Aeromonas hydrophila with plasmid-borne class A extended-spectrum β-lactamase TEM-24 and three chromosomal class B, C, and D β-lactamases, isolated from a patient with necrotizing fasciitis.

We report a case of necrotizing fasciitis with probable in vivo transfer of a TEM-24 plasmid-borne extended-spectrum -lactamase (ESBL) gene from Enterobacter aerogenes to Aeromonas hydrophila. The patient was an 87-year-old female with a leg lesion following a trauma. She had a history of rheumatoid polyarthritis treated by 10 mg of prednisone per day, refractory anemia, and chronic venous insufficiency of the lower limbs. Within 5 days, the infection grew worse and the initial amoxicillin-clavulanic acid antibiotic therapy was replaced with ceftriaxone-metronidazole (1 to 1.5 g daily). Surgical debridement revealed extensive necrosis, and 3 days later, the lesion evolved toward typical necrotizing fasciitis (1, 6), leading to a second surgical intervention for above-knee amputation followed by complete healing. Routine bacteriological procedures revealed (i) Escherichia coli NI-202 susceptible to most -lactam compounds, (ii) E. aerogenes NI-203 resistant to all -lactam antibiotics except imipenem, (iii) A. hydrophila NI-204 resistant to ceftazidime, and (iv) A. hydrophila NI-205 susceptible to ceftazidime (Table 1). Pulsed-field electrophoresis confirmed that A. hydrophila NI-204 and NI-205 derived from a single clone. For -lactamase analysis, the E. aerogenes isolate was grown in brain heart infusion broth with and without cefoxitin or ceftazidime induction (10 g/ml) at 37°C before analytical isoelectric focusing with crude sonic cell extracts on polyacrylamide gels (2, 4). Two bands of -lactamase activity were detected with iodine gel with cefazolin (500 g/ml) as the substrate, which was suggestive of the production of an inducible cephalosporinase (pI 8.8) and an ESBL (pI 6.5). Aeromonas isolates were grown at 30°C with cefoxitin (10 g/ml), imipenem (1 g/ml), or tobramycin (1 g/ml) induction (5). Analytical isoelectric focusing with penicillin and cefazolin as substrates revealed three bands (pI 7, 7.8, and 8.2) probably corresponding to previously described cephalosporinase-, imipenemase-, and oxacillinasetype inducible -lactamases (5, 11, 12). A. hydrophila NI-204 produced an additional enzyme similar to E. aerogenes NI-203 ESBL (pI 6.5). Taking into account resistance to ceftazidime, pI determination, and local epidemiology, the ESBL was presumed to be the plasmid-mediated TEM-24 lactamase (2–4, 7). The plasmid was transferred from E. aerogenes NI-203 to A. hydrophila NI-205 and to E. coli C1a at a high frequency (10 ). Recipient strains (NI-206 and NI-207, Table 1) presented the same acquired resistance pattern. After plasmid extraction and gel electrophoresis, both wild-type strains (E. aerogenes NI-203, A. hydrophila NI-204) and recipient strains (A. hydrophila NI-206, E. coli C1a NI-207) showed a common 180-kb band, as previously characterized with Enterobacteriaceae, Pseudomonas aeruginosa, and recently A. caviae (4, 7–9). The capacity of Aeromonas salmonicida to maintain either or both of the Pseudomonas and Enterobacteriaceae R factors has already been observed (10). PCR amplification with TEM family-specific primers was applied to E. aerogenes NI-203 and A. hydrophila NI-204 and showed a deduced protein sequence with 100% identity to that of TEM-24 (3, 7). This report demonstrates probable in vivo transfer of ESBL TEM-24 from E. aerogenes to the genus Aeromonas. It was observed in a wild-type strain of A. hydrophila simultaneously producing the class A, B, C, and D -lactamases.