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Regulation of the environmental stress response and virulence of  Shigella flexneri  may involve multiple signaling pathways; however, these mechanisms are not well-defined. In bacteria, small regulatory RNAs (sRNAs) regulate bacterial growth, metabolism, virulence, and environmental stress response. Therefore, identifying novel functional sRNAs in  S. flexneri  could help elucidate pathogenic adaptations to host micro-environmental stresses and associated virulence. The aim of this study was to confirm the presence of an sRNA,  Ssr54 , in  S. flexneri  and to determine its functions and possible mechanism of action.  Ssr54  was found to regulate tolerance and virulence under hyperosmotic pressure. Its expression was verified by qRT-PCR and Northern blotting, and its genomic position was confirmed by 5′-rapid amplification of cDNA ends.  Ssr54  expression was significantly decreased (~ 80%) under hyperosmotic conditions (680 mM NaCl), and the survival rate of the  Ssr54  deletion strain increased by 20% under these conditions. This suggested that  Ssr54  has been selected to promote host survival under hyperosmotic conditions. Additionally, virulence assessment, including guinea pig Sereny test and competitive invasion assays in mouse lungs, revealed that  Ssr54  deletion significantly decreased  S. flexneri  virulence. Two-dimensional gel analyses suggest that  Ssr54  may modulate the expression of  tolC, ompA , and  treF  genes, which may affect the virulence and survival of  S. flexneri  under osmotic pressures. Furthermore,  treF  expression has been shown to improve the survival of  S. flexneri  under osmotic pressures. These results suggest that  Ssr54  has a broad range of action in  S. flexneri  response to hyperosmotic environmental stresses and in controlling its virulence to adapt to environmental stresses encountered during host infection.

A Novel sRNA in Shigella flexneri That Regulates Tolerance and Virulence Under Hyperosmotic Pressure