Project description:Pseudomonas aeruginosa infections can be virtually impossible to eradicate and the evolution of resistance during antibiotic therapy is a significant concern. In this study, we use DNA microarrays to characterize the global transcriptional response of P. aeruginosa to clinical-like doses of the antibiotic ciprofloxacin and also to determine the component that is regulated by LexA cleavage and the SOS response. We find that genes involved in virtually every facet of metabolism are down-regulated in response to ciprofloxacin. The LexA-controlled SOS regulon identified by microarray analysis includes only fifteen genes, but does include several genes that encode proteins involved in recombination and replication, including two inducible polymerases known to play a role in mutation and the evolution of antibiotic resistance in other organisms. The data suggests that the inhibition of LexA cleavage during therapy might help combat this pathogen by decreasing its ability to adapt and evolve resistance. Keywords: Time course of response of P. aeruginosa to the antibiotic ciprofloxacin
Project description:Pseudomonas aeruginosa infections can be virtually impossible to eradicate and the evolution of resistance during antibiotic therapy is a significant concern. In this study, we use DNA microarrays to characterize the global transcriptional response of P. aeruginosa to clinical-like doses of the antibiotic ciprofloxacin and also to determine the component that is regulated by LexA cleavage and the SOS response. We find that genes involved in virtually every facet of metabolism are down-regulated in response to ciprofloxacin. The LexA-controlled SOS regulon identified by microarray analysis includes only fifteen genes, but does include several genes that encode proteins involved in recombination and replication, including two inducible polymerases known to play a role in mutation and the evolution of antibiotic resistance in other organisms. The data suggests that the inhibition of LexA cleavage during therapy might help combat this pathogen by decreasing its ability to adapt and evolve resistance. Experiment Overall Design: Sample preparation and data analysis. For each strain (PAO1 and the lexA uninducible mutant), 5 clones were inoculated in LB and grown 18 h. Cultures were diluted 1:500 and grown to mid-log phase (OD600 ~0.4-0.5) at which point ciprofloxacin was added to a final concentration of 1 μg/ml. At 0, 30 and 120 minutes following ciprofloxacin addition, appropriate volumes from each of the 5 cultures per strain were pooled and added to 2 volumes of RNAprotect reagent (Qiagen); cell pellets were stored at 4 ºC until RNA extraction. Total RNA was extracted using the RNeasy Mini kit (Qiagen) at the end of the sample collection period. This procedure was repeated three independent times to generate three samples each just prior to and 120 minutes post ciprofloxacin addition.
Project description:We report the transcriptomic analysis of the response of P. aeruginosa AG1 to Ciprofloxacin to identify molecular determinants related to its tolerance to the antibiotic and the enriched pathways along time
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.
Project description:Gene expression of P. aerruginosa changes after short-term exposure to ciprofloxacin at sub-inhibitory concentrations but the effect of long-term exposure which select for the most fitted subpopulations is not known. We used microarrays to investigate the changes in gene expression of P.aeruginosa PAO1 and mutator (Î? mutS) after long-term evolution (94 daily passages) in LB in the presence and absence of ciprofloxacin Three different colonies from the ancestral populations of PAO1 and mutator (Î? mutS) as well as from the evolved populations (day 94) of each of the three lineages (A;B;C) in the presence or absence of ciprofloxacin at a concentration of 0.05 µg/ml were used for overnight cultures in LB and total RNA was extracted at OD600nm=1 and hybridized on P. aeruginosa Affymetrix chip.
Project description:Using in vitro directed evolution, we show that mismatch repair-deficient Pseudomonas aeruginosa can engage novel resistance mechanisms to ceftazidime-avibactam.
Project description:Previous synthesized Pt NPs were selected to evaluate the influences on bacterial resistance, and a typical pathogenic microbe P. aeruginosa was chosen as model bacteria. After 60-day PtNPs exposure, we found under 12.5 μg/mL of platinum nanoparticles (PtNPs) exposure for ~7200 generations, the IC50 of evolved Pseudomonas aeruginosa PAO1 to imipenem (IPM) and ciprofloxacin (CIP) reduced 77.0% and 87.8%, respectively. Interestingly, long-term of PtNPs exposure arose the bacterial susceptibility on antibiotics. We then performed gene expression profiling analysis using data obtained from RNA-seq.
Project description:Pseudomonas aeruginosa PAO1 persister and normal cells were treated with and without Granulocyte Macrophage Colony-Stimulating Factor (GM-CSF) to understand the effect of GM-CSF on gene expression of PAO1. We used DNA microarrays to identify the down-regulated and up-regulated genes after GM-CSF treatment. Pseudonmonas aeruginosa PAO1 normal cells were obtained from stationary phase culture and persister cells were isolated by lysing normal cells with ciprofloxacin. The normal cells and isolated persister cells were treated with GM-CSF and selected for RNA extraction and hybridization on Affymetrix microarrays. The experiment was performed in two biological replicates. The samples for the biological replciates were: Control C1, Treatment T1, Control C2, and Treatment T2.