| Salmonellosis is a contagious disease caused by Salmonella in various animals,which lead to can cause sepsis,enteritis,can lead to pregnancy maternal abortion,food poisoning and acute gastroenteritis.Salmonella enteritidis is one of the major Gram-negative bacteria which can cause human food-borne diseases and the death of poultry.The disease results in serious economic losses,and the products carry Salmonella enteritidis,seriously endangering human health and becoming a public health concern.Therefore,to study an effective vaccine to control this disease has become an urgent task for researchers.In this study,a lysozyme plasmid based on pyroptosis-associated protein-mediated new bacterial ghost was constructed and then transformed into E.coli DH5α,in which the lysis rate can reach 99.99%.The lysozyme plasmid was transformed into Salmonella enteritidis by electroporation and then lysed under the induction of arabinose to prepare Salmonella ghost.The mice were immunized with the prepared salmonella ghost vaccine.There are 100mice aged 6-8 weeks randomly divided into five groups:Salmonella inactivated vaccine,Salmonella inactivated vaccine with adjuvant,ghost vaccine,new Salmonella ghost vaccine and control group,immunized three times.After immunization,blood was collected from the eyes and serum was separated.Salmonella ultrasound supernatant was used as envelop antigen,and serum IgG levels were detected by indirect enzyme-linked immunosorbent assay(iELISA).The splenocytes of immunized mice were stimulated.The stimulator was Salmonella enteritidis lipopolysaccharide(LPS).The secretion of cytokines IL-2,IL-4 and IFN-γwere detected by kit,and the dynamic changes of CD4~+and CD8~+cell subsets were detected by flow cytometry.After 1 week of challenge,clinical symptoms and number of surviving animals were observered protective effect was evaluated and histopathological observations were made.The results of IgG serum on immunized mice showed that with the increase of immunization times,the level of serum specific antibody in each experimental group gradually increased.The difference between the control group and the experimental group was extremely significant(P<0.001).The difference between the new ghost group and the inactivated vaccine group was extremely significant(P<0.001),and the new ghost group was significantly different from the inactivated vaccine+adjuvant group and the inactivated bacteria group(P>0.05).The results of cytokines IL-2,IL-4and IFN-γshowed that the cytokines of IL-2,IL-4 and IFN-γalso increased compared with the control group,which could cause strong cellular immunity.The dynamic changes of CD4~+and CD8~+subsets indicate that the vaccine group can induce lymphocyte proliferation.The difference between the experimental group and the control group was extremely significant(P<0.001),the difference between the new ghost group and the inactivated vaccine group was extremely significant(P<0.001),and there was no significant difference between the inactivated vaccine+adjuvant group and the inactivated vaccine group(P>0.05).After challenge,the protective effects of each expermental group were better than that of control group.The rate of protection was the same between new Salmonella ghost vaccine and Salmonella ghost vaccine,was 100%.And no obvious symptom was found after challenge.The protection rate of inactivated vaccines and inactivated vaccines+adjuvant was 30%and 70%,respectively.The results showed that the new Salmonella ghost vaccine can induce higher cellular and humoral immunity,and have good protective effect.The results provide a basis for the development of a new vaccine for Salmonella enteritidis. |
