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:Shiga toxin-producing Escherichia coli (STEC) O157:H7 is a notorious foodborne pathogen capable of causing severe gastrointestinal infections in humans. The bovine rectoanal junction (RAJ) has been identified as a primary reservoir of STEC O157:H7, playing a critical role in its transmission to humans through contaminated food sources. Despite the relevance of this host-pathogen interaction, the molecular mechanisms behind the adaptation of STEC O157:H7 in the bovine RAJ and its subsequent infection of human colonic epithelial cells remain largely unexplored. This study aimed to unravel the intricate dynamics of STEC O157:H7 in two distinct host environments: bovine RAJ squamous epithelial (RSE) cells and human colonic epithelial cells. Comparative transcriptomics analysis was employed to investigate the differential gene expression profiles of STEC O157:H7 during its interaction with these cell types. The bacterial cells were cultured under controlled conditions to simulate the microenvironments of both bovine RAJ and human colonic epithelial cells. Using high-throughput RNA sequencing, we identified key bacterial genes and regulatory pathways that are significantly modulated in response to each specific host environment. Our findings reveal distinct expression patterns of virulence factors, adhesion proteins, and stress response genes in STEC O157:H7 grown in bovine RAJ cells as opposed to human colonic epithelial cells. Additionally, the comparative analysis highlights the potential role of certain genes in host adaptation and tissue-specific pathogenicity. Furthermore, this study sheds light on the potential factors contributing to the survival and persistence of STEC O157:H7 in the bovine reservoir and its ability to colonize and cause disease in humans.
Project description:Two outbreak strains of E. coli O157:H7 differ phylogenetically, in gene content, and in epidemiological characteristics. The working hypothesis in this experiment was that these strains will also differ in the transcription of shared virulence genes. Indeed, following a 30 minute exposure to epithelial cells, strain TW14359 overexpressed major and ancillary virulence genes, relative to strain Sakai.
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. Twelve condition experiment, 12 E. coli O157:H7 isolates. Two biological replicates for isolates RM6067, RM6069, RM6101, RM6102, RM6103, RM6149, RM6655, RM6658, RM9992, RM9997, RM9998 and RM10002 independently grown to stationary phase in LB at 37°C and harvested. One replicate per array. A type 2 gene expression experimental design was used, with fluorescently labeled genomic DNA as a reference channel in each experiment as described by Lucchini, S., et al. 2005. Infect Immun 73:88-102.
Project description:Two outbreak strains of E. coli O157:H7 differ phylogenetically, in gene content, and in epidemiological characteristics. The working hypothesis in this experiment was that these strains will also differ in the transcription of shared virulence genes. Indeed, following a 30 minute exposure to epithelial cells, strain TW14359 overexpressed major and ancillary virulence genes, relative to strain Sakai. E. coli O157:H7 strains were physiologically normalized by growth to stationary phase, twice, in MOPS minimal media. Cultures were then transferred to DMEM media for adaptation. After 3 h of growth in DMEM, O157:H7 cultures were used to infect monolayers of MAC-T epithelial cells. 30 min following incubation, aliquots of suspended, non-adherent bacteria were used for RNA extraction. Five biological replications of the experiment were performed with each strain and, together with dye-swaps, 10 array hybridizations were carried out. Array data were fitted to a mixed model ANOVA using the following linear model: array+dye+sample (biological replicate)+strain+error.
Project description:Deletion of yedL was found to signifcantly decrease type three secretion in EHEC O157:H7. Transcriptional profiles of Escherichia coli O157: H7 and the isogenic yedL mutant were generated and compared.
Project description:Deletion of yhaO was found to signifcantly decrease type three secretion in EHEC O157:H7. Transcriptional profiles of Escherichia coli O157: H7 and the isogenic yhaO mutant were generated and compared.
