Activity of the carbapenem panipenem and role of the OprD (D2) protein in its diffusion through the Pseudomonas aeruginosa outer membrane

Abstract

Evidence of permeation of panipenem through the OprD (D2) channel of Pseudomonas aeruginosa outer membrane was shown by using OprD protein-producing and -nonproducing strains which contained plasmid pHN4, which codes for L-1 13-lactamase of Xanthomonas maltophilia. Permeation by panipenem was determined by measuring hydrolysis of the carbapenem by 13-lactamase in the periplasmic space. Permeation by panipenem was also determined by counting uptake of [14C]panipenem into P. aeruginosa PAO1 and its OprD protein-deficient mutant, and this permeation of PAO1 was inhibited by L-lysine. These results indicate that panipenem, as well as imipenem, uses the OprD channel, which functions as a specific channel for diffusion of basic amino acids. Panipenem and imipenem showed stronger activities against PAO1 and clinical isolates in human serum than in Mueller-Hinton broth, which contains more amino acids than human serum does. The activities of the carbapenems were reduced by addition of L-lysine to human serum. Similar results were obtained with mouse serum and ascitic fluid. In contrast, such a change in the activities of carbapenems was not observed with an OprD protein-deficient mutant, suggesting that the main reason for the strong activities of carbapenems in biological fluids is a decrease in competition between the antibiotics and basic amino acids for permeation through the OprD channel. Panipenem and imipenem showed much stronger therapeutic efficacies against experimental infections with P. aeruginosa in mice than did the reference antibiotics. Their in vivo activities were more consistent with their MICs in biological fluids than with those in Mueller-Hinton broth.Infections by Pseudomonas aeruginosa are serious clinical problems, particularly in compromised hosts. In recent years, many new extended-spectrum penicillins and cephalosporins have been introduced into clinical practice. Some of them show potent antipseudomonal activity in vitro. However, only a few antibiotics show adequate clinical efficacy against pseudomonal infections. Carbapenems, such as panipenem (formerly RS-533 or CS-533) and imipenem, have been shown to have potent antipseudomonal activity both in vitro and in vivo (2,4,8,9,12,14,15,17,22).We reported previously that panipenem and imipenem showed much stronger antipseudomonal activities in lowamino-acid media, such as a minimal medium, than in Mueller-Hinton medium (5). This mechanism was investigated by using P. aeruginosa PAO1 and its OprD (D2) protein-deficient mutants, and it was assumed that the increase in the antipseudomonal activities of the carbapenems in low-amino-acid media relates to lessened competition between the carbapenems and basic amino acids for permeation through the OprD channel of the P. aeruginosa outer membrane. Trias and Nikaido reported that imipenem permeated the outer membrane of P. aeruginosa through the OprD channel (20) and that the permeation was competitively inhibited by basic amino acids (21). Concentrations of free basic amino acids in human serum are ...

Phenotypic Heterogeneity Enables Uropathogenic Escherichia coli To Evade Killing by Antibiotics and Serum Complement

“…At the same time, growth in serum has been shown to induce expression of antibiotic efflux pumps in P. aeruginosa and Acinetobacter, suggesting enhanced antibiotic tolerance during infection (52). Beside induction of efflux pumps, the growth in serum has been shown to enhance expression of OprD porin in P. aeruginosa (53). Recent study has shown the mutation in oprD gene in P. aeruginosa conferred resistance to carbapenem antibiotics, as well as enhanced resistance to killing by human serum (54).…”

OmpA Is the Principal Nonspecific Slow Porin of Acinetobacter baumannii

“…aeruginosa also shows high intrinsic resistance levels to earlier cephalosporins, but it used to be quite susceptible to imipenem. It was discovered that imipenem bypasses the inefficient nonspecific porin OprF and uses a basic amino acid channel, OprD (14,45,46). In the organisms with slow major porins, we expect to see numerous specific channels for the uptake of various nutrients (16), and imipenem "hijacks" one of them.…”

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

“…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).…”

“…To compensate for this low permeability and to permit the uptake of essential nutrients available at low concentrations in the environment, certain substrate-specific transport systems are present in the P. aeruginosa outer membrane. For example, OprP allows the facilitated permeation of phosphate (4), OprB (formerly called protein D1) has a binding site for glucose (13,34), iron-repressible outer membrane proteins (IROMP) permit uptake of Fe3+-siderophore complexes when produced under the conditions of iron deprivation (35), and OprD (formerly called protein D2) (12,14) facilitates the diffusion of basic amino acids and small peptides containing these amino acids (32). These channels may be utilized by compounds which are structural analogs of the natural substrate.…”