A pleiotropic, posttherapy, enoxacin-resistant mutant of Pseudomonas aeruginosa

Abstract

An enoxacin-resistant Pseudomonas aeruginosa mutant (G49) isolated during patient therapy was characterized in detail. The G49 mutant was cross resistant to several classes of antibiotics including quinolones, I-lactams, chloramphenicol, and tetracycline, but not imipenem or aminoglycosides. Compared with its paired pretherapy isolate G48, this mutant had several alterations in outer membrane proteins including a complete loss of the major porin protein OprF and a substantially altered lipopolysaccharide profile. Revertants were selected at a frequency of approximately 1% after enrichment for OprF+ cells on low-salt proteose peptone no. 2 medium. Ninety-seven of these OprF+ revertants were as susceptible to carbenicillin and norfloxacin as the pretherapy isolate. One of these revertants was characterized in more detail and shown to be indistinguishable in all properties from the pretherapy isolate. It is proposed that the multiple-antibiotic-resistance (Mar) phenotype of this mutant resulted from a single pleiotropic mutation.Enoxacin is a difluorinated quinolone with strong activity for gram-negative bacteria (7,41). Enoxacin is now marketed in 11 countries, including South America, the United Kingdom, and other parts of Europe, for use in the therapy of urinary tract infections and in some countries for respiratory tract infections. As in other bacteria, the primary target for quinolones in Pseudomonas aeruginosa is DNA gyrase. nalA, nfxA, norA, and cipA are alleles ofgyrA and encode A subunits that are less susceptible to inhibition by quinolones (14,16,33,34). Other mutations affecting quinolone activity, but not DNA gyrase, in P. aeruginosa have also been described. Many laboratory mutants contain alleles of nalB and require nalidixic acid MICs of >500 ,ug/ml and are cross resistant to carbenicillin, ureidopenicillins, chloramphenicol, and novobiocin (33, 34). Resistant strains have arisen during experimental P. aeruginosa infections, with several classes of mutants isolated with mutations resembling naL4 and nfxC (10,22,23). Quinolone-resistant P. aeruginosa isolated from patients may also have the nal4 or nalB phenotype (43) or a nalB-like phenotype lacking OprF. Such strains have been isolated from patients with chronic obstructive airway disease (30), burn wound sepsis (17), cystic fibrosis (6), and empyema (20). While several strains with decreased susceptibilities to several agents have been isolated, which suggests a mutation in nalB, there are biochemical differences that are commensurate with different mutations encoding multiple resistance. Some quinolone-resistant P. aeruginosa strains contain a new 54-kDa outer membrane protein (14, 18) or lack a 31.5-kDa outer membrane protein (9, 17), possibly OprF, or have decreased levels of proteins D2 and Hi or Gl (5,20,23). Several workers have also reported changes in lipopolysaccharide (LPS) (6,18,34).An open study to evaluate the efficacy of enoxacin in respiratory tract infections with particular attention to those caused by P. aeruginosa was perform...

The Amino Terminus of Pseudomonas aeruginosa Outer Membrane Protein OprF Forms Channels in Lipid Bilayer Membranes: Correlation with a Three-Dimensional Model

“…OprF is a major outer membrane protein in Pseudomonas aeruginosa that has been studied extensively due to its proposed utility as a vaccine component, its role in antimicrobial drug resistance, and its porin function (3,6,7,13,20). It has been shown to be required for cell growth in low-osmolarity medium and for the maintenance of cell shape (21).…”

Clinically Relevant Chromosomally Encoded Multidrug Resistance Efflux Pumps in Bacteria

“…Evidence that may support a global regulator involved in efflux-mediated antibiotic resistance in P. aeruginosa is provided by several studies with clinical isolates. First, an isolate for P. aeruginosa from a chronic obstructive airways disease patient overexpressed not only the MexAB-OprM efflux pump but also the MexEF-OprN pump and had decreased expression of a porin protein (OprF), plus a mutation in a topoisomerase gene (161,173,174). By complementing various genes, the role of each mechanism was determined.…”

Differential Impact of MexB Mutations on Substrate Selectivity of the MexAB-OprM Multidrug Efflux Pump of Pseudomonas aeruginosa

“…To date, seven RND multidrug efflux systems have been characterized in P. aeruginosa, including 27,46,47), MexCD-OprJ (45), MexEFOprN (24), MexXY-OprM (2, 36, 63), MexJK-OprM (4), and, most recently, MexHI-OpmD (1, 56) and MexVW-OprM (31). The major system contributing to intrinsic multidrug resistance is encoded by the mexAB-oprM operon, which is also hyperexpressed in so-called nalB-type multidrug-resistant mutants (18,19,23,33,34,41,49,52,68). These mutants carry mutations in the mexR gene (19,49,53,60,68), which encodes a repressor of mexAB-oprM (60).…”