OXA-28, an extended-spectrum variant of OXA-10 beta-lactamase from Pseudomonas aeruginosa and its plasmid- and integron-located gene

Pseudomonas aeruginosa ED-1, isolated from a pulmonary brush of a patient h ospitalized in a suburb of Paris, France, was resistant to ceftazidime and of intermediate susceptibility to ureidopenicillins and to cefotaxime. Clon ing and expression of the beta -lactamase gene content of this isolate in E scherichia coil DH10B identified a novel OXA-10 variant, OXA-28, with a pI value of 8.1 and a molecular mass of 29 kDa. It differed from OXA-10 by 10 amino acid changes and from OXA-13 and OXA-19 by 2 amino acid changes, incl uding a glycine instead of tryptophan at position 164, which is likely invo lved in its resistance to ceftazidime. Like OXA-11, -14, -16, and -19 and a s opposed to OXA-17, OXA-28 predominantly compromised ceftazidime and had o nly marginal effect on the MICs of aztreonam and cefotaxime in P. aeruginos a. Once expressed in E. coli, OXA-28 raised the MIC of ceftazidime to a muc h higher level than those of amoxicillin, cephalothin, and cefotaxime (128, 16, 8, and 4 mug/ml, respectively). OSA-28 beta -lactamase had a broad spe ctrum of activity, including ceftazidime. Its activity was partially antago nized by clavulanic acid (50% inhibitory concentration, 10 muM) and NaCl ad dition. The oxa28 gene cassette nas inserted in the variable region of a cl ass 1 integron, In57, immediately downstream of an amino 6'-N-acetyltransfe rase gene cassette, aac(6')Ib. The structures of the integrons carrying eit her oxa28, oxa13, or oxa19 gene cassettes were almost identical, suggesting that they may have derived from a common ancestor as a result of the commo n European origin of the P. aeruginosa isolates. In57 was located on a self -transferable plasmid of ca. 150 kb that was transferred from P. aeruginosa to P. aeruginosa.