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

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Antimicrob. Agents Chemother. doi:10.1128/AAC.00427-09
Copyright (c) 2009, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Molecular and biochemical characterization of the natural chromosome-encoded class A -lactamase from Pseudomonas luteola

Benoît Doublet, Frédéric Robin, Isabelle Casin, Laëtitia Fabre, Anne Le Fleche, Richard Bonnet, and François-Xavier Weill^*

Institut Pasteur, Laboratoire des Bactéries Pathogènes Entériques, Paris, France; INRA, UR1282 Infectiologie Animale Santé Publique, Nouzilly, France; CHU de Clermont-Ferrand, Laboratoire de bactériologie, Clermont-Ferrand, France; Université Clermont 1, UFR de Médecine, Laboratoire de Bactériologie, JE2526 usc INRA2018, Clermont-Ferrand, France; Hôpital St-Louis, Service de Microbiologie, Paris, France; Institut Pasteur, Unité de Biodiversité des Bactéries Pathogènes Emergentes, Paris, France

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

Pseudomonas luteola (formerly classified as CDC group Ve-1 and named Chryseomonas luteola) is an unusual pathogen implicated in rare but serious infections in humans. A novel -lactamase gene, bla_LUT-1 was cloned from the whole-cell DNA of the P. luteola clinical isolate LAM, which had a weak narrow-spectrum -lactam-resistant phenotype, and expressed in Escherichia coli. This gene encoded LUT-1, a 296-amino acid Ambler class A -lactamase with a pI value of 6 and a theoretical molecular mass of 28.9 kD. The catalytic efficiency of this enzyme was higher for cephalothin, cefuroxime, and cefotaxime than for penicillins. It was found to be 49 % to 59 % identical to other Ambler class A -lactamases from Burkholderia sp. (PenA to PenL), Ralstonia eutropha (REUT), Citrobacter sedlakii (SED-1), Serratia fonticola (FONA and SFC-1), Klebsiella sp. (KPC and OXY), and CTX-M extended-spectrum -lactamases. No gene homologous to the regulatory ampR genes of class A -lactamases was found in the vicinity of the bla_LUT-1 gene. The entire bla_LUT-1 coding region was amplified by PCR and sequenced in five other genetically unrelated P. luteola strains (including the P. luteola type strain). A new variant of bla_LUT-1 was found for each strain. These genes (named bla_LUT-2 to bla_LUT-6) had nucleotide sequences 98.1 to 99.5% identical to that of bla_LUT-1, and differing from this gene by two to four nonsynonymous single nucleotide polymorphisms. The bla_LUT- gene was located on a 700 to 800 kb chromosomal I-CeuI fragment, the precise size of this fragment depending on the P. luteola strain.