ABSTRACT
This study evaluated, in vitro, the role of different Pseudomonas aeruginosa exopolysaccharides (EPS) in mediating adherence to human respiratory epithelial cells. Two mucoid and non-mucoid isogenic pairs of P aeruginosa strains isolated from patients with cystic fibrosis (CF) and bronchiectasis were used. Adherence was tested with human tracheal epithelial cell lines from CF and normal fetuses. The CF cells bound significantly more bacteria than the normal cells. The strain from the bronchiectasis patient was significantly more adherent than that from the CF patient and this difference was consistently most marked with the non-mucoid variant and with normal epithelial cells. The differing behaviour of mucoid CF and non-mucoid bronchiectasis strains reflected the chemical composition of their EPS: mainly alginate in the former and neutral polysaccharides in the latter. Additive inhibition experiments with chemically characterised EPS indicated that neutral polysaccharides associated with alginate may act as ligands for the adherence of P. aeruginosa to CF epithelial cells.
Subject(s)
Bacterial Adhesion , Polysaccharides, Bacterial/physiology , Pseudomonas aeruginosa/metabolism , Trachea/microbiology , Alginates/analysis , Alginates/chemistry , Bronchiectasis/microbiology , Carbohydrates/analysis , Cystic Fibrosis/microbiology , Humans , Polysaccharides, Bacterial/chemistry , Pseudomonas aeruginosa/chemistry , Sputum/microbiology , Time FactorsABSTRACT
There is evidence that exopolysaccharides (EPS) contribute to the persistence of Pseudomonas aeruginosa in cystic fibrosis lung. However, the relationship between the chemical composition of EPS and the modulation of phagocytic cells is poorly understood. In order to evaluate the role of the chemical composition of EPS in macrophage behavior changes, we pretreated macrophages with characterized EPS and assessed P. aeruginosa phagocytosis and reactive oxygen intermediate (ROI) production. The results showed that alginate and neutral polysaccharides are involved in phagocytic impairment of P. aeruginosa. Moreover, alginates were able to prime macrophages for increased P. aeruginosa-induced macrophage oxidative burst as determined by chemiluminescence. In contrast, neutral polysaccharides are responsible for the decrease of ROI by a scavenging effect evaluated by the xanthine-xanthine oxidase system. This study showed that the content of P. aeruginosa EPS in alginate, but also in neutral polysaccharides, influences the behavior of strains towards phagocytosis and macrophage oxidative burst.
Subject(s)
Alginates/pharmacology , Biocompatible Materials/pharmacology , Macrophages, Peritoneal/drug effects , Macrophages, Peritoneal/physiology , Polysaccharides, Bacterial/pharmacology , Pseudomonas Infections/immunology , Pseudomonas aeruginosa/immunology , Animals , Female , Glucuronic Acid , Hexuronic Acids , Luminescent Measurements , Mice , Phagocytosis/drug effects , Polysaccharides, Bacterial/isolation & purification , Pseudomonas aeruginosa/chemistry , Reactive Oxygen Species/metabolism , Respiratory Burst/drug effects , Xanthine Oxidase/metabolismABSTRACT
We used two PCR methods for epidemiological typing of Stenotrophomonas (Xanthomonas) maltophilia with either arbitrary primers (random amplified polymorphic DNA) or enterobacterial repetitive intergenic consensus sequences as primers (ERIC-PCR). The analysis was performed with 38 isolates of S. maltophilia, comprising 9 nosocomial isolates from a burn unit, 20 other clinical isolates epidemiologically unrelated, and 9 isolates from one cystic fibrosis patient. Both methods indicated that all of the nosocomial episodes were independent. In contrast, the nine isolates from the cystic fibrosis patient were assigned to very closely related profiles, especially by ERIC-PCR. We conclude that random amplified polymorphic DNA and ERIC-PCR have comparable reproducible and discriminatory powers for epidemiological typing of S. maltophilia, but ERIC-PCR profiles can be more easily evaluated.
Subject(s)
Polymerase Chain Reaction/methods , Xanthomonas/classification , Xanthomonas/genetics , Bacterial Typing Techniques/statistics & numerical data , Base Sequence , Burns/complications , Burns/microbiology , Cross Infection/epidemiology , Cross Infection/microbiology , Cystic Fibrosis/complications , Cystic Fibrosis/microbiology , DNA Fingerprinting/methods , DNA Fingerprinting/statistics & numerical data , DNA Primers/genetics , DNA, Bacterial/genetics , Epidemiologic Methods , France/epidemiology , Gram-Negative Bacterial Infections/complications , Gram-Negative Bacterial Infections/epidemiology , Gram-Negative Bacterial Infections/microbiology , Humans , Molecular Sequence Data , Opportunistic Infections/complications , Opportunistic Infections/epidemiology , Opportunistic Infections/microbiology , Polymerase Chain Reaction/statistics & numerical data , Reproducibility of Results , Xanthomonas/isolation & purificationABSTRACT
Two mucoid Pseudomonas aeruginosa strains and their non-mucoid revertants isolated from two different clinical origins (cystic fibrosis and bronchiectasis) were grown in various chemically defined media. The extracted exopolysaccharide was characterized by gas-liquid chromatography and 1H-NMR spectroscopy. The exopolysaccharide was always heterogeneous, with an alginate fraction and a neutral fraction essentially composed of glucose, galactose, rhamnose and hexosamines. The alginate composition (mannuronate/guluronate ratio and O-acetylation degree) changed according to the carbon source in nutrient media and whether the strains tested were responding differently to these environmental stimuli. In all cases, the best carbon source for the alginate production was glycerol: the two cystic fibrosis strains produced a predominantly O-acetylated alginate whereas only the mucoid bronchiectasis strain produced a polymannuronate exopolysaccharide.
Subject(s)
Polysaccharides, Bacterial/chemistry , Pseudomonas aeruginosa/chemistry , Alginates/chemistry , Bronchiectasis/microbiology , Culture Media , Cystic Fibrosis/microbiology , Glucuronic Acid , Hexuronic Acids , Humans , Mucus/chemistry , Pseudomonas aeruginosa/growth & development , Pseudomonas aeruginosa/isolation & purification , Species SpecificityABSTRACT
The aim of this study was to evaluate the in vitro effect of five antibiotics at sub-inhibitory concentrations on the adhesive and haemagglutinating properties of Pseudomonas aeruginosa isolated from cystic fibrosis sputa. Eleven isolates (mucoid and non-mucoid) from cystic fibrosis, and four isolates (mucoid and non-mucoid) from other chronic respiratory infections were tested. The adhesion test was performed on human lymphoblastoid cell-lines; the haemagglutination test used human O+ and guinea-pig erythrocytes. The antibiotics were tested at six sub-inhibitory concentrations, from MIC/2 to MIC/64. Among the five antibiotics, cefsulodin and pefloxacin were the most active in decreasing the adhesive properties: this effect was statistically significant at MIC/2 and MIC/4 for cefsulodin and at all sub-inhibitory concentrations for pefloxacin. No differences appeared between mucoid and non-mucoid strains, and no correlation was noted with their clinical origins. The three other antibiotics (ceftazidime, latamoxef and imipenem) had no significant effect on the adhesion of all the strains tested, but their effect was rather strain-dependent. This fact and the heterogeneity found in adherence and haemagglutinating activity of each strain suggest that the adhesins and the haemagglutinins of P. aeruginosa are very complex structures.