title: OXA-18, a class D clavulanic acid-inhibited extended-spectrum beta-lactamase from Pseudomonas aeruginosa.
abstract: Clinical isolate Pseudomonas aeruginosa Mus showed resistance both to extended-spectrum cephalosporins and to aztreonam. We detected a typical double-disk synergy image when ceftazidime or aztreonam was placed next to a clavulanic acid disk on an agar plate. This resistance phenotype suggested the presence of an extended-spectrum beta-lactamase. Isoelectric focusing revealed that this strain produced three beta-lactamases, of pI 5.5, 7.4, and 8.2. A 2.6-kb Sau3A fragment encoding the extended-spectrum beta-lactamase of pI 5.5 was cloned from P. aeruginosa Mus genomic DNA. This enzyme, named OXA-18, had a relative molecular mass of 30.6 kDa. OXA-18 has a broad substrate profile, hydrolyzing amoxicillin, ticarcillin, cephalothin, ceftazidime, cefotaxime, and aztreonam, but not imipenem or cephamycins. Its activity was totally inhibited by clavulanic acid at 2 microg/ml. Hydrolysis constants of OXA-18 (Vmax, Km) confirmed the MIC results. Cloxacillin and oxacillin hydrolysis was noticeable with the partially purified OXA-18. The blaOXA-18 gene encodes a 275-amino-acid protein which has weak identity with all class D beta-lactamases except OXA-9 and OXA-12 (45 and 42% amino acid identity, respectively). OXA-18 is likely to be chromosomally encoded since no plasmid was found in the strain and because attempts to transfer the resistance marker failed. OXA-18 is peculiar since it is a class D beta-lactamase which confers high resistance to extended-spectrum cephalosporins and seems to have unique hydrolytic properties among non-class A enzymes.
authors.affiliation: Service de Bactériologie-Virologie, Hôpital Antoine Béclère, Faculté de Médecine Paris-Sud, Clamart, France.
authors.forename: L N
authors.initials: LN
authors.lastname: Philippon
authors.forename: T
authors.initials: T
authors.lastname: Naas
authors.forename: A T
authors.initials: AT
authors.lastname: Bouthors
authors.forename: V
authors.initials: V
authors.lastname: Barakett
authors.forename: P
authors.initials: P
authors.lastname: Nordmann
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D002876:Chromosomes, Bacterial; D019818:Clavulanic Acid / Q000494:pharmacology*; D003001:Cloning, Molecular; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D007525:Isoelectric Focusing; D007700:Kinetics; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009693:Nucleic Acid Hybridization; D010068:Oxacillin / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D011550:Pseudomonas aeruginosa / Q000201:enzymology* / Q000235:genetics; D012689:Sequence Homology, Nucleic Acid; D065093:beta-Lactamase Inhibitors; D001618:beta-Lactamases / Q000235:genetics / Q000378:metabolism*
chemical_list: D004791:Enzyme Inhibitors; D065093:beta-Lactamase Inhibitors; D019818:Clavulanic Acid; C108862:beta-lactamase OXA-18; D001618:beta-Lactamases; D010068:Oxacillin