Investigation of the regulation of expression of E. Coli Common Pilus subunit, ECPA, of enterohaemorrhagic E. Coli 0157:H7 under acid stress
thesisposted on 2021-05-23, 17:05 authored by Hannah Tollman
Escherichia coli (EHEC) 0157:H7 is a pathogenic bacterial species that is most commonly linked to severe diarrhea, but is also the leading cause of the potentially fatal hemolytic-uremic syndrome (HUS). In order to establish infection in the colon, EHEC must endure different stresses encountered in the gastrointestinal (GI) tract, such as acid stress in the stomach, bile salt stress in the small intestine, and short-chain fatty acid (SCFA) stress in the colon. These bacteria are likely able to use GI stresses as indicators of their location, impacting gene expression of adhesion, motility, and virulence factors. The E. coli Common Pilus (ECP) has been shown to be an important factor for EHEC adhesion to epithelial cells, which is increased after either acid or SCFA stress. It has also been demonstrated via microarray that genes of this operon are upregulated after acid stress. The aim of this study is to determine how expression of the main subunit of this structure, EcpA, is regulated upon exposure of EHEC 0157:H7 to acid or SCFA-stress. Both transcriptional and translational regulation are hypothesized to be involved. Isogenic mutants have been constructed that lacked key regulators suspected to be important for each system. Two approaches are used to determine if the predicted regulatory systems are playing a role in response to stress: observing EcpA protein expression analysis through Western blotting with anti-EcpA antibodies, and examining differences in ecp operon promoter activity in regulatory mutants. In this study Western blots reconfirmed H-NS does not modulate ecpA expression in direct response to acute acid stress. This suggests an alternate regulatory response in EHEC 0157:H7 to acute acid stress resulting in the upregulation of ecpA expression previously observed with microarray analysis.