The Shigella ProU system is required for osmotic tolerance and virulence.

To cope with hyperosmotic stress encountered in the environments and in the host, the pathogenic as well as non-pathogenic microbes use diverse transport systems to obtain osmoprotectants. To study the role of Shigella sonnei ProU system in response to hyperosmotic stress and virulence, we constructed deletion and complementation strains of proV and used an RNAi approach to silence the whole ProU operon. We compared the response between wild type and the mutants to the hyperosmotic pressure in vitro, and assessed virulence properties of the mutants using gentamicin protection assay as well as Galleria mellonella moth larvae model. In response to osmotic stress by either NaCl or KCl, S. sonnei highly up-regulates transcription of proVWX genes. Supplementation of betaine greatly elevates the growth of the wild type S. sonnei but not the proV mutants in M9 medium containing 0.2 M NaCl or 0.2 M KCl. The proV mutants are also defective in intracellular growth compared with the wild type. The moth larvae model of G. mellonella shows that either deletion of proV gene or knockdown of proVWX transcripts by RNAi significantly attenuates virulence. ProU system in S. sonnei is required to cope with osmotic stress for survival and multiplication in vitro, and for infection.