Efflux-Mediated Resistance to Tigecycline (GAR-936) in Pseudomonas aeruginosa PAO1

Abstract

Pseudomonas aeruginosa strains are less susceptible to tigecycline (previously GAR-936; MIC, 8 g/ml) than many other bacteria (P. J. Petersen, N. V. Jacobus, W. J. Weiss, P. E. Sum, and R. T. Testa, Antimicrob. Agents Chemother. 43:738-744, 1999). To elucidate the mechanism of resistance to tigecycline, P. aeruginosa PAO1 strains defective in the MexAB-OprM and/or MexXY (OprM) efflux pumps were tested for susceptibility to tigecycline. Increased susceptibility to tigecycline (MIC, 0.5 to 1 g/ml) was specifically associated with loss of MexXY. Transcription of mexX and mexY was also responsive to exposure of cells to tigecycline. To test for the emergence of compensatory efflux pumps in the absence of MexXY-OprM, mutants lacking MexXY-OprM were plated on medium containing tigecycline at 4 or 6 g/ml. Resistant mutants were readily recovered, and these also had decreased susceptibility to several other antibiotics, suggesting efflux pump recruitment. One representative carbenicillin-resistant strain overexpressed OprM, the outer membrane channel component of the MexAB-OprM efflux pump. The mexAB-oprM repressor gene, mexR, from this strain contained a 15-bp in-frame deletion. Two representative chloramphenicol-resistant strains showed expression of an outer membrane protein slightly larger than OprM. The mexCD-OprJ repressor gene, nfxB, from these mutants contained a 327-bp in-frame deletion and an IS element insertion, respectively. Together, these data indicated drug efflux mediated by MexCD-OprJ. The MICs of the narrower-spectrum semisynthetic tetracyclines doxycycline and minocycline increased more substantially than did those of tigecycline and other glycylcyclines against the MexAB-OprM-and MexCD-OprJ-overexpressing mutant strains. This suggests that glycylcyclines, although they are subject to efflux from P. aeruginosa, are generally inferior substrates for P. aeruginosa efflux pumps than are narrower-spectrum tetracyclines.Pseudomonas aeruginosa is a clinically important gram-negative opportunistic pathogen causing serious acute and chronic infections (8, 11). The exceptional array of intrinsic and acquired drug resistance mechanisms employed by P. aeruginosa renders antibiotic treatment of these infections problematic. One important resistance mechanism is efflux mediated by the so-called resistance nodulation division (RND) family of efflux pumps. Four RND pumps have been described in P. aeruginosa: MexAB-OprM (16), MexCD-OprJ (31), , and 25,40). RND pumps consist of an inner membrane transporter (MexB, MexD, MexF, and MexY), an outer membrane channel-forming component (OprM, OprJ, and OprN), and a membrane fusion protein (MexA, MexC, MexE, and MexX) (27). RND pumps show broad specificity, and their tripartite architecture allows extrusion of compounds directly from the cytoplasm to the external environment. Efflux pump action in P. aeruginosa leads to particularly high levels of drug resistance as a result of apparent synergism with the atypically impermeable outer membrane, which limits influx of antimic...

Effects of Efflux Transporter Genes on Susceptibility of Escherichia coli to Tigecycline (GAR-936)

“…It is unlikely that this inhibition is due to disruption of ⌬pH by tigecycline because tigecycline did not significantly affect the formation of the proton motive force in the membrane vesicle study, as already described for the quinacrine fluorescence assay. These results are in good agreement with those in a recent paper by Dean et al (6), who reported the tigecycline resistance conferred by MexXY and possibly MexAB, which are Pseudomonas homologues of AcrAB.…”

“…Tigecycline is a broadspectrum glycylcycline derivative (2) and is efficacious against highly resistant bacteria (1), including methicillin-resistant Staphylococcus aureus and penicillin-resistant Streptococcus pneumoniae. However, it is less active against clinically problematic opportunistic pathogens, such as Pseudomonas aeruginosa (6) and Proteus mirabilis (3,28).…”

Efflux pumps may play a role in tigecycline resistance in Burkholderia species

“…It is effective against many Gram-positive and Gram-negative microorganisms but has little activity against the non-fermenting CF pathogens P. aeruginosa and Burkholderia spp. The mechanism of resistance has been clearly demonstrated for P. aeruginosa, but in BCC it remains to be determined [4,10]. The data presented here indicate a role for efflux pumps.…”

“…A c c e p t e d M a n u s c r i p t 4 The aim of this study was to determine whether tigecycline resistance in BCC is medicated by efflux pumps and, if so, whether EPIs can be used to augment the efficacy of the glycylcycline molecule.…”

“…Tigecycline, a novel expanded broad-spectrum glycylcycline antibiotic, has been shown to be active against a broad range of Gram-negative, Gram-positive, anaerobic and atypical bacteria [3]. However, its activity against P. aeruginosa is poor, with the microorganism's inherent resistance attributed to the activity of broad-substrate efflux pumps [4]. The activity of tigecycline against B. cepacia complex (BCC) has also been reported to be poor [5], but the mechanisms of resistance are poorly defined.…”