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Novel resistance to imipenem associated with an altered PBP-4 in a Pseudomonas aeruginosa clinical isolate
F Bellido1, C Veuthey, J Blaser
1Department of Microbiology, Medical School, University of Geneva, Switzerland.
Abstract:
A Pseudomonas aeruginosa (isolate 416) from a patient with pneumonia, was initially susceptible to imipenem (MIC: 2 mg/l) but became resistant to this antibiotic (isolate 470, MIC: 32 mg/l) during imipenem therapy. Treatment failed. No parallel increases in MIC were observed for other antimicrobials tested. Isolates 416 and 470 shared the same pyocin type and serotype, produced small amounts of an inducible beta-lactamase, and had similar lipopolysaccharide compositions. On electrophoresis of outer membrane proteins, the porin F, identified by the monoclonal antibody MA4-4, was expressed similarly by the two isolates but the production of one band (apparent molecular weight: 47,000) was diminished in isolate 470. [14C]-Imipenem labelling of intact cells proceeded more slowly in 470 than in 416, especially when bacterial cells were treated by antibody MA4-4 to block the porin F channel. [14C]-Imipenem labelling of penicillin binding proteins (PBP) showed that the band identified as PBP-4 bound markedly less radioactivity in isolate 470 than in 416. After isolate 470 was passaged several times in antibiotic-free broth, the imipenem MIC was decreased from 32 to 8 mg/l, and the [14C]-imipenem PBP pattern recovered the initial profile as exhibited by isolate 416. Two resistance mechanisms, affecting imipenem electively, could have combined their effect in the post-therapy isolate, altered target protein and reduced permeability.
Insights
Pseudomonas aeruginosa developed imipenem resistance during therapy due to reduced permeability and altered penicillin-binding protein 4. This resistance reversed upon antibiotic withdrawal, suggesting combined mechanisms.
Area of Science:
- Microbiology
- Antibiotic Resistance
- Molecular Biology
Background:
- Pseudomonas aeruginosa can develop antibiotic resistance during treatment.
- Imipenem is a critical antibiotic for treating serious infections.
Purpose of the Study:
- To investigate the mechanisms of imipenem resistance development in Pseudomonas aeruginosa during therapy.
- To identify specific molecular changes associated with acquired imipenem resistance.
Main Methods:
- Comparative analysis of pre- and post-therapy Pseudomonas aeruginosa isolates.
- Minimum Inhibitory Concentration (MIC) determination for imipenem and other antimicrobials.
- Outer membrane protein electrophoresis and identification.
- Radiolabeling studies with [14C]-Imipenem to assess cell permeability and penicillin-binding protein (PBP) interactions.
Main Results:
- The post-therapy isolate (470) showed significantly increased imipenem MIC compared to the pre-therapy isolate (416).
- Reduced expression of a 47,000 MW outer membrane protein band and decreased [14C]-imipenem labeling of PBP-4 were observed in isolate 470.
- Imipenem resistance and PBP-4 labeling patterns were partially reversed after passaging isolate 470 in antibiotic-free broth.
Conclusions:
- Acquired imipenem resistance in Pseudomonas aeruginosa during therapy likely resulted from a combination of reduced bacterial permeability and alterations in the penicillin-binding protein 4 target.
- These resistance mechanisms appear to be reversible, offering potential insights for future therapeutic strategies.