First characterization of inhibitor-resistant TEM (IRT) beta-lactamases in Klebsiella pneumoniae strains

Abstract

Two clinical strains of Klebsiella pneumoniae, TP 01 and TP 02, presented resistance to amoxicillin-clavulanate and were fully susceptible to cephalothin. These strains produced two ␤-lactamases, SHV-1 and a TEM enzyme with a pI of 5.2. The previously described changes Arg-2443Cys and Arg-2443Ser in IRT-1 and IRT-2 (A. Belaaouaj, C. Lapoumeroulie, M. M. Caniça, G. Vedel, P. Nevot, R. Krishnamoorthy, and G. Paul, FEMS Microbiol. Lett. 120:75-80, 1994) were found in TEM enzymes from the TP 01 and TP 02 strains, respectively. This is the first report of inhibitor-resistant TEM (IRT) in species other than Escherichia coli from the family Enterobacteriaceae.Most Klebsiella pneumoniae strains are moderately resistant to amino-and carboxy-penicillins by synthesis of a class A chromosomally mediated penicillinase (SHV-1), and clavulanic acid has a synergistic effect when associated with these penicillins. These strains are generally susceptible to cephalothin and all other cephalosporins. Some K. pneumoniae isolates which are highly resistant to penicillins because of the production of plasmid-mediated TEM-like penicillinases also have reduced susceptibility to cephalosporins.Previous reports have established that the susceptibility of Escherichia coli to ␤-lactamase inhibitors can be affected by the overproduction of TEM-1 ␤-lactamase (14, 18). In K. pneumoniae, overproduction of either TEM-1 or SHV-1 increased resistance to amoxicillin-clavulanic acid and to cephalothin (16).In this report, we describe the characterization of the ␤-lactamases produced by two clinical isolates of K. pneumoniae that exhibited high-level resistance to amoxicillin-clavulanate and susceptibility to cephalothin.K. pneumoniae TP 01 and TP 02 were isolated from the urine and respiratory tracts of patients hospitalized in the geriatric and pneumology services, respectively, of a teaching hospital in Toulouse. The MICs of amoxicillin, amoxicillin-clavulanate, ticarcillin, ticarcillin-clavulanate, piperacillin, piperacillin-tazobactam (clavulanic acid and tazobactam were at fixed concentrations of 2 g/ml and 4 g/ml, respectively), and cephalothin were determined by dilution in Mueller-Hinton agar (Sanofi Diagnostics Pasteur, Marnes-la-Coquette, France) with an inoculum of 10 4 CFU per spot. A K. pneumoniae strain (CF 004) producing only the chromosomal SHV-1 enzyme, a K. pneumoniae strain (CF 014) producing both SHV-1 and TEM-1, and a K. pneumoniae strain (CF 024) producing SHV-1 enzyme at a high level were studied as comparisons for the MIC determinations (Table 1).K. pneumoniae CF 004 (SHV-1) was characterized by susceptibility to all ␤-lactams tested except for amoxicillin and ticarcillin. The MIC for both of these drugs was 128 g/ml. The resistance patterns of K. pneumoniae TP 01 and TP 02 were characterized by high MICs of amoxicillin-clavulanate (with clavulanate at a fixed concentration of 2 g/ml) of 256 and 512 g/ml, respectively, while that for TEM-1-producing strain CF 014 is only 32 g/ml. MICs of ticarcillin-clavulanate for inhibitor-...

Analysis of the plasticity of location of the Arg244 positive charge within the active site of the TEM‐1 β‐lactamase

“…Similar results were obtained in previous studies examining TEM-1 Arg244 substitutions and the observation is consistent with the proposed role of the arginine side chain stabilizing substrate through interactions with the carboxylate group of b-lactam antibiotics. 10,26,27 Site-directed mutagenesis was used to combine each of the Leu220Arg, Met272Arg, and Asn276Arg mutations with the Arg244Ala mutation with the intent of returning a guanidinium group to the area left vacant from the removal of arginine 244. The effect of the arginine substitutions at 220, 272, and 276 in the context of the Arg244Ala mutant varied widely with the mutants (Table II).…”

Extended-Spectrum β-Lactamases in the 21st Century: Characterization, Epidemiology, and Detection of This Important Resistance Threat

“…There are at least 19 distinct inhibitorresistant TEM ␤-lactamases (for amino acid sequences for TEM, SHV and OXA extended-spectrum and inhibitor resistant ␤-lactamases, see http://www.lahey.org/studies/webt.htm). Inhibitor-resistant TEM ␤-lactamases have been found mainly in clinical isolates of E. coli, but also some strains of K. pneumoniae, Klebsiella oxytoca, P. mirabilis, and Citrobacter freundii (31,83). Although the inhibitor-resistant TEM variants are resistant to inhibition by clavulanic acid and sulbactam, thereby showing clinical resistance to the ␤-lactam-␤-lactamase inhibitor combinations of amoxicillin-clavulanate, ticarcillin-clavulanate, and ampicillin-sulbactam, they remain susceptible to inhibition by tazobactam and subsequently the combination of piperacillin and tazobactam (23,37).…”

In vitrosynergistic antibacterial activities of helvolic acid on multi-drug resistantStaphylococcus aureus

“…The resistance problems of other antibiotics and from other resistant mechanisms have not yet been solved. Moreover, clavulanic acid does not always prove effective now because of the emergence of the inhibitor resistant -lactamase (Bret, Chanel, Sirot, Labia, & Sirot, 1996;Lemozy et al, 1995). *Corresponding author.…”