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Broad-host-range gyrase A gene probe.

N J Robillard1

  • 1Pharmaceutical Division, Miles Inc., West Haven, Connecticut 06516.

Antimicrobial Agents and Chemotherapy
|October 1, 1990
PubMed
Summary
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The cloned Escherichia coli gyrase A gene, expressed in Pseudomonas aeruginosa, restores quinolone susceptibility in specific mutants. This DNA gyrase A probe effectively detects quinolone resistance mutations in clinical P. aeruginosa isolates.

Area of Science:

  • Microbiology
  • Molecular Biology
  • Genetics

Background:

  • Quinolone resistance in Pseudomonas aeruginosa is a significant clinical concern.
  • Understanding the genetic basis of quinolone resistance is crucial for developing effective treatments.
  • The DNA gyrase A subunit is a primary target for quinolone antibiotics.

Purpose of the Study:

  • To investigate the expression and function of the Escherichia coli gyrase A gene in Pseudomonas aeruginosa.
  • To develop a tool for detecting quinolone resistance mechanisms in P. aeruginosa.
  • To differentiate between gyrase A mutations and permeability alterations as causes of quinolone resistance.

Main Methods:

  • Cloning of the E. coli gyrase A gene into a broad-host-range cosmid vector (pLA2917).

Related Experiment Videos

  • Mobilization of the resulting plasmid (pNJR3-2) into various P. aeruginosa strains, including wild-type and quinolone-resistant mutants.
  • Assessment of quinolone susceptibility in engineered P. aeruginosa strains.

    • Main Results:

    • The E. coli gyrase A gene (on pNJR3-2) conferred wild-type quinolone susceptibility on P. aeruginosa gyrA mutants.
    • Intermediate susceptibility was observed in cfxA-cfxB double mutants, suggesting a role for both genes.
    • pNJR3-2 did not affect quinolone susceptibility in wild-type or permeability mutants (cfxB).
    • The probe successfully analyzed quinolone resistance in clinical P. aeruginosa isolates.

    Conclusions:

    • The E. coli DNA gyrase A gene is expressed in P. aeruginosa and functionally complements gyrA mutations.
    • The pNJR3-2 plasmid serves as a valuable probe for identifying gyrase A mutations conferring quinolone resistance.
    • This approach can distinguish between gyrase A-mediated resistance and resistance due to altered permeability.