TraDIS-Xpress analysis of gene functions in Escherichia coli
Ontology highlight
ABSTRACT: This experiment was conducted to understand the gene functions of Escherichia coli in different concentrations of induction of Isopropyl β-D-1-thiogalactopyranoside (IPTG). Experiments were performed with no induction, or with induction using 0.2mM or 1 mM of IPTG.
Project description:In past, resistance mechanisms have been identified by analysis of resistant isolates or defined mutants. Recently, high-throughput transposon mutagenesis coupled with sequencing (TraDIS-Xpress) is another approach proving useful for elucidating the roles of genes involved in the overall cellular response to a particular stress. In this study, we used TraDIS-Xpress to determine the role played by genes following exposure to colistin stress. Approximately 10^7 cells from the mutant library were inoculated into LB broth at a range of doubling concentrations of colistin ( 0.25 x MIC, 0.5 x MIC, 1 x MIC, 2 X MIC). Experiments were performed with no induction, or with induction using 0.2 or 1 mM of Isopropyl β-D-1-thiogalactopyranoside (IPTG). All experiments were performed in duplicate.
Project description:Found how E. coli responds to the commonly prescribed antibiotic-inhibitor combination piperacillin and tazobactam, separately and in combination. TraDIS-Xpress was performed following the protocol outlined by Yasir et al 2020 Genome Research. The results suggest tazobactam triggers the activity of multidrug efflux systems that have been previously seen to confer clinical resistance to multiple classes of antibiotics.
Project description:The mechanisms of action of common food preservatives are poorly understood. As there is a drive to develop alternative preservatives, understanding the mechanisms of action of current preservatives can inform development of novel food preservatives to ensure their efficacy. Here we used TraDIS-Xpress, a large-scale, genome-wide unbiased screen to determine the mechanisms of action of common food preservatives by determining the genes that affect preservative susceptibility in Salmonella enterica serovar Typhimurium.
Project description:We used TraDIS-Xpress to determine the mechanism of action of a novel antimicrobial compound. We found that it inhibits lipid IVA biosynthesis in both Escherichia coli and Salmonella enterica serovar Typhimurium. We also were able to determine mechanisms of synergy with colistin, through ATP biosynthesis and the BasSR signalling system.
Project description:To assay every gene in the E. coli genome to identify those that contribute to increased or decreased susceptibility to the antibiotics trimethoprim and sulfamethoxazole. This will help to define more accurately those bacterial cell mechanisms that contribute to these phenomena and provide information that will contribute to the development of new antibiotics, or compounds or known antibiotics that synergise with those already in clinical use. Thus, this set of experiments confirmed that AZT, widely known for its antiviral activity, acts synergistically with trimehoprim.
Project description:This study investigates the mechanisms employed by Salmonella to colonise and establish itself on fresh produce at critical timepoints following infection. We established an alfalfa infection model and compared the findings to those obtained from glass surfaces. Our research revealed dynamic changes in the pathways associated with biofilm formation over time, with distinct plant-specific and glass-specific mechanisms for biofilm formation, alongside the identification of shared genes playing pivotal roles in both contexts.
Project description:The aim of the experiment was to assay every gene in the Escherichia coli genome to identify those that contribute to competition with Pseudomonas aeruginosa. A library of transposon-insertion mutants was grown overnight either alone or together with an equal starting number of P. aeruginosa cells. The experiment was also conducted in the presence or absence of glucose to interrogate the effect of a plentiful supply of carbon source. DNA sequencing was then used to reveal the locations of the transposons in every mutant. By comparing the numbers of each mutant between conditions, information can be gained about the relative fitness of that mutant under the conditions tested.
Project description:The aim of the experiment was to assay every gene in the E. coli genome to identify those that contribute to increased or decreased susceptibility to the beta-lactam antibiotic meropenem. A library of transposon-insertion mutants was grown overnight in the presence or absence of a range of concentrations of meropenem. Different concentrations of IPTG were also used to promote expression from a transposon-encoded promoter to assay the effects of increased transcription for each gene. DNA sequencing was then used to reveal the locations of the transposons in every mutant. By comparing the numbers of each mutant between conditions, information can be gained about the relative fitness of that mutant under the conditions tested.
Project description:Creation of Transposon libraries within 6 C. jejuni, 4 C. coli, 1 C. hyointestinalis and1 C. lari species. A mariner transposon was used to create libraries, these were sequenced and analysed using the Biotradis pipeline. This allowed us to investigate essential genes for each strain, across strains and between species.
Project description:The experiment aimed to find how Campylobacter responds to oxidative stress using hydrogen peroxide. This was done by using previously made TraDIS libraries (https://bmcmicrobiol.biomedcentral.com/articles/10.1186/s12866-023-02835-8) and putting them under oxidative stress. The strains used were C. jejuni 11168, C. coli 15-537360 and C. coli CCN182.