| The molecular pathogenic mechanism will be deeply understood with the identification of Shigella flexneri 2a virulence-related genes complicated. This study might lead to develop many better methods or means of controlling or preventing this disease such as dysenteric diarrhea. Since some Shigella flexneri 2a genes, including yphF wcaJ dam alkA cysG yaiC IS600 citC aroD CPO198, possibly related to virulence were isolated by using in vivo expression technology (IVET), these genes mutants were constructed and the relationship between these genes and the virulence of this pathogen was researched. First of all, we built a preliminary technology to quick knock out genes in Shigella flexneri 2a by using Red system. Along with serial experiments, we successfully knocked out Shigella flexneri 2a alkA gene. However, this system was firstly used in Shigella flexneri 2a and has some flaws needed to consummate. In order to accomplish the study within two years we constructed all these genes' insertion mutants with RecA system. To test the mutant strains for invasion ability and alterations in virulence to the wild type, the competition assays were carried out by using HeLa cells invasion model and BALB/c mice lung infection model. The dates we obtained indicates that most of the mutants strains' have lower competition indexes, lower capability of invasion and much more significant virulence defects, that might show that these genes related to the virulence of Shigella flexneri 2a and probably are the virulence genes of this pathogen. |
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