Application of TraDIS to define the core essential genome of Campylobacter jejuni and Campylobacter coli
Ontology highlight
ABSTRACT: 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.
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:Expression data for Desulfovibrio alaskensis strain G20 and mutants in regulator proteins grown on lactate sulfate media. Pellets collected at mid-log growth phase. We aimed to confirm or expand the predicted regulons of each of the transposon interrupted regulator mutants. 12 samples were collected: 10 regulator mutants, 1 mutant in a secretory protein and 1 control sample Desulfovibrio alaskensis G20. 1 replicate for each condition.
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: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:Expression data for Desulfovibrio alaskensis strain G20 and mutants in regulator proteins grown on lactate sulfate media. Pellets collected at mid-log growth phase. We aimed to confirm or expand the predicted regulons of each of the transposon interrupted regulator mutants. 8 samples were collected: 7 regulator mutants and 1 control sample Desulfovibrio alaskensis G20. 1 replicate for each condition.
Project description:Expression of efflux pumps is a key feature of most cells which are resistant to multiple antibiotics. This study used TraDIS-Xpress, a genome wide transposon mutagenesis technology to identify genes in Escherichia coli and Salmonella Typhimurium involved in drug efflux and its regulation. We exposed mutant libraries to the canonical efflux substrate acriflavine in the presence and absence of the efflux inhibitor phenylalanine-arginine β-naphthylamide. Comparisons between conditions identified efflux specific and drug specific responses. Known efflux associated genes were easily identified including: AcrAB-TolC, MarA, RamA and SoxS confirming specificity of the response. Further genes encoding cell envelope maintenance enzymes and products involved with stringent response activation, DNA housekeeping, respiration and glutathione biosynthesis were also identified as affecting efflux activity in both species. We identified a conserved set of pathways crucial for efflux activity in these experimental conditions which expands the list of genes known to impact efflux efficacy.
Project description:Most bacteria can form biofilms, which typically have a life cycle from cells initially attaching to a surface before aggregation and growth produces biomass and an extracellular matrix before finally cells disperse. To maximise fitness at each stage of this life cycle, and given the different events taking place within a biofilm, temporal regulation of gene expression is essential. We recently described the genes required for optimal fitness over time during biofilm formation in Escherichia coli using a massively parallel transposon mutagenesis approach called TraDIS-Xpress. We have now repeated this study in Salmonella enterica serovar Typhimurium to determine the similarities and differences in biofilm formation through time between these species. This work deepens understanding of the core requirements for biofilm formation in the Enterobacteriaceae whilst also identifying some genes with specialised roles in biofilm formation in each species.
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: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.