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In vitro evolution of Pseudomonas aeruginosa AA2 biofilms in the presence of cystic fibrosis lung microbiome members

ABSTRACT: In vitro evolution of Pseudomonas aeruginosa AA2 biofilms in the presence of cystic fibrosis lung microbiome members

PROVIDER: PRJEB29375 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	ERR2856770_1.fastq.gz	Fastqsanger.gz
	ERR2856770_2.fastq.gz	Fastqsanger.gz
	ERR2856780_1.fastq.gz	Fastqsanger.gz
	ERR2856780_2.fastq.gz	Fastqsanger.gz
	ERR2856771_1.fastq.gz	Fastqsanger.gz

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In vitro evolution of Pseudomonas aeruginosa AA2 biofilms in the presence of cystic fibrosis lung microbiome members

Project description:P. aeruginosa was cultured in a MultiScreen-Mesh plate, which has a filter at the bottom of the wells. The plate was immersed in either in medium alone (control) or in medium inoculated with a mixture of five bacterial strains commonly found in cystic fibrosis sputum (\"microbiome\"). The filter prevented physical contact between P. aeruginosa and the other bacteria, yet soluble products could migrate through the filter into the P. aeruginosa biofilm. P. aeruginosa was then allowed to form biofilms in the wells for 72h, then the biofilm was harvested and a fraction of the harvested cells were used for re-inoculations. This was repeated for 18 cycles for a total of 54 days.

2019-09-13 | E-MTAB-7331 | biostudies-arrayexpress

In vitro evolution of Pseudomonas aeruginosa AA2 biofilms in the presence of cystic fibrosis lung microbiome members.

Project description:In cystic fibrosis (CF) airways, the opportunistic pathogen Pseudomonas aeruginosa evolves from an acute to a chronic infection phenotype. Yet, the in vivo factors influencing the evolutionary trajectory of P. aeruginosa are poorly understood. This study aimed at understanding the role of the CF lung microbiome in P. aeruginosa evolution. Therefore, we investigated the in vitro biofilm evolution of an early CF P. aeruginosa isolate, AA2, in the presence or absence of a synthetic CF lung microbiome. Whole genome sequencing of evolved populations revealed mutations in quorum sensing (QS) genes (lasR, pqsR) with and without the microbiome. Phenotypic assays confirmed decreased production of the QS molecule 3-O-C12-homoserine lactone, and QS-regulated virulence factors pyocyanin and protease. Furthermore, a mixture of lasR and lasR pqsR mutants was found, in which double mutants showed less pyocyanin and protease production than lasR mutants. While the microbial community did not influence the production of the tested P. aeruginosa virulence factors, we observed a trend towards more mutations in the transcriptional regulators gntR and mexL when P. aeruginosa was grown alone. P. aeruginosa developed resistance to β-lactam antibiotics during evolution, when grown with and without the microbiome. In conclusion, in an experimental biofilm environment, the early P. aeruginosa CF isolate AA2 evolves towards a CF-like genotype and phenotype, and most studied evolutionary adaptations are not impacted by CF microbiome members.

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Expression data from Pseudomonas aeruginosa wild type and ∆anr grown as biofilms on ∆F508 cystic fibrosis bronchial epithelial cells (CFBEs)

Project description:The transcription factor Anr regulates the response to low oxygen in P. aeruginosa and is inhibited by oxygen. We used microarrys to compare gene expression in P. aeruginosa PAO1 wild-type with an isogenic anr mutant in order to determine which transcripts are affected by Anr. We grew P. aeruginosa cells as biofilms on CFBE cells in order to model cystic fibrosis airways infections.

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Transcriptome Analysis of Candida albicans Biofilms Grown in the Presence of Arachidonic Acid and Fluconazole

Project description:Candida albicans is a commensal yeast within the human microbiota with significant medical importance because of its pathogenic potential. The yeast produces biofilms, which are highly resistant to available antifungals. High level of antifungal resistance by C. albicans biofilms has resulted in the need for alternative treatment. Polyunsaturated fatty acids such as arachidonic acid has been reported to increase the susceptibility of C. albicans biofilms to azole. However, the underlining mechanism is unknown. To unravel the mechanism behind this phenomenon, identification of differentially regulated genes in C. albicans biofilms grown in the presence of arachidonic acid, fluconazole, and the combination of both compounds was conducted using RNAseq.

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Experimental evolution of Pseudomonas aeruginosa biofilms exposed to furanone C-30 with and without tobramycin

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.

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Project description:Pseudomonas aeruginosa was repeatedly and intermittently exposed to tobramycin. Bacteria were grown in synthetic cystic fibrosis medium in wells of a 96-well microtiter plate. After 24 hours, more medium with or without tobramycin was added. After another 24 hours of incubation, a subsample of the well content was used to inoculate fresh synthetic cystic fibrosis medium in a 96-well microtiter plate. This was repeated for a total of 15 cycles. Evolved lineages were then DNA-sequenced to screen for genome changes.

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The presence of Cystic Fibrosis Related Diabetes modifies the sputum microbiome in Cystic Fibrosis disease.

Project description:The presence of Cystic Fibrosis Related Diabetes modifies the sputum microbiome in Cystic Fibrosis disease.

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Cystic leukoencephalopathy without megalencephaly

Project description:The cystic leukoencephalopathy without megalencephaly has been defined as distinct autosomal-recessive syndrome of quasi-static encephalopathy with normo- or microcephaly, impaired psychomotor development and characteristic pattern on brain MRI. We identified mutations in the gene encoding the RNASET2 glycoprotein in seven affected individuals as the cause of disease. The results suggest that RNASET2 plays an important role in central nervous system myelination and development. 2-Color dye swap design independent for two different families including technical and biologial replicates for affected family members (disease) and not affected family members (controls).

2009-06-02 | E-GEOD-11964 | biostudies-arrayexpress

Identification of adaptive mechanisms leading to reduced antibiotic susceptibility in bacterial biofilms using experimental evolution. Part 1: Pseudomonas aeruginosa strain AA2 biofilms on beads

Project description:Pseudomonas aeruginosa AA2 was repeatedly and intermittently exposed to tobramycin, ciprofloxacin or meropenem. Bacteria were grown on cryobeads submerged in liquid BHI medium. After 24 hours, the beads were washed and fresh medium with of without antibiotics added. After another 24 hours of incubation, the beads were washed, the bacteria removed from the beads, and used for inoculation of fresh beads. This was repeated to a total of up to ten cycles. Evolved lineages were then DNA-sequenced to screen for genome changes.

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Identification of adaptive mechanisms leading to reduced antibiotic susceptibility in bacterial biofilms using experimental evolution. Part 1: Pseudomonas aeruginosa strain AA2 biofilms on beads

Project description:Identification of adaptive mechanisms leading to reduced antibiotic susceptibility in bacterial biofilms using experimental evolution. Part 1: Pseudomonas aeruginosa strain AA2 biofilms on beads

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