Project description:Our aim was to identify candidate transcripts that distinguish AIEC from non-invasive E. coli (NIEC) strains and might be useful for rapid and accurate identification of AIEC by culture-independent technology. We performed comparative RNA-Sequence (RNASeq) analysis using AIEC strain LF82 and NIEC strain HS during exponential and stationary growth.
Project description:DNA microarrays were conducted on E. coli K12 cells stressed with 10 μM in N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine (TPEN). Overall, 260 genes varied in expression, 114 up-regulated and 146 down-regulated by Zn deprivation Keywords: TPEN stress
Project description:Strains of urinary tract associated E. coli both recent isolates and from the ECOR collection and non pathogenic E. coli strains were analyzed. Replicates were performed to establish the reproduciblity, then single experiments were performed there on.
Project description:The complex reservoir of metabolite-producing bacteria in the gastrointestinal tract contributes tremendously to human health and disease. Bacterial composition, and by extension gut metabolomic composition, is undoubtably influenced by the use of modern antibiotics. Herein, we demonstrate that polymyxin B, a last resort antibiotic used for chronic multidrug resistant infections infections, influences the production of the genotoxic metabolite colibactin from adherent-invasive Escherichia coli (AIEC) NC101. Colibactin can augment colorectal cancer (CRC) through DNA double stranded breaks and interstrand crosslinks. While the structure and biosynthesis of colibactin has been elucidated, chemical-induced regulation of its biosynthetic gene cluster and subsequent production of the genotoxin by pathogenic E. coli are largely unexplored. This research highlights the regulation of the colibactin-producing biosynthetic gene cluster under polymyxin stress. Using a multi-omic approach, we have identified that polymyxin stress enhances the abundance of colibactin biosynthesis proteins (Clb’s) in multiple pks+ E. coli strains, including pro-carcinogenic AIEC: NC101, the probiotic strain: E. coli Nissle 1917, and the antibiotic testing strain: E. coli ATCC 25922. Expression analysis via qPCR revealed that increased transcription of clb genes likely contributes to elevated Clb protein levels in NC101. Enhanced production of Clb’s by NC101 under polymyxin stress matched an increased production of the colibactin prodrug motif, a proxy for the mature genotoxic metabolite. Furthermore, E. coli with heightened tolerance for polymyxin antibiotics induced greater DNA damage, assessed by quantification of γH2AX staining in cultured intestinal epithelial cells. This study establishes a key link between the polymyxin B stress response and colibactin production in pks+ E. coli. Ultimately, our findings will inform future studies investigating colibactin regulation, the microbial response to antibiotics in the gut, and the ability of seemingly innocuous commensal microbes to induce host disease.
Project description:These E. coli strains were grown with various signaling molecules and the expression profiles were determined. Keywords: addition of quorum and host hormone signals
Project description:Expression and use of motility genes is a potentially beneficial but costly process in bacteria. Interestingly, many isolate strains of Escherichia coli possess motility genes but have lost the ability to activate them in conditions in which motile cells are advantageous, raising the question of how they respond to these situations. Through transcriptome profiling of strains in the E. coli single-gene knockout Keio collection, we noticed drastic upregulation of motility genes in many of the deletion strains as compared to its typically non-motile parent strain (BW25113). This switch to a motile phenotype is not a direct consequence of the genes deleted, but is instead due to a variety of secondary mutations that increase the synthesis of the major motility regulator, FlhDC. We found that a phenotypic switch to motility at a population level can be induced in non-motile E. coli strains by incubation in non-shaking liquid medium overnight but not in shaking media. Individual isolates after the overnight incubation acquired distinct mutations upstream of the flhDC operon, including different insertion sequence (IS) elements and, to a lesser extent, point mutations. The rapid sweep in the non-shaking population shows that non-motile strains without existing regulatory mechanisms can quickly adapt to a motile lifestyle by quickly acquired genetic changes.
Project description:DNA microarrays were conducted on E. coli K12 cells stressed with 10 μM in N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine (TPEN). Overall, 260 genes varied in expression, 114 up-regulated and 146 down-regulated by Zn deprivation Keywords: TPEN stress Array hybridizations were carried out for three RNA samples prepared from three independent cultures (control or TPEN-treated).
Project description:PhoP is considered a regulator of virulence despite being conserved in both pathogenic and non-pathogenic Enterobacteriaceae. While Escherichia coli strains represent both non-pathogenic commensal isolates and numerous virulent pathotypes, the PhoP virulence regulator has only been studied in commensal E. coli. To better understand how conserved transcription factors contribute to virulence, we characterized PhoP in pathogenic E. coli. Loss of phoP significantly attenuated E. coli during extraintestinal infection. This was not surprising since we demonstrated that PhoP differentially regulated the transcription of >600 genes. In addition to survival at acidic pH and resistance to polymyxin B, PhoP was required for repression of motility and oxygen-independent changes in the expression of primary dehydrogenase and terminal reductase respiratory chain components. All phenotypes have in common a reliance on an energized membrane. Thus, we hypothesized that PhoP mediated these effects by regulating genes that generate a proton motive force. Indeed, bacteria lacking PhoP exhibited a hyper-polarized membrane, and dissipation of the transmembrane electrochemical gradient increased the susceptibility of the phoP mutant to acidic pH, while inhibiting respiratory generation of the proton gradient restored resistance to antimicrobial peptides independent of lipopolysaccharide modification. These findings demonstrate a connection between PhoP, virulence, and the energized state of the membrane.
Project description:Imbalance in beneficial and harmful bacteria underlies gastrointestinal diseases, such as inflammatory bowel disease. Here, we demonstrated that certain E. coli strains, specifically adherent-invasive E. coli (AIEC), utilize a serine metabolism pathway to outcompete other E. coli strains in the inflamed gut. In contrast, amino acid metabolism has a minimal effect on their competitive fitness in the healthy gut. The availability of luminal serine used for the competition of E. coli is largely dependent on dietary intake, as the inflammation-induced blooms of AIEC are significantly blunted when amino acids, particularly serine, are removed from the diet. Thus, intestinal inflammation regulates the intraspecific competition between Enterobacteriaceae by eliciting their metabolic reprogramming.
Project description:Transcription profiles in BL21, BL21/pOri1 and BL21/pOri2 were analysed using DNA microarray technology. BL21, BL21/pOri1 or BL21/pOri2 strains were cultured at chemostat status and harvested after the cultivation arrived steady status. Keywords: Effects of plasmid DNA on Escherichia coli metabolism