Unknown,Transcriptomics,Genomics,Proteomics

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Plasticity of transcriptional regulation under antibiotic stress

ABSTRACT: Evolution of antibiotic resistance in microbes is frequently achieved by acquisition of spontaneous mutations during antimicrobial therapy. Here we demonstrate that inactivation of a central regulator of iron homeostasis (fur) facilitates laboratory evolution of ciprofloxacin resistance in Escherichia coli. To decipher the underlying molecular mechanisms, we first performed a global transcriptome analysis and demonstrated a substantial reorganization of the Fur regulon in response to antibiotic treatment. We hypothesized that the impact of Fur on evolvability under antibiotic pressure is due to the elevated intracellular concentration of free iron and the consequent enhancement of oxidative damage-induced mutagenesis. In agreement with expectations, over-expression of iron storage proteins, inhibition of iron transport, or anaerobic conditions drastically suppressed the evolution of resistance, while inhibition of the SOS response-mediated mutagenesis had no such effect in fur deficient population. In sum, our work revealed the central role of iron metabolism in de novo evolution of antibiotic resistance, a pattern that could influence the development of novel antimicrobial strategies. We used microarrays to identify genotype specific transcriptional changes under severe DNA damaging conditions (antibiotic ciprofloxacin). We treated Escherichia coli cells with a highly toxic level of ciprofloxacin (gyrase inhibitor) for RNA extraction and hybridization on Affymetrix microarrays. We planned to find genotype specific transcriptional responses using WT control (BW25113) and fur-knockout mutant (selected from the KEIO collection) strains during antibiotic treatments. For each treatment type we used two biological replicates.

ORGANISM(S): Escherichia coli K-12

SUBMITTER: Balazs Bogos

PROVIDER: E-GEOD-55662 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Perturbation of iron homeostasis promotes the evolution of antibiotic resistance.

Méhi Orsolya O Bogos Balázs B Csörgő Bálint B Pál Ferenc F Nyerges Akos A Papp Balázs B Pál Csaba C

Molecular biology and evolution 20140724 10

Evolution of antibiotic resistance in microbes is frequently achieved by acquisition of spontaneous mutations during antimicrobial therapy. Here, we demonstrate that inactivation of a central transcriptional regulator of iron homeostasis (Fur) facilitates laboratory evolution of ciprofloxacin resistance in Escherichia coli. To decipher the underlying molecular mechanisms, we first performed a global transcriptome analysis and demonstrated that the set of genes regulated by Fur changes substantia ...[more]

PMID: 25063442

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Project description:Treatment failures of antibiotic therapy are of major concern and can be caused by a misalignment of the antibiotic susceptibility determined in vitro with the behaviour of the pathogen in the patient. The aim of this study was to investigate the transcriptomic response of the uropathogenic strain E. coli CFT073 to antibiotic treatment in blood stream infection (BSI) models in order to understand and avoid antibiotic therapy failures in urosepsis treatments. Blood stream infection models were established by growing E. coli CFT073 in pooled human serum with and without ciprofloxacin and compared to Iso-sensitest medium. The antibiotic challenge was introduced at mid-logarithmic phase of growth of the organism to depict a clinical scenario . Global gene expression profiling of these conditions was examined using commercial DNA microarrays. The organismâs metabolic genes appeared to be regulated differently in each medium, this indicated that the bacterial growth regulation were different between the models. Bacterial growth in human serum mainly involved regulations of amino acid synthesis/utilisation such as glycine, arginine, thiamine, regulations of fimbrial proteins and bacteriophage genes. When comparing the responses to antibiotic challenge, bacteria grown in the respective medium displayed specific responses to the antibiotic challenge which were not seen in the other media. The common functions of genes that responded to the ciprofloxacin challenge were SOS response, DNA repair, DNA replication, fimbrial genes and bacteriophage initiation. A subset of the bacteriophage genes showed similar responses between the three models. From genes that were differentially regulated, responses observed in the serum model appeared to have the highest fold changes. In this study we established new models to investigate blood stream infections. They have been used to identify previously unknown differences in the molecular response to antibiotic treatment by the uropathogenic E. coli CFT073 depending on the media. These unique responses will help to unravel the complexity of bloodstream infection and can help to improve the antibiotic therapy that is used. A 20 array study using total RNA recovered from bacteria from bacteria that were either grown in human serum or Iso-Sensitest (IST) broth, with and without ciprofloxacin challenge. Arrays were performed in 5 biological replicates from each condition.Upon QC checks, certain biological repeats were excluded due to poor hybridisation results.

Project description:Treatment failures of antibiotic therapy are of major concern and can be caused by a misalignment of the antibiotic susceptibility determined in vitro with the behaviour of the pathogen in the patient. The aim of this study was to investigate the transcriptomic response of the uropathogenic strain E. coli CFT073 to antibiotic treatment in blood stream infection (BSI) models in order to understand and avoid antibiotic therapy failures in urosepsis treatments. Blood stream infection models were established by growing E. coli CFT073 in pooled human blood with and without ciprofloxacin. The antibiotic challenge was introduced at mid-logarithmic phase of growth of the organism to depict a clinical scenario. The responses were quantified by comparing to the responses at a given time point without the challenge. Global gene expression profiling of these conditions was examined using commercial DNA microarrays. The organismâ??s metabolic genes appeared to be regulated differently in each medium, this indicated that the bacterial growth regulation were different between the models. Bacterial growth in human serum mainly involved regulations of amino acid synthesis/utilisation such as glycine, arginine, thiamine, regulations of fimbrial proteins and bacteriophage genes. When comparing the responses to antibiotic challenge, bacteria grown in the respective medium displayed specific responses to the antibiotic challenge which were not seen in the other media. The common functions of genes that responded to the ciprofloxacin challenge were SOS response, DNA repair, DNA replication, fimbrial genes and bacteriophage initiation. A subset of the bacteriophage genes showed similar responses between the three models. From genes that were differentially regulated, responses observed in the serum model appeared to have the highest fold changes. In this study we established new models to investigate blood stream infections. They have been used to identify previously unknown differences in the molecular response to antibiotic treatment by the uropathogenic E. coli CFT073 depending on the media. These unique responses will help to unravel the complexity of bloodstream infection and can help to improve the antibiotic therapy that is used. A 10 array study using total RNA recovered from bacteria that were grown in human whole blood, with and without ciprofloxacin challenge. Arrays were performed in 5 biological replicates from each condition.

Dataset Information

Plasticity of transcriptional regulation under antibiotic stress

Publications

Perturbation of iron homeostasis promotes the evolution of antibiotic resistance.

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