A chromosome-encoded -lactamase gene from Shewanella algae clinical isolate KB-1 was cloned and expressed in Escherichia coli. It encoded the Ambler class D enzyme OXA-55, sharing less than 55% identity with any other oxacillinases. Although conferring a narrow-spectrum -lactam resistance phenotype, OXA-55 had carbapenem-hydrolyzing activity that mirrored the reduced susceptibility to imipenem observed in S. algae KB-1. Very similar oxacillinases were found in other S. algae isolates.Bacteria of the genus Shewanella are gram-negative bacilli, facultatively anaerobic, belonging to the family Alteromonadaceae (25), that are widely distributed in marine and freshwater environments (23). Shewanella algae is a bacterial species recently identified (23, 24) that is closely related to Shewanella putrefaciens (formerly Pseudomonas putrefaciens) (25). Both S. putrefaciens and S. algae species are rare human pathogens (12,15,25).Recently, we have characterized -lactamase OXA-54 from S. oneidensis and we have shown that this Ambler class D enzyme hydrolyzed imipenem significantly (19). Three S. algae isolates were obtained from clinical samples during the last 2 years from our hospital, and the aim of the present study was to determine their -lactamase content.
MATERIALS AND METHODSBacterial strains and plasmids. S. algae clinical isolate KB-1 was isolated from an arm wound of a 10-year-old girl hospitalized at the Bicêtre hospital in 2001 (Kremlin-Bicêtre, France). S. algae KB-2 and KB-3 were from a peritoneum aspiration and from a deep pus sample of the flank, respectively, from patients also hospitalized at Bicêtre hospital. These isolates were identified by the API 32GN system (bioMérieux, Marcy l'Etoile, France) and by sequencing of 16S rRNA genes using primers 16S 8-27 (5Ј-AGAGTTTGATCHTGGYTYAGA-3Ј; H is A, T, or C and Y is C or T) and 16S 1512-1491 (5Ј-ACGGYTACCTTGT TACGACTTC-3Ј; Y is C or T) (9,11,23).S. putrefaciens CIP 8040 (Institut Pasteur strain collection, Paris, France) was used as reference strain. Escherichia coli reference strain DH10B, and plasmid pBK-CMV (Stratagene, Amsterdam, The Netherlands) were used for cloning experiments, whereas rifampin-resistant E. coli JM109 was used for conjugation experiments (3).Antimicrobial agents and MIC determinations. The agents and their sources have been referenced elsewhere (17). MICs were determined by an agar dilution technique on Mueller-Hinton agar (Sanofi Diagnostics Pasteur, Paris, France) with an inoculum of 10 4 CFU per spot and were interpreted according to the guidelines of the National Committee for Clinical Laboratory Standards (14).Plasmid DNA content and conjugation. Plasmid DNA extraction of S. algae KB-1 was performed according to different methods as previously described (18). Direct transfer of -lactam resistance markers into rifampin-resistant E. coli JM109 was attempted by liquid and solid mating-out assays and by electroporation of a putative plasmid DNA suspension of S. algae KB-1 into E. coli DH10B. Transconjugants and electroporants were...