Elucidating the Role of the Cpx Envelope Stress Response in Citrobacter rodentium colonization and virulence

Gastrointestinal attaching and effacing pathogens (A/E), such as Enteropathogenic and Enterohemorrhagic Escherichia coli (EPEC and EHEC), are prominent food-borne pathogens associated with severe diarrheal disease. Citrobacter rodentium, a murine A/E pathogen with genetic similarities to EPEC and EHEC, is used as a model to study these pathogens in vivo. While infecting, C. rodentium must adapt to stresses in the host environment such as antimicrobial peptides, changes in pH, host immune responses and competition with commensal microorganisms. The Cpx envelope stress response (ESR) is a two-component system used by C. rodentium to mediate stress by modulating gene expression and is necessary for colonization and virulence. To determine in what way the Cpx ESR contributes to the pathogenesis of C. rodentium, RNA-Seq data from previous research was analyzed comparing the expression profiles of genes in wildtype C. rodentium DBS100 and a ΔcpxRA mutant. Genes that were up-regulated in the presence of CpxRA and had a potential to promote virulence or sustain colonization were then chosen. lux-reporters were constructed to confirm the regulation of these genes in the presence of CpxRA. The genes yebE, ygiB, bssR, and htpX were determined to be strongly up-regulated by CpxRA and had further increased expression when subjected to alkaline pH, which is thought to induce the Cpx ESR. In addition, their expression over time resembled that of the positive control, cpxP, further indicating a reliance of their expression on the presence of CpxRA. Using sacB¬-based allelic exchange, deletion mutants for yebE, ygiB, bssR, and htpX were generated. The mutants will also be tested in vivo to determine their relative pathogenicity in mice compared to that of wildtype C. rodentium DBS100. The goal of this research is to determine if there are specific genes controlled by the Cpx ESR that are required for successful colonization of C. rodentium in mice.