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Emergence of resistance to imipenem in Pseudomonas aeruginosa
M J Lynch1, G L Drusano, H L Mobley
1Department of Medicine, University of Maryland School of Medicine, Baltimore 21201.
Abstract:
The emergence of resistance to imipenem by Pseudomonas aeruginosa was investigated with four pairs of isolates. Each pair represented pretherapy (susceptible) and posttherapy (resistant) specimens. In all cases, the imipenem-resistant isolates did not demonstrate changed susceptibilities to other beta-lactams. Agarose gel electrophoresis revealed no change in plasmid profiles between any pair of isolates. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the Sarkosyl-insoluble membrane protein revealed the loss of an outer membrane protein of apparent molecular mass 48 to 49 kilodaltons in posttherapy strains when grown with imipenem selection (5 micrograms/ml). There was no significant difference in the binding of [14C]imipenem to the penicillin-binding proteins of the pre- and posttherapy strains. Trichloroacetic acid precipitation of membranes isolated after growth in the presence of [14C]imipenem revealed that significantly less drug was bound to Sarkosyl-soluble membrane protein in three of the four posttherapy strains than the membrane proteins of the respective pretherapy strains. beta-Lactamase activity against imipenem at 100 or 3 microM was not detected in any isolate either with or without induction. These data suggest that resistance to imipenem is associated with the loss of a 48- to 49-kilodalton outer membrane protein accompanied by, in three of four cases, decreased penetration of the antibiotic across the outer membrane.
Insights
Pseudomonas aeruginosa developed imipenem resistance through the loss of a key outer membrane protein, hindering antibiotic entry. This resistance did not affect susceptibility to other beta-lactams.
Area of Science:
- Microbiology
- Molecular Biology
- Drug Resistance
Background:
- Pseudomonas aeruginosa is an opportunistic pathogen that frequently develops antibiotic resistance.
- Imipenem is a critical carbapenem antibiotic used to treat serious P. aeruginosa infections.
- Understanding resistance mechanisms is crucial for effective antimicrobial therapy.
Purpose of the Study:
- To investigate the mechanisms of imipenem resistance in Pseudomonas aeruginosa.
- To compare susceptible (pretherapy) and resistant (posttherapy) clinical isolates.
Main Methods:
- Agarose gel electrophoresis to analyze plasmid profiles.
- Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) to examine outer membrane proteins.
- Radioisotope labeling ([14C]imipenem) to assess drug binding to membrane proteins and penicillin-binding proteins.
- Beta-lactamase activity assays.
Main Results:
- Imipenem-resistant isolates showed no change in susceptibility to other beta-lactams or plasmid profiles.
- Loss of a 48- to 49-kilodalton outer membrane protein was observed in resistant strains.
- Reduced imipenem binding to Sarkosyl-soluble membrane proteins occurred in three of four resistant strains.
- No significant difference in beta-lactamase activity against imipenem was detected.
Conclusions:
- Imipenem resistance in P. aeruginosa is primarily associated with the downregulation or loss of a specific outer membrane protein.
- This protein loss likely contributes to decreased imipenem penetration into the bacterial cell.
- Resistance mechanisms do not involve altered beta-lactamase activity or changes in penicillin-binding proteins.