Characterisation of the Pathogenesis and Immunogenicity of Two Novel Attenuated Mutants of Salmonella Typhimurium

Salmonella enterica causes a wide range of diseases in many different hosts. Previously we used novel technology to screen the genome of S. enterica serovar Typhimurium to identify genes required for full virulence of the bacterium in a mouse model of invasive salmonellosis. <P>
We must now follow up this study by characterising thoroughly some of the more interesting mutants. We thus propose to characterise atpA and trxA mutants, which we have shown in preliminary studies to be highly attenuated and protective when used as live vaccines. Both of these mutants induce minimal to no inflammation in the spleens of infected animals, thus using these mutations in vaccine strains may be a way of reducing adverse reactions. <P>
In this study the basis for the attenuation, lack of inflammation and immunogenicity will be thoroughly characterised using microbiological, cell biological, and immunological techniques. Mutants in the other genes in the atp operon and in other trx genes will be constructed and tested for their degree of attenuation. Mutants will be subjected to a range of tests and assays, for example resistance to acid and reactive oxygen species, to investigate whether their lack of virulence results from reduced resistance to these challenges. The ability of the mutants to survive in primary macrophages from wild-type and knock-out mouse strains will be tested. The ability of the mutants to induce pro-inflammatory signals in cells and animals of various genetic types will be tested to investigate the mechanism by which inflammation is not induced. Finally a full range of immunological parameters will be tested and correlated with the different degrees of protection provide by these mutants.<P>
This project will provide detailed information about these new candidate vaccine strains, as well as considerable data concerning basic parameters and mechanisms of attenuation, virulence, inflammation and protective immunity in invasive salmonellosis.