Immune response induced by Salmonella typhimurium defective in ppGpp synthesis.
ABSTRACT Systemic infection by Salmonella typhimurium requires coordinated expression of virulence genes found primarily in Salmonella Pathogenecity Islands (SPIs). We have previously reported that the intracellular signal that induces these virulence genes is a stringent signal molecule, ppGpp [Song et al. J Biol Chem 2003;279:34183]. In this study, we found that relA and spoT double mutant Salmonella (DeltappGpp strain), which is defective in ppGpp synthesis, was virtually avirulent in BALB/c mice. Subsequently, the live vaccine potential of the avirulent DeltappGpp Salmonella strain was determined. A single immunization with live DeltappGpp Salmonella efficiently protected mice from challenge with wild-type Salmonella at a dose 10(6)-fold above the LD50 30 days after immunization. Various assays revealed that immunization of mice with the DeltappGpp strain elicited both systemic and mucosal antibody responses, in addition to cell-mediated immunity.
- Journal of Bacteriology and Virology 01/2009; 39(2).
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ABSTRACT: During the last decade, an increasing number of papers have described the use of various genera of bacteria, including E. coli and S. typhimurium, in the treatment of cancer. This is primarily due to the facts that not only are these bacteria capable of accumulating in the tumor mass, but they can also be engineered to deliver specific therapeutic proteins directly to the tumor site. However, a major obstacle exists in that bacteria because the plasmid carrying the therapeutic gene is not needed for bacterial survival, these plasmids are often lost from the bacteria. Here, we report the development of a balanced-lethal host-vector system based on deletion of the glmS gene in E. coli and S. typhimurium. This system takes advantage of the phenotype of the GlmS(-) mutant, which undergoes lysis in animal systems that lack the nutrients required for proliferation of the mutant bacteria, D-glucosamine (GlcN) or N-acetyl-D-glucosamine (GlcNAc), components necessary for peptidoglycan synthesis. We demonstrate that plasmids carrying a glmS gene (GlmS(+)p) complemented the phenotype of the GlmS(-) mutant, and that GlmS(+) p was maintained faithfully both in vitro and in an animal system in the absence of selection pressure. This was further verified by bioluminescent signals from GlmS (+)pLux carried in bacteria that accumulated in grafted tumor tissue in a mouse model. The signal was up to several hundred-fold stronger than that from the control plasmid, pLux, due to faithful maintenance of the plasmid. We believe this system will allow to package a therapeutic gene onto an expression plasmid for bacterial delivery to the tumor site without subsequent loss of plasmid expression as well as to quantify bioluminescent bacteria using in vivo imaging by providing a direct correlation between photon flux and bacterial number.PLoS ONE 01/2013; 8(3):e60511. · 3.53 Impact Factor
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ABSTRACT: Burkholderia pseudomallei is a Gram-negative soil bacterium and the causative agent of melioidosis, a disease of humans and animals. It is also listed as a category B bioterrorism threat agent by the U.S. Centers for Disease Control and Prevention, and there is currently no melioidosis vaccine available. Small modified nucleotides such as the hyperphosphorylated guanosine molecules ppGpp and pppGpp play an important role as signaling molecules in prokaryotes. They mediate a global stress response under starvation conditions and have been implicated in the regulation of virulence and survival factors in many bacterial species. In this study, we created a relA spoT double mutant in B. pseudomallei strain K96243, which lacks (p)ppGpp-synthesizing enzymes, and investigated its phenotype in vitro and in vivo. The B. pseudomallei ΔrelA ΔspoT mutant displayed a defect in stationary-phase survival and intracellular replication in murine macrophages. Moreover, the mutant was attenuated in the Galleria mellonella insect model and in both acute and chronic mouse models of melioidosis. Vaccination of mice with the ΔrelA ΔspoT mutant resulted in partial protection against infection with wild-type B. pseudomallei. In summary, (p)ppGpp signaling appears to represent an essential component of the regulatory network governing virulence gene expression and stress adaptation in B. pseudomallei, and the ΔrelA ΔspoT mutant may be a promising live-attenuated vaccine candidate.Infection and immunity 07/2012; 80(9):3247-55. · 4.21 Impact Factor