Bacterial pathogens undergo profound physiological changes when they infect their hosts, requiring coordinated regulation of gene expression in response to the stresses encountered during infection. A number of environmental stresses (such as osmotic shock and anaerobiosis) have been shown to induce changes in DNA supercoiling that can directly affect the transcription of a specific subset of bacterial genes, at least some of which (the outer-membrane porins and type 1 fimbriae) play a part in bacterial virulence. Here, we demonstrate that the virR gene of S. flexneri, implicated in the temperature regulation of plasmid-encoded virulence genes, is equivalent to the osmZ gene of Escherichia coli, which has previously been shown to mediate its regulatory effects through changes in DNA supercoiling. Our results imply that environmentally induced changes in DNA supercoiling are important in the coordinated control of virulence gene expression in S. flexneri and have general implications for the control of bacterial virulence.