The CpxA/R system mediates the regulation of TolC on biofilm formation by extraintestinal pathogenic Escherichia coli in response to medium osmolarity

AIMS: Hyperosmolarity is widely used for food preservation by inhibiting bacterial survival and growth. Therefore, it is of great significance to reveal bacterial osmotic-response mechanism. Biofilm formation presents a significant challenge for the control and prevention of pathogenic bacteria. Our previous study showed that inactivation of the efflux protein TolC in extraintestinal pathogenic Escherichia coli (ExPEC) decreased biofilm formation by affecting curli production in a medium osmolarity-dependent manner. This study aims to explore the role of the two-component CpxA/R system in mediating TolC regulation of ExPEC biofilm formation in response to osmolarity. METHODS AND RESULTS: Various mutants derived from the parental ExPEC ΔtolC strain were constructed, and their abilities to form biofilms and produce curli fimbriae in different osmotic media were evaluated using crystal violet staining, scanning electron microscopy, Congo red assay, and real-time quantitative polymerase chain reaction. The results showed that the disruption of CpxA/R system by deleting the gene encoding histidine kinase-CpxA or response regulator-CpxR, or by introducing a point mutation at the phosphorylation site of CpxA, significantly compromised the effect of TolC inactivation on ExPEC biofilm formation and curli biosynthesis under both NaCl- and sucrose-induced osmotic stresses. CONCLUSIONS: Our study firstly demonstrate that the CpxAR system mediated the regulation of TolC inactivation on ExPEC biofilm formation and curli production in response to both NaCl- and sucrose-induced osmotic stresses. These findings expand the regulatory network of bacterial biofilm formation and osmotic-responsiveness, contributing to exploring potential targets for preventing and controlling pathogenic bacteria.