Resistance of <i>Pseudomonas aeruginosa</i> to imipenem is independent of β-lactamase production

Büscher, K.-H.; Cullmann, W.; Opferkuch, W.

doi:10.1093/jac/19.5.700

Cited by 6 publications

(7 citation statements)

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“…It has been suggested that a protein belonging to the D group of outer membrane proteins is responsible for the permeation of imipenem (7)(8)(9)19). The synthesis of protein Dl is induced by the presence of glucose in growth medium "The hydrolysis rates by leaked-out enzymes (measured with supernatants obtained by centrifugation of cell suspensions) were between 20 to 40% of the rates measured with intact cells of strain 3-C for the hydrolysis of cephaloridine and no more than 10% for the hydrolysis of imipenem.…”

Section: Resultsmentioning

confidence: 99%

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Decreased outer membrane permeability in imipenem-resistant mutants of Pseudomonas aeruginosa

Trias

Dufresne

Levesque

et al. 1989

Antimicrob Agents Chemother

103

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The outer membrane of imipenem-resistant mutants of Pseudomonas aeruginosa was shown to have decreased permeability to imipenem but not to cephaloridine. These experiments were performed with intact cells and liposomes containing imipenem-hydrolyzing beta-lactamase derived from Pseudomonas maltophilia, in both cases utilizing an imipenem concentration of 50 microM. In contrast, liposome swelling assays using imipenem at 8 mM detected no significant difference between the imipenem-resistant mutants and their parents. It was found that the outer membrane of P. aeruginosa has a saturable specific channel through which imipenem travels mainly at low concentrations, whereas at high concentrations this pathway is obscured by diffusion through nonspecific porin channels.

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Section: Resultsmentioning

confidence: 99%

“…These specifically imipenem-resistant strains cannot hydrolyze imipenem and are not altered in the binding of imipenem to penicillin-binding proteins (9,25). In contrast, they exhibit diminished production of a 45-to 49-kilodalton outer membrane protein when compared with the suscepti-* Corresponding author.…”

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confidence: 99%

Decreased outer membrane permeability in imipenem-resistant mutants of Pseudomonas aeruginosa

Trias

Dufresne

Levesque

et al. 1989

Antimicrob Agents Chemother

103

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“…However, during clinical therapy of P. aeruginosa, imipenem-resistant isolates arise at a significant rate (22). The major cause of resistance is loss of a specific outer membrane protein, OprD (5)(6)(7)15 such as meropenem and panipenem (12,31,33). Nevertheless, it was suggested that OprD had significant nonspecific permeability to monosaccharides (and disaccharides) and was generally involved as a porin for antibiotic (13-lactam) uptake (24,33,38,39).…”

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confidence: 99%

Genetic definition of the substrate selectivity of outer membrane porin protein OprD of Pseudomonas aeruginosa

Huang

Hancock

1993

J Bacteriol

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Earlier studies proved that Pseudomonas aeruginosa OprD is a specific porin for basic amino acids and imipenem. It was also considered to function as a nonspecific porin that allowed the size-dependent uptake of monosaccharides and facilitation of the uptake of quinolone and other antibiotics. In the present study, we utilized P. aeruginosa strains with genetically defined levels of OprD to characterize the in vivo substrate selectivity of this porin. An oprD::fQ interposon mutant was constructed by gene replacement utilizing an in vitro mutagenized cloned oprD gene. In addition, OprD was overexpressed from the lac promoter by cloning the oprD gene into the broad-host-range plasmid pUCP19. To test the substrate selectivity, strains were grown in minimal medium with limiting concentrations of the carbon sources glucose, gluconate, or pyruvate. In minimal medium with 0.5 mM gluconate, the growth rates of the parent strain H103 and its oprD::fQ mutant H729 were only 60 and 20%, respectively, of that of the OprD-overexpressing strain H103(pXH2). In contrast, no significant differences were observed in the growth rates of these three strains on glucose or pyruvate, indicating that OprD selectively facilitated the transport of gluconate. To determine the role of OprD in antibiotic uptake, nine strains representing different levels of OprD and OprF were used to determine the MICs of different antibiotics. The results clearly demonstrated that OprD could be utilized by imipenem and meropenem but that, even when substantially overexpressed, it could not be significantly utilized by other P-lactams, quinolones, or aminoglycosides. In addition, competition experiments confirmed that imipenem had common binding sites with basic amino acids in the OprD channel, but not with gluconate or glucose.

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“…These are characteristics of proteins of the D group in the P. aeruginosa outer membrane (10,11), and it has been suggested that imipenem uses protein(s) belonging to this group for influx into the periplasm (4)(5)(6). Two major proteins belong to the D group, Dl and D2.…”

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confidence: 99%

“…Liposomes were made by suspending a dried film containing 6 ,umol of egg with Triton X-100-EDTA buffer and eluted from an anionexchange column with a gradient of NaCl. Protein D2 eluted first at a lower ionic strength (0.1 M NaCe), and we were able to remove it completely from the column before the elution of protein Dl.…”

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confidence: 99%

Outer membrane protein D2 catalyzes facilitated diffusion of carbapenems and penems through the outer membrane of Pseudomonas aeruginosa

Trias

Nikaido

1990

Antimicrob Agents Chemother

276

198

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The outer membrane of imipenem-resistant mutants ofPseudomonas aeruginosa with decreased permeability to imipenem was shown by Western (immuno-) blotting to contain protein Dl and to lack protein D2. Protein D2 was purified and was shown to allow the permeation of imipenem at a rate higher than expected from its molecular weight. Spontaneous imipenem-resistant mutants of P. aeruginosa PAO1 appeared at a frequency of 10-8 in the laboratory and did not synthesize protein D2. Experiments performed with intact cells carrying plasmid pHN4 containing the gene for L-1 ,-lactamase from Pseudomonas maltophiia showed that this channel could also be used by SM-7338, Sch 33755, and Sch 33440 but apparently not by Sch 34343 or Sch 29482.Nutrients and waste products have to go through the outer membrane of gram-negative bacteria to reach the cytoplasm and the outer medium, respectively. This membrane is semipermeable and acts as a molecular sieve allowing the passage of small hydrophilic molecules (21). The overall permeability of the outer membrane usually depends on the number and properties of the pore-forming proteins that are generically called porins (21). Although most of the small hydrophilic molecules utilize this nonspecific pathway, there are some specific channels that allow the passage of compounds that have an insufficient or negligible rate of permeation through the porin channels and of compounds that are found in the external medium at low concentrations (21). Some of these specific channels are synthesized at higher levels when the substrate is present in the medium of growth: for example, the well-known lambda phage receptor of Escherichia coli, protein LamB, which allows the passage of maltose and maltodextrins (16), and the Dl protein of Pseudomonas aeruginosa, which allows the permeation of glucose (26), are induced by maltose and glucose, respectively. Other specific permeation systems, such as the Tsx protein that shows a high specificity for nucleosides in E. coli (18) and the TonB-related iron and cobalamine uptake systems of the outer membrane, function in the uptake of building blocks for the cell rather than of substrates for energy generation. They are derepressed when the substrate is missing from the medium or present at very low concentrations. The TonB-dependent transport systems in E. coli are energy dependent and include the following outer mem-

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Resistance of Pseudomonas aeruginosa to imipenem is independent of β-lactamase production

Cited by 6 publications

References 0 publications

Decreased outer membrane permeability in imipenem-resistant mutants of Pseudomonas aeruginosa

Decreased outer membrane permeability in imipenem-resistant mutants of Pseudomonas aeruginosa

Genetic definition of the substrate selectivity of outer membrane porin protein OprD of Pseudomonas aeruginosa

Outer membrane protein D2 catalyzes facilitated diffusion of carbapenems and penems through the outer membrane of Pseudomonas aeruginosa

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