Two clinical strains of Enterobacter aerogenes that exhibited phenotypes of multiresistance to ␤-lactam antibiotics, fluoroquinolones, chloramphenicol, tetracycline, and kanamycin were investigated. Both strains showed a porin pattern different from that of a susceptible strain, with a drastic reduction in the amount of the major porin but with an apparently conserved normal structure (size and immunogenicity), together with overproduction of two known outer membrane proteins, OmpX and LamB. In addition, the full-length Opolysaccharide phenotype was replaced by a semirough Ra phenotype. Moreover, in one isolate the intracellular accumulation of chloramphenicol was increased in the presence of the energy uncoupler carbonyl cyanide m-chlorophenylhydrazone, suggesting an energy-dependent efflux of chloramphenicol in this strain. The resistance strategies used by these isolates appear to be similar to that induced by stress in Escherichia coli cells.Bacteria have developed various regulatory systems which coordinate their adaptive responses with the different environmental stresses to which they are exposed. Recently, the wholegenome transcriptional profiles, or "transcriptomes," were determined in vitro for an Escherichia coli strain exposed to an inducer of the soxRS or marRAB system and an E. coli strain constitutively expressing MarA (7,39). Under these conditions, the expression of several genes appeared to be significantly activated or down-regulated. These modulations of gene expression alter the sensitivities of the bacteria to a broad range of antibiotics (1, 2).Enterobacter aerogenes is one of the more frequently described gram-negative bacteria responsible for nosocomial respiratory tract infections (5, 17). In the last 5 years, it has been shown that clinical isolates of this species, which are naturally resistant to aminopenicillins through their production of a chromosomal cephalosporinase, often express an extendedspectrum ␤-lactamase, TEM-24, which gives rise to resistance to ␤-lactam antibiotics (5,17,19,31). Moreover, E. aerogenes exhibits acquired resistance to other families of antimicrobial agents. Previous studies have reported that clinical strains exhibiting an efflux process are resistant to ␤-lactam antibiotics, quinolones, tetracycline, and chloramphenicol (12, 29). Drug efflux can be coincident with a drastic reduction in drug uptake due to the loss of porin content (16,22,23,29).Two E. aerogenes clinical isolates, isolates 117 and 119, were selected from up to 100 strains isolated in our laboratory (12). These two strains exhibited a phenotype of multiresistance to ␤-lactam antibiotics, fluoroquinolones, chloramphenicol, tetracycline, and kanamycin comparable to that of E. aerogenes strains lacking nonspecific porins or expressing mutated porins (12, 29).The aim of this work was to examine some factors that may contribute to the resistance to antimicrobial agents in these two E. aerogenes clinical strains. MATERIALS AND METHODSBacterial strains, growth conditions, and antibiotic suscepti...

The role of the AcrAB-TolC pump in macrolide and ketolide susceptibility in Escherichia coli and Enterobacter aerogenes was studied. Efflux pump inhibitor restored erythromycin, clarithromycin, and telithromycin susceptibilities to multidrug-resistant isolates. No modification of telithromycin accumulation was detected in E. aerogenes acrAB or tolC derivatives compared to that in the parental strain. Two independent efflux pumps, inhibited by phenylalanine arginine β-naphthylamide, expel macrolides and telithromycin in E. aerogenes

Multidrug resistance (MDR) in Enterobacter aerogenes can be mediated by induction of MarA, which is triggered by certain antibiotics and phenolic compounds. In this study, we identified the gene encoding RamA, a 113-amino-acid regulatory protein belonging to the AraC-XylS transcriptional activator family, in the Enterobacter aerogenes ATCC 13048 type strain and in a clinical multiresistant isolate. Overexpression of RamA induced an MDR phenotype in drug-susceptible Escherichia coli JM109 and E. aerogenes ATCC 13048, as demonstrated by 2-to 16-fold-increased resistance to ␤-lactams, tetracycline, chloramphenicol, and quinolones, a decrease in porin production, and increased production of AcrA, a component of the AcrAB-TolC drug efflux pump. We show that RamA enhances the transcription of the marRAB operon but is also able to induce an MDR phenotype in a mar-deleted strain. We demonstrate here that RamA is a transcriptional activator of the Mar regulon and is also a self-governing activator of the MDR cascade.Multidrug resistance (MDR) relates to all bacterial species but has been studied mostly in members of the Enterobacteriaceae, particularly in E. coli with the study of the mar operon (1,17,18,32). The MDR phenotype often associates a decrease in porin synthesis with an increased activity of efflux pumps to restrict the intracellular concentration of various antibiotics, including ␤-lactams, tetracyclines, chloramphenicol, and quinolones. Resistance results from activation of the chromosomal genes by induction or mutation and by DNA transfer (21). A single drug treatment can lead to cross-resistance to other unrelated antibiotics (16). In clinical isolates which exhibit high resistance to broad-spectrum antibiotics, MDR is the result of enzymatic responses, mutations in the antibiotic target, and modifications in envelope permeability, including porin alteration and induction of drug efflux (7,20,23). In such a case, the existence of clinical strains resistant to several structurally unrelated antibiotics contributes to treatment failures in human infections, bacterial dissemination, outbreaks, and changes in patients' flora due to the selective advantage of the microorganism.The mar regulon identified in E. coli plays a key role in the expression of the multidrug resistance phenotype. MarA expression induces the synthesis of the micF antisense RNA that downregulates the OmpF outer membrane porin, on one hand, and, on the other hand, overproduction of the AcrAB-TolC drug efflux pump (1,3,17). Mutations located in the MarR repressor leading to constitutive expression of the marRAB operon have been described in fluoroquinolone-resistant isolates of E. coli and Enterobacter cloacae (20,25). The Rob and SoxS regulatory proteins are MarA homologues and participate in E. coli MDR regulation (1,16,28).Among the emerging resistant bacteria, E. aerogenes is now the third leading cause of nosocomial respiratory tract infections (6, 11). A TEM-24-producing clone was the agent of a hospital pandemic (6, 13). Moreover, a c...

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The ompX gene of Enterobacter aerogenes was cloned. Its overexpression induced a decrease in the major porin Omp36 production and consequently a b-lactam resistance was noted. Purified outer membrane protein X (OmpX) was reconstituted into artificial membranes and formed ion channels with a conductance of 20 pS in 1 M NaCl and a cationic selectivity. Both MarA expression and high osmolarity induced a noticeable increase of the OmpX synthesis in the E. aerogenes ATCC 13048 strain. In addition, OmpX synthesis increased under conditions in which the expression of the E. aerogenes major non-specific porins, Omp36 and Omp35, decreased.