Biology subjects

O'Boyle, N.

Publications and source records attributed to O'Boyle, N..

3 recordsLinked to original sources

bioRxiv2023

Enhancing a multipurpose artificial urine for culture and gene expression studies of uropathogenic Escherichia coli strains

Uropathogenic Escherichia coli (UPEC) are the most common cause of urinary tract infections, which pose a great burden on global health and the economy through morbidity, mortality, healthcare costs and loss of productivity. Pooled human urine can be used as a growth medium for in vitro studies, however even if the same donors are used, composition can vary depending largely on diet and fluid intake. There have been a number of artificial urine formulae used as alternatives to pooled human urine. However, we observed that a recently reported multipurpose artificial urine was unable to support the growth of prototypic UPEC strains suggesting it lacked key metabolites. We therefore used liquid chromatography mass spectrometry to identify and adjust the metabolic profile of multipurpose artificial urine closer to that of pooled human urine. Modification in this way facilitated growth of UPEC strains with growth rates similar to those obtained in pooled human urine. Transcriptomic analysis of UPEC strains cultured in enhanced artificial urine and pooled human urine showed that the gene expression profiles are similar, with less than 7% of genes differentially expressed between the two conditions. The data support this enhanced artificial urine as a robust media to study aspects of UPEC physiology in vitro.

microbiology↗

bioRxiv2022

The therapeutic potential of D-Serine in reducing expression of the cytopathic genotoxin colibactin

Some Escherichia coli strains belonging mainly to the B2 phylogroup harbour the pks island, a 54 kb genomic island encoding the biosynthesis genes for a genotoxic compound named colibactin. In eukaryotic cells, colibactin can induce DNA damage, cell cycle arrest and chromosomal instability. Moreover, production of colibactin has been implicated in the development of colorectal cancer. In this study, we demonstrate the inhibitory effect of D-Serine on the expression of the pks island in two colibactin-producing strains, CFT073 and Nissle 1917, and determine the implications for cytopathic effects on host cells. To investigate the specificity of the inhibitory effect of D-Serine, we also tested a comprehensive panel of proteinogenic L-amino acids and corresponding D-enantiomers for their ability to modulate clbB transcription using RT-qPCR. Several D-amino acids exhibited the ability to inhibit expression of clbB, with D-Serine exerting the strongest repressing activity (3.81-fold in CFT073; 3.80-fold in Nissle 1917) and thus, we focussed additional experiments on D-Serine. To investigate the cellular effect, we investigated if repression of colibactin by D-Serine could reduce the cytopathic responses normally observed during infection of HeLa cells with pks+ strains. Levels of {gamma}-H2AX (a marker of DNA double strand breaks) were reduced 2.75-fold in cells infected with D-Serine treatment. Moreover, exposure of pks+ E. coli to D-Serine during infection caused a reduction in cellular senescence that was observable at 72 h post infection. The recent finding of an association between pks-carrying commensal E. coli and CRC, highlights the necessity for the development of colibactin targeting therapeutics. Here we show that D-Serine can reduce expression of colibactin, and inhibit downstream cellular cytopathy, illuminating its therapeutic potential to prevent colibactin-associated disease.

microbiology↗

bioRxiv2021

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. ImportanceThe pathogenic Escherichia coli comprise a group of highly specialized bacteria, some of which are capable of disseminating from the intestine and causing disease at other sites within the human host. Chemicals (metabolites) derived from the host and other microorganisms shape the behaviour of E. coli in different environments. Here we investigate the changes in gene expression that occur in E. coli strains capable (UPEC and NMEC) and incapable (EHEC) of metabolizing D-ser - a metabolite specifically enriched in the urine and in regions of the brain. We show that EHEC, UPEC and NMEC - distinct pathotypes associated with disease in the gut, bladder and brain, respectively, respond in a distinct manner to D-ser. Many of the genes affected by D-ser have been shown to be important during disease, highlighting the importance of varied responses to this common signal in host infection.

molecular biology↗