Role of RpoS in the Virulence of Citrobacter rodentium

Abstract

Citrobacter rodentium is a mouse enteropathogen that is closely related to Escherichia coli and causes severe colonic hyperplasia and bloody diarrhea. C. rodentium infection requires expression of genes of the locus of enterocyte effacement (LEE) pathogenicity island, which simulates infection by enteropathogenic E. coli and enterohemorrhagic E. coli in the human intestine, providing an effective model for studying enteropathogenesis. In this study we investigated the role of RpoS, the stationary phase sigma factor, in virulence in C. rodentium. Sequence analysis showed that the rpoS gene is highly conserved in C. rodentium and E. coli, exhibiting 92% identity. RpoS was critical for survival under heat shock conditions and during exposure to H 2 O 2 and positively regulated the expression of catalase KatE (HPII). The development of the RDAR (red dry and rough) morphotype, an important virulence trait in E. coli, was also mediated by RpoS in C. rodentium. Unlike E. coli, C. rodentium grew well in the mouse colon, and the wild-type strain colonized significantly better than rpoS mutants. However, a mutation in rpoS conferred a competitive growth advantage over the wild type both in vitro in Luria-Bertani medium and in vivo in the mouse colon. Survival analysis showed that the virulence of an rpoS mutant was attenuated. The expression of genes on the LEE pathogenicity island, which are essential for colonization and virulence, was reduced in the rpoS mutant. In conclusion, RpoS is important for the stress response and is required for full virulence in C. rodentium.Intestinal diseases caused by bacterial infection, such as the infection caused by Escherichia coli O157:H7, are major threats to public health (40). It is imperative to fully understand how the pathogens that cause these diseases are transmitted, propagate, and cause disease in the host. During transmission, pathogens likely have to survive many stresses, including environmental stresses (e.g., nutrient limitation) and host internal stresses (e.g., acidic exposure in the stomach and host defense), implying that stress response systems are important. One of the most important regulators in the stress response is RpoS, an alternative sigma factor of RNA polymerase that is present primarily in gammaproteobacteria, including E. coli and Salmonella (11,20).RpoS is important for cell survival in stress conditions, such as oxidative stress and exposure to acid, in many pathogens, including Salmonella sp. (15), Vibrio cholerae (59), Pseudomonas aeruginosa (50), and Yersinia enterocolitica (22). However, RpoS has distinct roles in the pathogenesis of these organisms. RpoS is essential for virulence in Salmonella (15) and is important for the invasion of brain microvascular endothelial cells in E. coli K1 strains (55), but it is not required for virulence in P. aeruginosa (50) and Y. enterocolitica (22). The effects of RpoS on virulence may also differ within a species. For example, RpoS has been found to be either important (35) or not required (26, 59) for colon...

The Serine Protease Autotransporter Pic Modulates Citrobacter rodentium Pathogenesis and Its Innate Recognition by the Host

“…This medium allows the assessment of bacterial growth under limiting nutrient conditions and the production of what is termed the RDAR morphotype. This phenotype reflects an aggregative and resistant physiology that has been linked to survival in nutrient-limited environments and previously was shown to indicate the production of extracellular components, such as curli fimbriae and cellulose (35)(36)(37)(38). Notably, the C. rodentium ⌬picC mutant exhibited a more pronounced RDAR morphotype than the WT strain (Fig.…”

“…An increased propensity of the ⌬picC C. rodentium mutant to aggregate in vivo may be linked to its in vitro ability to produce the RDAR morphotype. As previously suggested, the RDAR phenotype represents an aggregative state due in part to the production of extracellular matrix components, such as curli and cellulose (35)(36)(37)(38). The C. rodentium ⌬picC mutant's RDAR morphotype (potentially linked to its ability to produce higher levels of cellulose and a modest increase in its ability to form biofilms in vitro) may be analogous to the pathogen's ability to form in vivo microcolony-like structures associated with the intestinal mucus layer, thereby decreasing the ability of the pathogen to shed into the lumen.…”

Phenotypic Diversity Caused by Differential RpoS Activity among Environmental Escherichia coli Isolates

“…Overnight cultures of environmental isolates in LB were washed with 0.9% NaCl and subcultured to ϳ10 7 cells into LB medium containing 15 mM H 2 O 2 . Cultures then were incubated at 37°C with shaking at 200 rpm, and CFU/ml was determined over time by serially plating onto LB plates (18,45). The percent survival was calculated as (final CFU/ml)/(initial CFU/ml) ϫ 100.…”

Role of RpoS in Virulence of Pathogens

“…Infection of mice using C. rodentium provides a promising alternative model to study enteropathogenesis in natural hosts (97,143). The rpoS mutant of C. rodentium is more sensitive to heat and oxidative stress than the wild type, indicating a conserved RpoS function (30). Colonization and virulence of C. rodentium are attenuated in rpoS mutants during infection in mice (30).…”