Plasmid-mediated gentamicin resistance of Pseudomonas aeruginosa and its lack of expression in Escherichia coli

Abstract

We isolated 11 nonconjugative plasmids mediating resistance to aminoglycoside antibiotics, including gentamicin, from Pseudomonas aeruginosa strains. Their genetic properties were investigated in both P. aeruginosa and Escherichia coli transformants. The plasmid molecular weights ranged from 11 x 106 to 24 x 106. A low level or complete absence of gentamicin resistance was observed when these plasmids were introduced into E. coli, but gentamicin resistance was restored when the plasmids were transferred back to P. aeruginosa from E. coli. Aminoglycoside-modifying enzyme activity was detected in P. aeruginosa harboring these plasmids, but was absent or greatly reduced in E. coli strains. This lack of expression may explain the observed decrease in aminoglycoside resistance.Plasmids isolated from Pseudomonas aeruginosa have been classified into 11 incompatibility groups. Most of the conjugative plasmids are nontransmissible to Escherichia coli strains, except for plasmids of the P-1 or P-3 group (12). Three reasons for this nontransferability to E. coli can be postulated: (i) nontransmissibility owing to some differences in cell surface structure; (ii) no replication of the plasmids after transmission to E. coli strains; and (iii) failure of resistance gene expression in the E. coli system. Some of the plasmids mobilized to E. coli by the presence of P-1 group plasmids can replicate by themselves in that host (12,13,16). In contrast, many plasmids are incapable of replication in E. coli, except by recombination with the mobilizing plasmid (13,20).Recently the isolation frequency of gentamicin-resistant P. aeruginosa strains has increased (3,4,14,18,21,24,26). We have also isolated plasmids mediating gentamicin resistance and transferable by conjugation to a recipient P. aeruginosa strain in 20% of the 91 gentamicinresistant P. aeruginosa isolates from clinical specimens in Japan. Gentamicin-resistance plasmids which were transfer deficient were also found in 56% of the gentamicin-resistant strains. In this report, we examined the genetic properties of such nonconjugative plasmids mediating gentamicin resistance in P. aeruginosa and also in E. coli after being introduced by transformation.MATERIALS AND METHODS Media. Heart infusion agar (Eiken Kagaku Co., Tokyo, Japan) and peptone water were used for the determination of drug resistance. Peptone water con-

AAC(6′)-Iaf, a Novel Aminoglycoside 6′- N -Acetyltransferase from Multidrug-Resistant Pseudomonas aeruginosa Clinical Isolates

“…Drug resistance was transferred from P. aeruginosa clinical isolates to a rifampin-resistant mutant of P. aeruginosa, ATCC 27853 Rfp r , using the broth mating method (26). The transconjugants were selected on Mueller-Hinton agar plates containing rifampin (200 g/ml) and IPM (16 g/ml) or AMK (20 g/ml).…”

Transferable imipenem resistance in Pseudomonas aeruginosa

“…Conjugation and transformation were carried out by the method of Kato et al (12). Plasmid pMS350 was transferred to P. aeruginosa PAOlNf, * Corresponding author.…”

“…Preparation of plasmid DNAs and agarose gel electrophoresis were carried out by the method of Kato et al (12). Plasmids RP4 (36 MDa), pMS108 (24 MDa), pMS101 (14 MDa) (10), and Rlb679 (5.6 MDa) were used as standards to determine the size of pMS350.…”

Mechanisms of high-level resistance to quinolones in urinary tract isolates of Pseudomonas aeruginosa

“…Competent cells of norfloxacin-resistant (norfloxacin MIC, >1.56 ,ug/ml) P. aeruginosa strains were prepared by using early-log-phase cells in L broth. Transformation was carried out with 100 mM MgCl2-treated P. aeruginosa (9). The transformants were selected on bromthymol blue agar containing 400 ,ug of carbenicillin per ml, which was the selective marker for pPAW207 and pNF111 plasmids.…”