Enhancing a multi-purpose artificial urine for culture and gene expression studies of uropathogenic Escherichia coli strains

Aims: The main objective of this study was to modify a recently reported multi-purpose artificial urine (MP-AU) for culture and gene expression studies of uropathogenic Escherichia coli (UPEC) strains. Methods and results: We used liquid chromatography mass spectrometry (LC-MS) to identify and adjust the metabolic profile of MP-AU closer to that of pooled human urine (PHU). Modification in this way facilitated growth of UPEC strains with growth rates similar to those obtained in PHU. Transcriptomic analysis of UPEC strains cultured in enhanced artificial urine (enhanced AU) and PHU showed that the gene expression profiles are similar, with less than 7% of genes differentially expressed between the two conditions. Conclusions: Enhancing an MP-AU with metabolites identified in PHU allows the enhanced AU to be used as a substitute for the culture and in vitro gene expression studies of UPEC strains

Control of resistance against bacteriophage killing by a metabolic regulator in meningitis-associated Escherichia coli

Ecologically beneficial traits in bacteria are encoded by intrinsic and horizontally acquired genes. However, such traits are not universal, and the highly mosaic nature of bacterial genomes requires control at the transcriptional level to drive these processes. It has emerged that regulatory flexibility is widespread in the Escherichia coli species, whereby preexisting transcription factors can acquire new and unrelated roles in regulating beneficial traits. DsdC is the regulator of D-serine tolerance in E. coli, is essential for D-serine catabolism, and is often encoded by two copies in neonatal meningitis-associated E. coli (NMEC). Here, we reveal that DsdC is a global regulator of transcription in NMEC and does not require D-serine for the control of novel beneficial traits. We show that DsdC binds the chromosome in an unusual manner, with many binding sites arranged in clusters spanning entire operons and within gene coding sequences, such as neuO. Importantly, we identify neuO as the most significantly down-regulated gene in a strain deleted for both dsdC copies, in both the presence and absence of D-serine. NeuO is prophage encoded in several NMEC K1 isolates and mediates capsule O-acetylation but has no effect on attachment to or invasion of human brain endothelial cells. Instead, we demonstrate that NeuO provides resistance against K1 bacteriophage attack and that this critical function is regulated by DsdC. This work highlights how a horizontally acquired enzyme that functions in cell-surface modulation can be controlled by an intrinsic regulator to provide a key ecological benefit to an E. coli pathotype

D-serine induces distinct transcriptomes in diverse Escherichia coli pathotypes

Appropriate interpretation of environmental signals facilitates niche specificity in pathogenic bacteria. However, the responses of niche-specific pathogens to common host signals are poorly understood. D-serine (D-ser) is a toxic metabolite present in highly variable concentrations at different colonisation sites within the human host that we previously found is capable of inducing changes in gene expression. In this study, we made the striking observation that the global transcriptional response of three Escherichia coli pathotypes - enterohaemorrhagic E. coli (EHEC), uropathogenic E. coli (UPEC) and neonatal meningitis associated E. coli (NMEC) - to D-ser was highly distinct. In fact, we identified no single differentially expressed gene common to all three strains. We observed the induction of ribosome-associated genes in extraintestinal pathogens UPEC and NMEC only, and the induction of purine metabolism genes in gut-restricted EHEC and UPEC indicating distinct transcriptional responses to a common signal. UPEC and NMEC encode dsdCXA – a genetic locus required for the detoxification and hence normal growth in the presence of D-ser. Specific transcriptional responses were induced in strains accumulating D-ser (WT EHEC and UPEC/NMEC mutants lacking the D-ser-responsive transcriptional activator DsdC), corroborating the notion that D-ser is an unfavourable metabolite if not metabolized. Importantly, many of the UPEC-associated transcriptome alterations correlate with published data on the urinary transcriptome, supporting the hypothesis that D-ser sensing forms a key part of urinary niche adaptation in this pathotype. Collectively, our results demonstrate distinct pleiotropic responses to a common metabolite in diverse E. coli pathotypes, with important implications for niche selectivity