AAC Accepts, published online ahead of print on 2 November 2009

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Antimicrob. Agents Chemother. doi:10.1128/AAC.01037-09
Copyright (c) 2009, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Screening and Quantification of the Expression of Antibiotic Resistance Genes in Acinetobacter baumannii with a Microarray

Sébastien Coyne, Ghislaine Guigon, Patrice Courvalin^*, and Bruno Périchon

Institut Pasteur, Unité des Agents Antibactériens, and Plateforme Génotypage des Pathogènes et Santé Publique, 75724 Paris Cedex 15, France

^* To whom correspondence should be addressed. Email: patrice.courvalin{at}pasteur.fr.

An oligonucleotide-based DNA microarray was developed to evaluate expression of genes for efflux pumps in Acinetobacter baumannii and to detect acquired antibiotic resistance determinants. The microarray contained probes for 205 genes including those for 47 efflux systems, 55 resistance determinants, and 35 housekeeping genes. The microarray was validated by comparative analysis of mutants overexpressing or deleted for the pumps relative to the parental strain. The performance of the microarray was also evaluated using in vitro single step mutants obtained on various antibiotics. Overexpression, confirmed by quantitative RT-PCR, of RND efflux pumps AdeABC, due to a G30D substitution in AdeS in a multidrug resistant (MDR) strain obtained on gentamicin and of AdeIJK in two mutants obtained on cefotaxime or tetracycline was detected. A new efflux pump, AdeFGH, was found to be overexpressed in a mutant obtained on chloramphenicol. Study of MDR clinical isolates, including the AYE strain whose entire sequence has been determined, indicated overexpression of AdeABC and of the chromosomally encoded cephalosporinase as well as the presence of several acquired resistance genes. The overexpressed and acquired determinants detected by the microarray could account for nearly the entire MDR phenotype of the isolates. The microarray is potentially useful for detection of resistance in A. baumannii and should allow detection of new efflux systems associated with antibiotic resistance.