Transferrin Binding Protein A From Serum Type 13 And 14 Of Haemophilus Influenzae Cross Immune Protection Study By Guinea Pigs

Haemophilus parasuis is a commensal bacteria of the upper respiratory tract of pigs,the bacteria can cause the main clinical symptoms of Haemophilus parasuis disease like neurological symptoms,serositis,arthritis,fever and dyspnea,the disease was distributed worldwide in recent years and showing on the upward trend.the disease has become an important bacterial infection,which could caused serious economic losses.Due to the irrational use of veterinary drugs for a long time,the resistance of HPS is increasing,and multi drug resistant strains are also began to be discovered.Therefore,one of the most effective ways to prevent Haemophilus influenzae disease is vaccine control.but,HPS has obvious regional characteristics.Moreover,the rate of mutual protection between different serotype strains has low effect.At present,commercial vaccines can not provide sufficient preventive effect to all serum types.Therefore,it is significant to reaserch new vaccines with high cross immunity protection.The previous of immune challenge test by guinea pigs in our lab was proved that HPS serum type 13 transferrin binding protein A have high immune protection to the same serum strains.The protein can also provide certain cross protection to serum type 4.However,It is unknown that if the other serum types of HPS's TbpA have high immunity protection and cross immune protection.This study explored the different serum types of HPS's TbpA immune protection and cross immunity protection by guinea pigs,so as to provide the basis for further research the animal?pigs?.Use the HPS serum type 13 and type 14 recombinant TbpA immune guinea pigs,then detecting after immunization in guinea pig serum IgG antibody levels and cytokine levels?TNF-?,IFN-?,IL-2?IL-5?IL-8,MCP-1?,observe the infected guinea pigs'food intake,mental state and mortality,pathological changes and pathological changes.First,Determine the median lethal dose(LD₅₀)of HPS serum types 13 and 14 to guinea pigs with the cumulative dose method?Reed-Muench?.Secondly,Screen 54healthy 40d HPS antibody negative female guinea pigs,each group were randomLy assigned to 6 guinea pigs,a total of 9 groups;A1,A2 groupimmune serum type 13HPS's TbpA with 50?g each;B1,B2 group immune serum type HPS 14 TbpA with50?g each;C1,C2 group injected 200?L PBS each as challenge control group;A3,B3,C3 group injected 200?L PBS each as control.Using Freund's adjuvant,subcutaneous multi-point injection,immune two times,second times the amount of immune 100?g each,immune interval of 20 days.10 days after immunization,Cardiac blood sampling and Isolated serum.measured the levels of antibodies and cytokines?IL-2,IL-5,IL-8,IFN-?,TNF-?,MCP-1?.In 21 days after the second immunization,A1,B1 and C1 were challenged with 1×LD₁₀₀00 HPS serum type 13.A2,B2,C2 were challenged with 1×LD₁₀₀00 HPS serum type 14.A1,B1,C1 injected PBS,statistical immune protection,necropsy of guinea pigs making pathological sections and observed pathological changes.The results showed that guinea pig has immuned HPS serum type 13,14 TbpA,both antibody levels in serum increased significantly,but no significant differences between them.In addition to MCP-1 and IL-2,the others cytokine levels were significantly increased after the two immunization,and the difference significantly between the two immunization,but it is no significant difference between serum type 13 and type 14 TbpA group;Serum type 13 TbpA immune group challenge protection on the same serotype and different serotypes of HPS were 83.3%and 50%;serum type 14 TbpA immune group challenge protection on the same serotype and different serotypes of HPS were 66.7%and 33.3%;immunity group survival in guinea pigs and blank control groups of guinea pigs were no significant gross lesions and the pathological changes of the lungs of guinea pigs,died of hemorrhage and necrosis of the liver surface,fibrinous exudation,pleural effusion,pleural adhesions,cerebral hemorrhage,lung,spleen and liver in a small amount of inflammatory cell infiltration or minor bleeding;controlgroup of guinea pig lung hemorrhage,necrosis,liver surface of cellulose effusion,pleural adhesions,pleural effusion,cerebral hemorrhage,lung,spleen,liver,inflammatory cell infiltration,hemorrhage and severe.The results show that the HPS serum type 13,type 14 TbpA protein to the same type and different types of HPS interaction provide protective immunity;serotype 13TbpA with immune protection against serotype HPS was higher than that of serotype14,serotype 13,14 type TbpA immune protection against different serotypes of HPS rate the same.