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Transferable production of PER-1 beta-lactamase in Pseudomonas aeruginosa
Abstract:
PER-1, an extended-spectrum class A beta-lactamase, has been described only from Pseudomonas aeruginosa RNL-1, which was obtained in France in 1991. During studies on ceftazidime-resistant P. aeruginosa collected from December 1991 to July 1993 in Ankara, Turkey, we found 14 further isolates with PER-1 enzyme, as recognised by isoelectric point (pI 5.4), hydrolytic activity, gene hybridization and DNA sequence. Five of these isolates also had OXA-10 (PSE-2)-related enzymes and one hyper-produced ampC beta-lactamase, whereas eight had PER-1 enzyme alone. The last group, from four wards, appeared to be identical, giving the same DNA restriction patterns and carrying the PER-1 gene on an 8.5 kb HincII fragment. Two more producers were related but the other four were unique. In several representatives of the group of eight replicates, the PER-1 gene was shown to be encoded on a plasmid, larger than 154 kb in size, which transferred to P. aeruginosa PU21. A further isolate had the gene on an even larger conjugative plasmid. By contrast, the PER-1 gene reportedly was chromosomally-inserted in strain RNL-1. The PER-1 producers and their transconjugants were highly resistant to ceftazidime and aztreonam (MIC > or = 128 mg/L) but not to carbapenems or latamoxet. Piperacillin insusceptibility was marginal (MIC 8 mg/L). Clavulanate 4 mg/L, but not tazobactam 4 mg/L, reversed resistance to ceftazidime and carbenicillin. Purification of the enzyme to homogeneity was achieved by three ion exchanges and one gel filtration. We found much lower Vmax rates for aminothiazolyl cephalosporins than reported previously for PER-1 enzyme. This reflected the present assays being in 0.1 M phosphate buffer pH 7.0, whereas the previous were pH-stat-regulated; concentrated phosphate reduced enzyme activity against ceftazidime, but not against cephaloridine.
Insights
Extended-spectrum beta-lactamase PER-1 was identified in 14 new Pseudomonas aeruginosa isolates in Turkey. These PER-1 producing bacteria showed high resistance to ceftazidime and aztreonam, with clavulanate partially reversing this resistance.
Area of Science:
- Microbiology
- Molecular Biology
- Biochemistry
Background:
- The extended-spectrum beta-lactamase PER-1 was previously identified only in Pseudomonas aeruginosa RNL-1 from France.
- The emergence and spread of antibiotic resistance in Pseudomonas aeruginosa is a significant clinical concern.
Purpose of the Study:
- To investigate the prevalence and characteristics of PER-1 beta-lactamase producing Pseudomonas aeruginosa in Turkey.
- To analyze the genetic basis and resistance profiles of these newly identified isolates.
Main Methods:
- Detection of PER-1 enzyme using isoelectric focusing, hydrolytic activity assays, gene hybridization, and DNA sequencing.
- Plasmid analysis, conjugation experiments, and DNA restriction profiling.
- Determination of antibiotic minimum inhibitory concentrations (MICs) and enzyme kinetics.
Main Results:
- Fourteen new isolates of Pseudomonas aeruginosa producing PER-1 beta-lactamase were identified in Ankara, Turkey.
- Eight isolates possessed PER-1 alone, with identical DNA restriction patterns and carrying the gene on a plasmid.
- PER-1 producers exhibited high resistance to ceftazidime and aztreonam, with clavulanate demonstrating partial efficacy in reversing resistance.
Conclusions:
- The study identified a significant prevalence of PER-1 beta-lactamase in Pseudomonas aeruginosa in Turkey, suggesting potential for wider dissemination.
- The findings highlight the role of plasmids in the spread of PER-1 and associated resistance mechanisms.
- Enzyme kinetics revealed lower activity against aminothiazolyl cephalosporins compared to previous reports, potentially influenced by assay conditions.