Novel plasmid-mediated beta-lactamase in clinical isolates of Klebsiella pneumoniae more resistant to ceftazidime than to other broad-spectrum cephalosporins

Abstract

Multiresistant Klebsiella pneumoniae strains isolated from three patients in the same intensive care unit were more resistant to ceftazidime than to cefotaxime and aztreonam but remained susceptible to moxalactam and imipenem. Resistance to I-lactams, kanamycin, streptomycin, sulfonamides, and tetracyclines was transferable to Escherichia coli by conjugation and was lost en bloc after treatment with ethidium bromide. Agarose gel electrophoresis of wild types and transconjugants indicated that these resistances were mediated by a 150-kilobase plasmid, pCFF14. The strains constitutively produced a ,-lactamase with isoelectric point close to 5.6 and which had a higher Vmax for ceftazidime and cephalothin than for cefotaxime. The substrate profile and isoelectric point of this enzyme thus differ from those of other known plasmid-mediated (-lactamases, including the broad-spectrum enzyme CTX-1. Hybridization studies support the derivation of the novel enzyme from a TEM-type ,I-lactamase.Klebsiella spp. are generally susceptible to cephalothin and most cephalosporins but are moderately resistant to penicillins by virtue of synthesis of a chromosomal penicillinase (14,20,21,27). Some Klebsiella isolates are resistant to high levels of penicillins by production of plasmid-mediated TEM-like penicillinases; these strains also have reduced susceptibility to the older cephalosporins (7,19,24). Recently, the emergence of resistance to broad-sprectrum cephalosporins has been reported in strains of Klebsiella oxytoca (6, 15) and Klebsiella pneumoniae (22). Transferable resistance to cefotaxime was demonstrated in the latter species (10). We previously described an outbreak of infections caused by K. pneumoniae resistant to all P-lactams except moxalactam and imipenem by production of the novel plasmid-mediated P-lactamase, . In this paper, we describe another novel plasmid-mediated P-lactamase, which is markedly active against ceftazidime and which was isolated in K. pneumoniae (25). MATERIALS AND METHODSBacterial strains. Three multiply resistant strains of K. pneumoniae, CF504, CF514, and CF524, were isolated from patients hospitalized in the same intensive care unit at St.-Jacques Hospital, Clermont-Ferrand, France. The biotypes of the strains determined by the API 50CH system were the same for CF504 and CF514. K. pneumoniae CF104 (CTX-1 producing) was included in this study for comparison. A rifampin-resistant mutant of Escherichia coli K-12, C600, was used in transfer experiments (1).Antibiotics Transfer of resistance characters and curing. Conjugation experiments were done at 37°C for 40 min, as previously described (16). Plasmids were transferred to mutants of E. coli K-12 strain C600 resistant to rifampin. Transconjugants CF604, CF614, and CF624 were selected on Mueller-Hinton agar containing rifampin (300 ,ug/ml) plus ceftazidime (4 ,ug/ml), kanamycin (20 ,ug/ml), tetracycline (2 pug/ml), or sulfonamide (200 p.g/ml). The frequency of transfer was expressed relative to the number of donor cells. Curing of antibiotic re...

Coexistence of SHV-4- and TEM-24-Producing Enterobacter aerogenes Strains before a Large Outbreak of TEM-24-Producing Strains in a French Hospital

“…The first nosocomial outbreaks caused by ESBL-producing strains occurred in 1985 in France (26,34). Klebsiella pneumoniae was the ESBL-producing enterobacterium most frequently isolated from clinical specimens, but E. aerogenes has recently emerged as an important hospital opportunist.…”

Development of "oligotyping" for characterization and molecular epidemiology of TEM beta-lactamases in members of the family Enterobacteriaceae

“…Molecular analysis (30) and nucleotide sequence determination (29) of blaTEM-3, the structural gene for the enzyme, indicated that the P-lactamase was a double point mutation of TEM-2 penicillinase and was therefore redesignated TEM-3 (30). Since then, similar enzymes of either the SHV type (5,9) or the TEM type (3,8,22,24) have been detected among members of the family Enterobacteriaceae. The epidemic aspect of this resistance, the fact that it involves recent cephalosporins and monobactams, and the mechanism of substrate range expansion by point mutations in well-known penicillinases prompted us to study the distribution of TEM P-lactamases in strains of enterobacteria and to develop techniques for the rapid detection and characterization of new variants.…”

“…Molecular analysis (30) and nucleotide sequence determination (29) of blaTEM-3, the structural gene for the enzyme, indicated that the P-lactamase was a double point mutation of TEM-2 penicillinase and was therefore redesignated TEM-3 (30). Since then, similar enzymes of either the SHV type (5, 9) or the TEM type (3,8,22,24) …”

Nosocomial Spread of Ceftazidime-Resistant Klebsiella pneumoniae Strains Producing a Novel Class A β-Lactamase, GES-3, in a Neonatal Intensive Care Unit in Japan

“…and members of the family Enterobacteriaceae have been documented (6). Since the late 1980s, extended-spectrum ␤-lactamases (ESBLs) derived from TEM-and SHV-type penicillinases capable of hydrolyzing the oxymino-cephalosporins have been spreading globally, mainly in the Enterobacteriaceae, including Klebsiella pneumoniae and Escherichia coli (5,23,29). Moreover, various non-TEM-, non-SHV-type class A ␤-lactamases exhibiting extended-spectrum activities, including CTX-M-type (13,31,38,39,41), SFO-type (18), VEB-type (12,20,25), and GES-type (10,11,19,24,28,37) ␤-lactamases, have also been reported in various gram-negative bacilli.…”