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:Avian pathogenic Escherichia coli strains frequently cause extra-intestinal infections and are responsible for significant economic losses in the poultry industry worldwide. APEC isolates are closely related to human extraintestinal pathogenic E.coli strains and may also act as pathogens for humans. In this work, three type VI secretion systems were deleted to analyze which pathogenicity characteristics would change in the mutants, compared to wild type strain (SEPT 362).
Project description:The antibiotic fosfomycin is widely recognized for treatment of lower urinary tract infections caused by Escherichia coli and lately gained importance as a therapeutic option to combat multidrug resistant bacteria. Still, resistance to fosfomycin frequently develops through mutations reducing its uptake. Whereas the inner membrane transport of fosfomycin has been extensively studied in E. coli, its outer membrane (OM) transport remains insufficiently understood. While evaluating minimal inhibitory concentrations in OM porin-deficient mutants, we observed that the E. coli ΔompCΔompF strain is five times more resistant to fosfomycin than the wild type and the respective single mutants. Continuous monitoring of cell lysis of porin-deficient strains in response to fosfomycin additionally indicated the relevance of LamB. Furthermore, the physiological relevance of OmpF, OmpC and LamB for fosfomycin uptake was confirmed by electrophysiological and transcriptional analysis. This study expands the knowledge of how fosfomycin crosses the OM of E. coli.
Project description:We measured the mRNA abundance in E.coli using RNAseq to calculate mRNA lifetimes. The data is used in support of a larger paper on the proteome and transcriptome of E.coli.
Project description:We compared the transcriptional profiles of 12 E. coli O157:H7 isolates grown to stationary phase in LB broth. These isolates possess the same two enzyme PFGE profile and are related temporally or geographically to the above outbreak. These E. coli O157:H7 isolates included three clinical isolates, five isolates from separate bags of spinach, and single isolates from pasture soil, river water, cow feces, and a feral pig.
Project description:Avian pathogenic Escherichia coli strains frequently cause extra-intestinal infections and are responsible for significant economic losses in the poultry industry worldwide. APEC isolates are closely related to human extraintestinal pathogenic E.coli strains and may also act as pathogens for humans. In this work, three type VI secretion systems were deleted to analyze which pathogenicity characteristics would change in the mutants, compared to wild type strain (SEPT 362). Four Avian Pathogenic Escherichia coli strains (one wild type and three deleted mutants) were grown at 37°C in Dulbecco´s Modified Eagle´s Media (DMEM) media until reach O.D 600 = 0.8, for RNA extraction and hybridization on Affymatrix microarrays.
