OXA-24, a Novel Class D β-Lactamase with Carbapenemase Activity in an Acinetobacter baumannii Clinical Strain

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Abstract
Acinetobacter baumannii RYC 52763/97, a clinical isolate involved in a prolonged nosocomial outbreak at our hospital, was resistant to all β-lactams tested, including imipenem and meropenem, which had MICs of 128 and 256 μg/ml, respectively. This strain synthesized three β-lactamases: a plasmid-mediated TEM-1 β-lactamase (pI 5.4), an AmpC-type chromosomal cephalosporinase (pI 9.4), and a novel, presumptively chromosomally mediated OXA-related enzyme (pI 9.0) named OXA-24. After cloning and sequencing, the deduced amino acid sequence of the OXA-24 β-lactamase showed 40% homology with the OXA-10 (PSE-2) and OXA-7 β-lactamases, 39% homology with the OXA-11 and OXA-5 enzymes, and 33% homology with the LCR-1 β-lactamase. The amino acid sequence of the OXA-24 β-lactamase contained the STFK motif found in serine β-lactamases, but the typical class D triad KTG was replaced by KSG and the motif YGN was replaced by FGN. The OXA-24 β-lactamase hydrolyzed benzylpenicillin and cephaloridine but lacked activity against oxacillin, cloxacillin, and methicillin. The enzymatic activity was inhibited by chloride ions and by tazobactam (50% inhibitory concentration [IC50], 0.5 μM), sulbactam (IC50, 40 μM), and clavulanic acid (IC50, 50 μM). Carbapenem MICs for an Escherichia coli transformant (pBMB-1) expressing the cloned OXA-24 enzyme had a fourfold increase. RelativeVmax/Km values of 13 and 6 were obtained with imipenem and meropenem, respectively, and a positive microbiological assay result with imipenem was obtained with a purified enzymatic extract of this transformant strain. Therefore, we consider this new β-lactamase to be involved in the carbapenem resistance of A. baumannii RYC 52763/97.