Etude du role du regulon Phosphate (Pho) dans la virulence de souches d'Escherichia coli pathogenes causant des maladies extra-intestinales (ExPEC) et de son influence sur la surface bacterienne

摘要

This work concerns the role of the Pho regulon in the virulence of extra-intestinal pathogenic Escherichia coli (ExPEC) strains as well as its influence on the bacterial cell surface. The Pho regulon is controlled by the two-component regulatory system PhoR/PhoB. It is the major gene circuit involved in adaptation to phosphate limitation. The Pst system is involved in phosphate specific transport. A mutation in Pst results in a constitutive expression of the Pho regulon. The ExPEC strains O115 and O78 cause important economical loss in the swine and poultry industry respectively. Our research group showed that in an O115 strain, a mutation in the Pst system results in pleiotropic effects and virulence loss.;Our laboratory is interested in elucidating the molecular basis behind these phenomena. We hypothesized that a mutation in the Pst system results in structural changes of the cell surface components. Thus, this project was to characterize cell surface structures using Pst mutants of ExPEC strains O115 and O78. We aimed to establish if a Pst mutation has similar effects on ExPEC strains from distinct animal origins and to determine which effects can be correlated to bacterial virulence.;The virulence of Pst mutants was evaluated in experimental infection models. We showed that a mutation in Pst results in an important virulence attenuation, including a decreased capacity to cause extra-intestinal lesions, in both O115 and O78 strains. Moreover, we showed that Pst mutants are sensitive to the bacteriolytic effects of serum, to cationic antimicrobial peptides (CAMP) and to acid shock. These stresses can be encountered within the host. The cell surface of Gram-negative bacteria acts as a major barrier to many environmental cues. More specifically, the lipid A which is the membrane anchor of the lipopolysaccharides (LPS), is a known structure involved in CAMP resistance. Our lipid A analyses, revealed a 66% reduction in the amount of the hexa-acylated 1-pyrophosphate lipid A in Pst mutants. This lipid A modification could contribute to serum and CAMP sensitivity by reducing cross-bridging between LPS molecules. In addition, our results showed that increased outer-membrane permeability as well as changes in fatty acids composition is likely to contribute to acid shock sensitivity in Pst mutants.;In conclusion, we showed that ExPEC strains O115 and O78 are similarly affected by a mutation in Pst. The phenotypes identified in this study confirmed that Pst mutants are indeed characterized by cell surface structural changes. Moreover, our results showed that the Pho regulon, which is constitutive in pst mutants, is involved in some of these modifications. Thus, the Pho regulon might be included in a more global network of regulation affecting phosphate homeostasis, bacterial virulence and stress responses. Finally, drugs that induce the expression of the Pho regulon may also be useful as therapeutic agents that would compromise bacterial virulence and facilitate elimination of the pathogen by host defences.;Key words: ExPEC, Pho regulon, TCS, Pst system, LPS, lipid A, Fatty acids, CAMP, acid shock, serum.