The Effects Of SseB Localizations In Salmonella Enterica On Induced Cross Protective Efficacies

Salmonella is a gram-negative,facultative intracellular parasitic bacteria that causes foodborne diseases in human and a variety of animals,SseB is a secreted protein that is considered one of the protective antigens of Salmonella.In this study,the protective antigen SseB was expressed on the extracellular,cell membrane and intracellular in attenuated Salmonella Typhimurium.The immunoprotective effect of three recombinant strains on Salmonella Typhimurium and Salmonella Enteritidis was compared in mouse models,then the vaccine strains with cross-immune protection were screened.1 Construction of recombinant strains of Salmonella TyphimuriumPlasmids expressing the Salmonella Typhimurium secreted protein SseB on extracellular,membrane and intracellular were constructed by seamless cloning,and three plasmids were transferred into attenuated Salmonella Typhimurium(Δaro A)to obtain three recombinant strains of Salmonella Typhimurium(Sec/Sur/Cyto-SseB-Δaro A).The protein of interest was detected in the supernatant of Sec-SseB-Δaro A culture by western blotting,in the cell lysate and membrane components of Sur-SseB-Δaro A,and in the cell lysate sample of Cyto-SseB-Δaro A.The above results indicate the successful construction of three recombinant strains expressing SseB on extracellular,membrane and intracellular areas.2 Study on biological characteristics of recombinant strains of Salmonella TyphimuriumThe results of biological characteristics showed that the growth rates of the three recombinant Salmonella Typhimurium were similar to the parent strain(Δaro A).The recombinant plasmids were stably preserved and expressed in the vector strain.Virulence and colonization tests showed that the virulence and colonization capacity of the three attenuated recombinant strains were similar to the parent strains,but still showing reduced compared with the wild type strains.The above results showed the high safety of three recombinant Salmonella Typhimurium and high stability of recombinant plasmids,and the expression of SseB had no significant effect on the growth of the strain itself.3 Evaluation of immunogenicity and cross-protection of vaccine strainsBALB/c mice were immunized and booster immunized with three recombinant strains,and the ELISA results showed that at the seventh weeks after first immunization,three immune groups produced high levels of serum Ig G response against homologous Salmonella Typhimurium whole bacteria antigen.Immunity of recombinant strains reduced the bacterial colonization of Salmonella Typhimurium in tissues and organs,and all three vaccine strains provided 100% homologous protection.Immunity of recombinant strains expressing SseB on the membrane and intracellular could not reduce the colonization of Salmonella Enteritidis in tissues and organs,and not protect mice against lethal infection after Salmonella Enteritidis challenge.However,the recombinant strain secreting SseB was significantly reduced the colonization amount of Salmonella Enteritidis in tissues and organs,resulting in a 37.5% protection rate.The above results show that the Sec-SseBΔaro A vaccine strain can prevent Salmonella Typhimurium and prevent Salmonella Enteritidis to some extent.