculaire, Université Catholique de Louvain, Brussels, Belgium,2 Laboratoire de Cristallographie, RMN Biologique, UMR CNRS 8015, Faculté de Pharmacie, Paris V, F-75270 France,3 Commissariat à l'Energie Atomique, Laboratoire de Biochimie et de Biophysique des Systèmes Intégrés, F-38054 Grenoble, France4
Received 31 July 2008/ Returned for modification 10 October 2008/ Accepted 18 February 2009
Retrospective analysis of 189 nonredundant strains of Pseudomonas aeruginosa sequentially recovered from the sputum samples of 46 cystic fibrosis (CF) patients over a 10-year period (1998 to 2007) revealed that 53 out of 189 (28%) samples were hypersusceptible to the β-lactam antibiotic ticarcillin (MIC 
4 µg/ml) (phenotype dubbed Tichs). As evidenced by trans-complementation and gene inactivation experiments, the mutational upregulation of the efflux system MexXY was responsible for various degrees of resistance to aminoglycosides in a selection of 11 genotypically distinct strains (gentamicin MICs from 2 to 64 µg/ml). By demonstrating for the first time that the MexXY pump may evolve in CF strains, we found that a mutation leading to an F1018L change in the resistance-nodulation-cell division (RND) transporter MexY was able to increase pump-promoted resistance to aminoglycosides, cefepime, and fluoroquinolones twofold. The inactivation of the mexB gene (which codes for the RND transporter MexB) in the 11 selected strains showed that the Tichs phenotype was due to a mutational or functional loss of function of MexAB-OprM, the multidrug efflux system known to contribute to the natural resistance of P. aeruginosa to β-lactams (e.g., ticarcillin and aztreonam), fluoroquinolones, tetracycline, and novobiocin. Two of the selected strains synthesized abnormally low amounts of the MexB protein, and 3 of 11 strains expressed truncated MexB (n = 2) or MexA (n = 1) polypeptide as a result of mutations in the corresponding genes, while 7 of 11 strains produced wild-type though nonfunctional MexAB-OprM pumps at levels similar to or even higher than that of reference strain PAO1. Overall, our data indicate that while MexXY is necessary for P. aeruginosa to adapt to the hostile environment of the CF lung, the MexAB-OprM pump is dispensable and tends to be lost or inactivated in subpopulations of P. aeruginosa.
* Corresponding author. Mailing address: Laboratoire de Bactériologie, EA 3186, UFR Sciences Médicales et Pharmaceutiques, 19 rue Ambroise Paré, 25041 Besançon Cedex, France. Phone: (33) 3 63 08 22 06. Fax: (33) 3 63 08 22 32. E-mail: patrick.plesiat{at}univ-fcomte.fr
Published ahead of print on 2 March 2009.
Antimicrobial Agents and Chemotherapy, May 2009, p. 1987-1997, Vol. 53, No. 5
0066-4804/09/$08.00+0 doi:10.1128/AAC.01024-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.
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