Transcriptional regulation of Salmonella enterica virulence plasmid genes in cultured macrophages

Abstract

The plasmid-carried spv genes promote virulence of salmonellae in mice by allowing bacterial growth in the reticuloendothelial tissue. When the bacteria are cultivated under normal laboratory conditions the spv genes appear dormant. This communication explores the transcriptional regulation of spv genes within murine macrophage-like J774-A.1 cells utilizing a new reporter system. Transcriptional fusions were constructed between promoter elements of the Salmonella enterica var. Typhimurium spv genes and the KS71A fimbrial gene cluster. The expression of KS71A fimbriae In fusion-carrying Escherichia coli strains was found to be under the control of the transcriptional activator gene spvR. In strains overproducing SpvR, KS71A fimbriae were assembled on the bacterial cell surface and could be detected by bacterial agglutination or immunofluorescence of intact bacteria; the reporter activity was quantified by estimating the percentage of fluorescent bacteria and by immunoblotting of cell lysates. The activity of the reporters, when transformed into the parent Typhimurium TML R66, was low and revealed less than 0.3% fimbriated cells under in vitro culture conditions. A 15–30-fold increase in fimbriation was observed when the bacteria were cultivated within J774-A.1 cells. No such increase occurred when the reporter fusions were transformed into TML R66 cured of the virulence plasmid. Insertional inactivation of the spvR gene of the virulence plasmid in Typhimurium TML R66 also abolished induction, whereas corresponding inactivation of spvA or spvB did not reduce induction. No increase in reporter activity was obtained in Typhimurium of line Q1, which is naturally avirulent for mice, although the strain was provided with virulence plasmid pEX102 of line TML R66. We conclude that the intracellular environment of J774-A.1 cells induces the spv genes and that this induction requires gene functions of both the bacterial chromosome and the virulence plasmid.