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Method for estimation of low outer membrane permeability to beta-lactam antibiotics
Bernard Lakaye1, Alain Dubus, Bernard Joris
1Laboratoire d'Enzymologie and Centre d'Ingénierie des Protéines, Université de Liège, Institut de Chimie, B-4000 Liège, Belgium.
Antimicrobial Agents and Chemotherapy
|August 17, 2002
Summary
A new method quantifies outer membrane permeability in gram-negative bacteria, crucial for understanding beta-lactam antibiotic resistance. This technique accurately measures low permeability, aiding in predicting antibiotic efficacy against resistant strains.
Area of Science:
- Microbiology
- Bacterial Physiology
- Antibiotic Resistance
Background:
- The outer membrane of gram-negative bacteria is a key factor in beta-lactam antibiotic resistance.
- Existing methods for quantifying outer membrane permeability are limited, especially for low permeability values.
- Secreted and cell surface-associated beta-lactamases complicate accurate permeability measurements.
Purpose of the Study:
- To develop and validate a novel method for assessing low outer membrane permeability in gram-negative bacteria.
- To measure the permeability coefficients of various beta-lactam antibiotics across the outer membranes of Enterobacter cloacae and Enterobacter aerogenes.
- To demonstrate the utility of the new method in predicting minimum inhibitory concentrations (MICs).
Main Methods:
- Utilized the acylation of a sensitive periplasmic penicillin-binding protein (PBP), BlaR-CTD, to measure beta-lactam penetration rates.
- Focused on conditions where outer membrane diffusion is the rate-limiting step for method validity.
- Applied the method to beta-lactamase-deficient strains of Enterobacter cloacae and Enterobacter aerogenes using six different beta-lactam antibiotics.
Main Results:
- Established a new, sensitive method for measuring low outer membrane permeability, with BlaR-CTD acylation as the indicator.
- Determined permeability coefficients for benzylpenicillin, ampicillin, carbenicillin, cefotaxime, aztreonam, and cephacetrile.
- Observed significantly lower outer membrane permeability in E. cloacae compared to E. coli for several antibiotics, including a record low for aztreonam (P = 3.2 x 10(-9) cm/s).
Conclusions:
- The BlaR-CTD acylation method is effective for quantifying low outer membrane permeability in gram-negative bacteria.
- Outer membrane permeability significantly impacts the efficacy of beta-lactam antibiotics against specific bacterial strains.
- The method successfully predicted MICs for a beta-lactamase-overproducing strain, validating its clinical relevance.