nfxC-type cells of Pseudomonas aeruginosa that produce the MexEF-OprN efflux pump exhibit resistance to fluoroquinolones and chloramphenicol and hypersusceptibility to most classical ␤-lactam antibiotics. We investigated the molecular mechanism of how the nfxC mutation causes ␤-lactam hypersusceptibility. The MexAB-OprM extrusion pump transports and confers resistance to ␤-lactam antibiotics. Interestingly, expression of the mexAB-oprM operon reached the highest level during the mid-stationary growth phase in both wild-type and nfxC-type mutant strains, suggesting that expression of the mexAB-oprM operon may be controlled by cell density-dependent regulation such as quorum sensing. This assumption was verified by demonstrating that exogenous addition of the quorum-sensing autoinducer N-butyryl-L-homoserine lactone (C4-HSL) enhanced the expression of MexAB-OprM, whereas N-(3-oxododecanoyl)-L-homoserine lactone had only a slight effect. Furthermore, this C4-HSL-mediated enhancement of mexAB-oprM expression was repressed by MexT, a positive regulator of the mexEF-oprN operon. It was concluded that ␤-lactam hypersusceptibility in nfxC-type mutant cells is caused by MexT-mediated cancellation of C4-HSL-mediated enhancement of MexABOprM expression.Pseudomonas aeruginosa is an opportunistic pathogen that causes infections in immunocompromised hosts and colonizes the lungs of individuals with cystic fibrosis. This organism shows broad resistance to structurally and functionally dissimilar antibiotics. This type of multidrug resistance is attributable mainly to the expression of the xenobiotic extrusion transporter MexAB-OprM coupled with tight outer membrane permeability (21,23,29). The mexAB-oprM operon encodes three protein subunits (21,26,29): the intrinsic inner membrane protein MexB (9), the inner membrane-associated periplasmic lipoprotein MexA (45), and the outer membrane lipoprotein OprM (13,22,44). The mexAB-oprM operon is negatively regulated by the product of the mexR gene, which is located upstream of the mexAB-oprM genes and is divergently transcribed (1,5,14,34,37). nalB-type mutants caused by the mexR mutation derepress MexAB-OprM production and are highly resistant to fluoroquinolones, chloramphenicol, and most classical ␤-lactam antibiotics (30,32,35,42).Recently, it was reported that the MexAB-OprM transporter exports quorum-sensing mediators, acylhomoserine lactones (AHSLs), which induce the production of cell density-dependent virulence factors, including proteases, rhamnolipids, exotoxin A, exoenzyme S, and pyocyanin (24, 25). The AHSLs control at least two quorum-sensing systems in P. aeruginosa, namely, LasR-LasI and RhlR-RhlI. LasI and RhlI catalyze the last steps in the syntheses of N-(3-oxododecanoyl)-L-homoserine lactone (3-oxo-C12-HSL) and N-butyryl-L-homoserine lactone (C4-HSL), respectively. LasR and RhlR are specifically activated by the diffusible signaling molecules 3-oxo-C12-HSL and C4-HSL, respectively.Four resistant-nodulation-division efflux pumps (MexABOprM, MexCD-OprJ, MexEF-OprN, ...

1 Zeolites were added to polyethersulphone (PES) membranes in order to enhance their capability of attracting creatinine. Zeolite was blended with PES by varying its doping concentration and time, and the mechanical properties of the resulting hollow fibre membranes produced under optimal concentration were characterised. The values of tensile stress, tensile strain and Young's modulus, as well as the flux and rejection of the membranes were determined.

Anaerobic growth of Pseudomonas aeruginosa PAO1 was affected by quorum sensing. Deletion of genes that produce N-acyl-L-homoserine lactone signals resulted in an increase in denitrification activity, which was repressed by exogenous signal molecules. The effect of the las quorum-sensing system was dependent on the rhl quorum-sensing system in regulating denitrification.Bacteria regulate their metabolism to adapt to various conditions by sensing environmental signals. Under anoxic conditions, many bacteria are able to use N-oxides as terminal electron acceptors. Pseudomonas aeruginosa is a denitrifying bacterium capable of anaerobic growth by utilizing N-oxides such as nitrate (NO 3 Ϫ ) and nitrite (NO 2 Ϫ ). Denitrification is induced under low-oxygen conditions when N-oxides are also present (1, 9, 11).Recent studies on bacteria have revealed new types of environmental signals known as cell-to-cell communication signals (25). P. aeruginosa is reported to control gene expression globally in response to cell density by utilizing N-acyl-L-homoserine lactone (AHL) signals. This cell-density-dependent regulation is termed quorum sensing (5). P. aeruginosa possesses at least two quorum-sensing systems: the LasR-LasI (las) and RhlR-RhlI (rhl) systems (20). LasI directs the synthesis of the AHL signal N-(3-oxododecanoyl)-L-homoserine lactone (3-oxo-C 12 -HSL) (17, 18), and RhlI directs the synthesis of another AHL signal, N-butyryl-L-homoserine lactone (C 4 -HSL) (19). The transcription regulatory proteins, LasR (6) and RhlR (16), are specifically activated by 3-oxo-C 12 -HSL and C 4 -HSL, respectively. Previous studies have indicated that production of virulence factors, such as protease, exotoxin A, rhamnolipids, and siderophores, is regulated by quorum sensing in P. aeruginosa (7,12,26), suggesting that quorum sensing is important in the pathogenesis of infection with this bacterium.Recent transcriptomic and proteomic studies indicate that quorum sensing is a global regulation system in P. aeruginosa (3,23,28). From these findings, it can be presumed that quorum-sensing systems have ecologically important roles in addition to the control of pathogenesis for the bacterium. For example, recent work suggests that quorum sensing regulates the activities of denitrification enzymes. In a recent study by Yoon et al. (31), the authors reported that levels of denitrifying enzyme activities of anaerobically grown P. aeruginosa cells are higher for an rhlR mutant than for its parent strain in an in vitro system. We were interested in further characterizing the potential anaerobic regulation of denitrification by the quorum-sensing system. Here we present a comprehensive analysis of the impact of quorum sensing on the denitrification pathway under anaerobic conditions by using in vivo and in vitro analyses.Effect of quorum sensing on denitrification activity. P. aeruginosa PAO1 was cultured anaerobically in 17-ml Hungate tubes containing 5 ml Luria-Bertani (LB) medium supplemented with 100 mM KNO 3 with shaking at 200 rpm at 37...

The opportunistic human bacterial pathogen Pseudomonas aeruginosa produces membrane vesicles (MVs) in its surrounding environment. Several features of the P. aeruginosa MV production mechanism are still unknown. We previously observed that depletion of Opr86, which has a role in outer membrane protein (OMP) assembly, resulted in hypervesiculation. In this study, we showed that the outer membrane machinery and alginate synthesis regulatory machinery are closely related to MV production in P. aeruginosa. Depletion of Opr86 resulted in increased expression of the periplasmic serine protease MucD, suggesting that the accumulation of misfolded OMPs in the periplasm is related to MV production. Indeed, the mucD mutant showed a mucoid phenotype and the mucD mutation caused increased MV production. Strains with the gene encoding alginate synthetic regulator AlgU, MucA, or MucB deleted also caused altered MV production. Overexpression of either MucD or AlgW serine proteases resulted in decreased MV production, suggesting that proteases localized in the periplasm repress MV production in P. aeruginosa. Deletion of mucD resulted in increased MV proteins, even in strains with mutations in the Pseudomonas quinolone signal (PQS), which serves as a positive regulator of MV production. This study suggests that misfolded OMPs may be important for MV production, in addition to PQS, and that these regulators act in independent pathways.