Project description:Pseudomonas aeruginosa is an opportunistic bacterial pathogen that possesses a plethora of virulence factors, including the public goods siderophores and exoproteases. Mutants unable to produce either exoprotease or siderophores are selected in competition with producers; hence, these two virulence factors are prone to exploitation by social cheaters. However, so far most of the studies of exoprotease exploitation by social cheaters had been done in the reference strains PAO1 and PA14 and the information available of these behaviors in clinical or environmental isolates is limited to a few studies. In this work, we studied the exoproteases secreted by two clinical isolates from cystic fibrosis patients belonging to the epidemic clone ST274-CC274. We found that the exoprotease exploitation in these strains is mild and not a result of the selection of lasR mutants during continuous growth in casein as sole carbon source, in contrast with the paradigm in the reference strains.
Project description:Pseudomonas aeruginosa is a versatile pathogen capable of causing illnesses that range from mild infections to life-threatening conditions. Its virulence is driven by a wide array of factors, among which extracellular vesicles (EVs) have gained recognition as important contributors to its pathogenicity. Despite this, the full scope of their roles remains unclear. A major barrier to EV characterization is the difficulty of vesicle isolation—procedures are often lengthy, yield is low, and specialized equipment is required. In this study, we assessed the effectiveness of a rapid vesicle extraction method from clinical strains of P. aeruginosa. To that end, we first selected and characterized six phenotypically diverse clinical strains of P. aeruginosa (two reference strains and 4 clinical isolates, including one strain from a cystic fibrosis patient) and used them to evaluate the vesicle extraction method. The results obtained through SDS-PAGE analysis, western blot, protein quantification, TEM, dynamic light scattering and mass spectrometry indicated the presence of vesicles in all samples; however, it was also possible to observe a large number of contaminants in some of them (mainly LS07 and Z37). Subsequent treatment with enzymes (DNase and/or alginate lyase) allowed for the elimination of the contaminants as observed by electron microscopy. Our results suggest that the method is suited for the vesicle extraction of clinical isolates of P. aeruginosa, where traditional methods, such as ultracentrifugation, are not available The phenotypic complexity of these strains presents challenges that current rapid purification methods are ill-equipped to handle, highlighting the need for improved or alternative approaches.
Project description:We report mRNA expression data from Pseudomonas fluorescens SBW25 wild type and two evolved strains (Beaumont et al., 2009). The evolution of one of these strains saw the emergence of colony switching; 1B4 switches rapidly between two different colony phenotypes. These two phenotypes were found to be genetically identical. Thus, in order to gain insight into epigenetic mechanisms of switching, we were interested in identifiying gene expression differences between ancestors and the 1B4 colony phenotypes.
Project description:35 P. aeruginosa clinical strains were cultivated under standard conditions, characterized in terms of virulence and biofilm phenotype, and their metabolomes were investigated by untargeted liquid chromatography-mass spectrometry.
Project description:To assess the role of two redox-sensitive transcriptional regulators, RoxSR and ANR, in Pseudomonas aeruginosa under aerobic conditions, microarray analysis was performed. Transcriptome profiles of roxSR mutant and anr mutant aerobically grown in LB medium were determined by Affymetrix GeneChip at both the exponential phase and early stationary phase and compared to that of the wild type strain. Experiment Overall Design: Pseudomonas aeruginosa wild type (PAO1ut), roxSR mutant (ROX1), and anr mutant (PAO6261) strains were cultivated aerobically in LB in Erlenmeyer flasks, and total RNAs were extracted at both the exponential phase (OD600 = 0.3) and early stationary phase (OD600 = 1.4). The experiment was performed in duplicate independent cultures.
Project description:The transcriptome of two different Pseudomonas aeruginosa mutant strains were compared to the Pseudomonas aeruginosa wild type strain in the stationary growth phase