| Pig erysipelas and Streptococcus suis(SS)are an acute,hot zoonotic infectious disease caused by E.rhusiopathiae and SS,which not only cause serious economic losses to the pig industry,but also threaten the health and safety of human beings.Serotype 1a E.rhusiopathiae and type 2 Streptococcus suis(SS2)are the strains with the highest separation rate and the strongest pathogenicity in Chinese pigs.Under the general trend of reducing resistance,limiting resistance and gradually moving towards prophylaxis,vaccination is undoubtedly the most effective measure to prevent and control swine erysipelas and Streptococcus suis.Inactivated seedlings have been widely used in clinical practice due to their advantages such as short development period,easy preservation,safe use and better preparation of multi-linked seedlings.The use of multi-linked vaccine can achieve the purpose of one needle to prevent multiple diseases.At present,there are no reports of commercial swine erysipelas and Streptococcus suis disease.This research is to early by pathogenicity,antigenicity and stability test,select the type 1 a"E.rhusiopathiae and SS2 as seedlings strains,develop swine erysipelas and pig streptococcus disease duplex inactivated seedlings and use the mice immune effect evaluation,aimed to further implement the duplex inactivated seedlings,laying a foundation for the clinical application for effective prevention and control to provide technical reserve swine erysipelas and pig streptococcus disease.This research use of seedlings strains screened type 1 A"E.rhusiopathiae(code AEr31)and SS2(code HF2)for the following test:(1)with formaldehyde for inactivating agent,ISA 201 VG mineral oil adjuvant,using two ways of duplex inactivated seedlings[A:(V-AEr31-HF2)+V+ISA 201 VG,B:(V-AEr31+ISA 201 VG)+(V-HF2+ISA 201 VG)],examined the traits,sterile test and safety test vaccine not approved or not.(2)the mice were immunized with two-line inactivated vaccines prepared by the two methods,and serum Ig G antibody levels and cytokine levels(IL-4,IL-10,IFN-γ,TNF-β,MCP-1)were detected by indirect ELISA.Serum bactericidal test to detect the level of functional antibody;The proliferation index of spleen lymphocytes was determined by MTT method.The percentages of CD4~+and CD8~+T lymphocyte subsets were determined by flow cytometry.Abdominal attack test was used to determine the immune protection rate,and the number of bacteria in tissues was used to determine the colonization of the attack strains in mice,and the pathological sections of lung,liver,spleen and kidney were prepared to observe the pathological changes.The results showed that:(1)The two-in-one inactivated seedlings prepared by the two methods were water-in-oil and water-in-water(W/O/W),which all met the qualification requirements of viscosity test,stability test,asepsis test and safety test.(2)During the whole immunization stage,there was no significant change in the detection indexes of the sterilized normal saline in the control group C,and both groups A and B could produce good humoral immunity and cellular immunity,with no significant difference between the two groups(P>0.05).Among them,the serum Ig G antibody levels of mice in groups A and B continued to increase and were significantly higher than those in group C(P<0.05).The serum functional antibody levels of mice in groups A and B continued to increase and were significantly higher than those in group C(P<0.05).The levels of related cytokines(IL-4,IL-10,IFN-γ,TNF-β,MCP-1),the proliferation index of spleen lymphocytes,and the percentages of CD4~+and CD8~+T cell subsets in group A and group B continued to increase and were significantly higher than that in group C(P<0.05).After the challenge,the protective rate of the mixed bacterial fluid of group A and group B to E.rhusiopathiae,SS and E.rhusiopathiae-ss was 100%.Both group A and group B could effectively prevent the infection of e.rhousiiae and SS on mice.The bacterial colonization amount of all organs(lung,liver,kidney,spleen)reached the minimum 14 days after erxun,and was significantly lower than that of group C(P<0.05).Compared with group C,all organ lesions in groups A and B were mild and there was no significant difference between the two groups.Results show that:(1)The research on AEr31 and HF2 seedlings strains,formaldehyde for inactivating agent,ISA 201 VG mineral oil adjuvant,the inactivated bacteria liquid and adjuvant prepared two different hybrids character stable,sterile test,safety test qualified swine erysipelas thermosphacta-streptococcus suis 2 inactivated seedlings,respectively named(V-AEr31+V-HF2)+ISA 201 VG and(V-AEr31+ISA201 VG)+(V-HF2+ISA 201 VG).(2)Kunming mice as experimental animal models,the(V-AEr31-HF2)+V+and ISA201VG(V-AEr31+ISA 201 VG)+(V-HF2+ISA201 VG)immune effect evaluation,there was no significant difference between the two,immune mice after all can produce good humoral immunity and cellular immunity was100%against"E.rhusiopathiae,SS virulent strain of attacks,for wire swine erysipelas bacteria-streptococcus suis duplex inactivated seedlings commercial basis,To provide new ideas and technical reserves for the prevention and control of swine erysipelas and Streptococcus suis. |
