Transcription profiling of Pseudomonas aeruginosa PAO1 virulent genes in the poplar rhizosphere
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ABSTRACT: Pseudomonas aeruginosa PAO1 contacted with and without poplar roots gene expression Poplar contacted with and without PAO1 gene expression. All samples cultured in 1 x hrp + 0.25 % sucrose Experiment Overall Design: Strains: P. aeruginosa PAO1 WT Experiment Overall Design: Medium: 1 x hrp + 0.25 % sucrose Experiment Overall Design: Biofilm grown on poplar root compared to biofilm grown on glasswool Experiment Overall Design: Poplar roots grown
Project description:Pseudomonas aeruginosa PAO1 contacted with and without poplar roots gene expression Poplar contacted with and without PAO1 gene expression. All samples cultured in 1 x hrp + 0.25 % sucrose Keywords: Contact with different species
Project description:P. aeruginosa PAO1 wild type and PA2663 mutant strains expression in biofilm cells relative to P. aeruginosa PAO1 wild type strain expression in biofilm cells. All samples cultured in LB with glass wool Keywords: Biofilm
Project description:P. aeruginosa PAO1 PA2663-UW expression in biofilm cells relative to P. aeruginosa PAO1 WT-UW expression in biofilm cells. All samples cultured in LB with glass wool. Keywords: Mutation
Project description:Biofilms are the most common cause of bacterial infections in humans and notoriously hard to treat due to their ability to withstand antibiotics and host immune defenses. To overcome the current lack of effective antibiofilm therapies and guide future design, the identification of novel gene targets is crucial. In this regard, transcriptional regulators have been proposed as promising targets for antimicrobial drug design, since they simultaneously affect multiple genes, typically lack human orthologs, and can be inactivated by small molecules that prevent dimerization. Therefore, a Transposon insertion sequencing approach was employed to systematically identify regulatory genes phenotypically affecting biofilm growth in Pseudomonas aeruginosa PA14. A screen of a pool of 300,000 transposon insertion mutants identified 349 genes involved in biofilm growth on hydroxy apatite, including 47 regulators. Detection of 19 regulatory genes participating in well-established biofilm pathways validated the results. An additional 28 novel prospective biofilm regulators suggested the requirement of multiple one-component transcriptional regulators. Defect phenotypes were confirmed for five one-component transcriptional regulators PA14_43720, PA14_56430, PA14_36180, arsR and merD as well as the protein kinase yeaG, which have not been implicated in biofilm growth before. Promisingly, the transcriptional regulator PA14_43720 displayed a conserved role in biofilm growth since its ortholog in P. aeruginosa strain PAO1 was also required for biofilm growth. Overall, our results highlighted that the gene network driving biofilm growth is complex and involves regulators beyond the primarily studied groups of two-component systems and cyclic diguanylate signaling proteins.
Project description:The PANarray design (GPL13324) contains the genes of eight P. aeruginosa genomes in non-redundant format, thus allowing identification of expression of non-PAO1 and other P. aeruginosa genes.
Project description:The aim of this study is to evaluate the evolutionary robustness of the quorum sensing inhibitor (QSI) furanone C-30 for the treatment of P. aeruginosa biofilm infections. We repeatedly exposed P. aeruginosa biofilms to furanone C-30 (with or without tobramycin) in the synthetic cystic fibrosis sputum medium (SCFM2) and characterized the genotype and phenotype of the evolved lineages. P. aeruginosa biofilms were grown in SCFM2 for 24 h after which the treatment in fresh SCFM2 was added to obtain a final concentration of 20 µg/ml tobramycin and 100 µg/ml furanone C-30. The negative control was treated with fresh SCFM2, including the same amount of DMSO (0.25%) as for the biofilms treated with C-30. After 24 h of static incubation at 37°C, biofilms were sonicated and vortexed in order to disintegrate the biofilm aggregates. After each cycle the number of CFU was determined and an aliquot of the culture was stored at -80°C in Microbank vials to allow further tests on the evolved strains. A sample from the treated biofilm was used to prepare a new overnight culture, in order to start a new cycle. For each treatment three independent lineages were established, that were each exposed for 16 cycles. Whole-genome sequencing was performed on the wild type P. aeruginosa PAO1 and on the exposed lineages after 5, 10 and 16 cycles.
Project description:We used DNA microarrays to investigate the impact of indole and 7-hdroxyindole on P. aeruginosa PAO1.For the microarray experiments, 10 g glass wool (Corning Glass Works, Corning, N.Y.) were used to form biofilms in 250 mL in 1 L Erlenmeyer shake flasks which were inoculated with overnight cultures of P. aeruginosa PAO1 diluted that were 1:100. For P. aeruginosa with 7-hydroxyindole and indole, 500 uM 7-hydroxyindole in 250 uL DMF, 1000 uM indole in 250 uL DMF, or 250 uL DMF alone were added to cells grown in LB. The cells were shaken at 250 rpm and at 37oC for 7 h to form biofilms on the glass wool, and RNA was isolated from the biofilm cells. Keywords: comparison with E. coli signal indole on P. aeruginosa PAO1