A clinical isolate of Pseudomonas aeruginosa was found to produce a clavulanic acid-inhibited extendedspectrum -lactamase with a pI of 6.4. PCR, cloning, and sequencing experiments showed that the corresponding bla TEM-21 gene was part of a chromosomally located Tn801 transposon disrupted by an IS6100 element and adjacent to an aac(3)-II gene.Most extended-spectrum -lactamases (ESBLs) produced by enterobacteria derive from the common plasmid-mediated penicillinases TEM-1, TEM-2, and SHV-1 (14). ESBLs have recently emerged in Pseudomonas aeruginosa, where they still remain infrequent. In this bacterial species, ESBLs are mainly OXA derivatives belonging to Ambler class D or newly emerging class A -lactamases such as PER-1 (23) and VEB-1 (9). However, occasionally some enzymes of the TEM or SHV families, previously (TEM-4, TEM-24, and SHV-2a) or newly (TEM-42) described, have been also found (11)(12)(13)19). These observations suggest that ESBLs widespread in the Enterobacteriaceae family may be increasingly found in P. aeruginosa (13), which could also be a reservoir for the dissemination of this kind of enzyme. We report here the production of an unusual TEM derivative, TEM-21, in a clinical strain of P. aeruginosa.(This work was presented in part at the 40th Interscience Conference on Antimicrobial Agents and Chemotherapy, Toronto, Ontario, Canada, 17 to 20 September 2000.) P. aeruginosa Pa141 was isolated in 1997 in a private laboratory in Bordeaux, France, from the urinary tract of a 68-yearold woman living in a nursing home, with Alzheimer's disease and carrying an indwelling urinary tract catheter. This strain belonged to serogroup P2, and by the disk diffusion method it was resistant to all antipseudomonal -lactams except for imipenem. A marked synergistic effect between clavulanic acid and cefotaxime, ceftazidime, or aztreonam was observed, suggesting the presence of an ESBL. Indeed, the isolate remained susceptible to the combinations of ticarcillin and clavulanic acid and of piperacillin and tazobactam. In addition, Pa141 was resistant to gentamicin, tobramycin, netilmicin, ciprofloxacin, and fosfomycin. These data were confirmed by MIC determination (Table 1) by an agar dilution method in Mueller-Hinton medium according to official guidelines (http://www.sfm.asso.fr). Isoelectric focusing of crude -lactamase extracts was performed on a pH 3.5 to 10 Ampholine polyacrylamide gel and revealed by the iodine procedure, with benzylpenicillin (75 g/ml) as substrate. By this method the Pa141 strain produced a single -lactamase of pI 6.4 cofocusing with the TEM-21 reference enzyme (22).Total DNA of P. aeruginosa Pa141 was extracted as previously described (18) and subjected to PCR amplification with primers specific for the bla TEM genes (22), and the PCR product was sequenced on both strands with the D Rhodamine dye terminator kit (Perkin-Elmer, Courtaboeuf, France) and an automatic sequencer (ABI 377; Perkin-Elmer). The nucleotide sequence of the ESBL-encoding gene differed from that of the bla TEM-21 g...