EXPONENTIAL-PHASE EXPRESSION OF SPVA OF THE SALMONELLA-TYPHIMURIUM VIRULENCE PLASMID - INDUCTION IN INTRACELLULAR SALTS MEDIUM AND INTRACELLULARLY IN MICE AND CULTURED-MAMMALIAN-CELLS

The spy genes of Salmonella typhimurium and other non-typhoidal Salmon ella serovars are essential for efficient systemic infection beyond th e intestines in orally inoculated mice as a model for enteric fever. T hese virulence genes are not significantly expressed by salmonellae du ring exponential growth in L broth but are induced when the bacteria e nter the stationary phase of growth. Using RNase protection analysis t o directly measure spvA mRNA from the virulence plasmid of S. typhimur ium, we found that spvA was maximally induced in an SpvR- and RpoS-dep endent manner during exponential growth in Intracellular Salts Medium, which mimics the intracellular environment of mammalian cells. A clon ed spvA-lacZ operon fusion in S. typhimurium was induced intracellular ly in peritoneal cells of mice, correlating in vivo intracellular gene expression with intracellular function of the spy genes in infected m ice. spvA was also induced intracellularly in vitro within both Henle- 407 intestinal epithelial cells and J774.A1 macrophage-like cells when the bacteria were replicating with exponential kinetics. Prevention o f invasion of salmonellae with cytochalasin D inhibited spvA induction within tissue culture cells, indicating that salmonellae must be inte rnalized for spvA to be induced. The spvA-lacZ fusion was not induced by salmonellae in extracellular fluid of the peritoneal cavity or in s erum. Since induction of the spy genes occurs intracellularly during e xponential growth of salmonellae. cessation of growth may not be the m ost relevant inducing signal for spy gene expression.