Development And Immune Effect Evaluation Of A Triplex Inactivated Vaccine Against Pasteurella Multocida,Mannheimia Haemolytica And Escherichia Coli In Goats

In recent years,the dairy goat industry has developed rapidly.With the expansion of breeding scale,respiratory diseases have become one of the most harmful diseases in dairy goat breeding.Pasteurella multocida,Mannheimia haemolytica and Escherichia coli are all pathogenic bacteria of respiratory diseases in goats.The three are often mixed infection,which makes it difficult to take targeted drugs and treat.Therefore,immunization should be taken as the main measure for prevention.At present,there is no vaccine for goats against such diseases,which has caused huge economic losses to the goat industry in Guanzhong area.In order to reduce the great harm caused by respiratory diseases,it is planned to develop a triplex inactivated vaccine against Pasteurella multocida,Mannheimia haemolytica and Escherichia coli,which aims to have a good preventive effect on the local epidemic pathogenic strains.In this study,the pathogenic bacteria were isolated from the diseased lung tissue of Guanzhong dairy goats.They were identified by staining microscopy,biochemical test and16S r RNA sequencing.Their drug resistance and pathogenicity were studied by drug sensitivity test and LD₅₀test.The three bacterial solutions were inactivated with a final concentration of 0.3%formaldehyde solution and then concentrated and purified,and two triplex inactivated vaccines were prepared by adding Freund’s adjuvant and aluminum hydroxide adjuvant respectively.After the character test and safety test,mice and goats were injected,the serum antibody titer was detected by MAT,and the protective effect of the vaccine on the body was observed through immune protection test.The test results are as follows:1.7 strains of Pasteurella multocida were isolated from 12 lung disease materials of Guanzhong dairy goats,and the isolation rate was 58.3%.2 strains of Mannheimia haemolytica were isolated,and the isolation rate was 16.7%.6 strains of Escherichia coli were isolated,and the isolation rate was 50.0%.Pasteurella multocida virulent strain PM01,Mannheimia haemolytica virulent strain MH01 and Escherichia coli virulent strain EC01were selected for vaccine preparation.In the drug susceptibility test,PM01 was resistant to3 of the 14 drugs,MH01 was sensitive to all drugs,EC01 was more resistant,and 8 of 14drugs were resistant.In the LD₅₀test in mice,the LD₅₀of PM01 was 6.339×10⁴CFU,the LD₅₀of MH01 was 6.325×10⁸CFU,and the LD₅₀of EC01 was 9.290×10⁷CFU.All of the isolated strains can cause lung lesions in mice.Mice are susceptible to Pasteurella multocida,but less susceptible to Mannheimia haemolytica.2.After concentrating the three antigens to 2.0×10¹⁰CFU/m L,the triplex inactivated vaccine against Pasteurella multocida,Mannheimia haemolytica and Escherichia coli with two adjuvants was prepared.The vaccine test is qualified.After injection,the serum antibody titers of mice and goats were detected.It was found that both vaccines can stimulate the body to produce immune response.The change of serum antibody titer after the first immunization is not obvious,and the serum antibody titer is significantly improved after enhanced immunization.The antibody level produced after Freund’s adjuvant vaccine injection is higher than that produced after aluminum hydroxide adjuvant vaccine injection.In the immune protection test of mice,after injection at a dose of 5 times LD₅₀,the protection rate of Freund’s adjuvant vaccine against Pasteurella multocida was 100%,the protection rate against Mannheimia haemolytica was 40%,and the protection against Escherichia coli was 80%.The protection rate of aluminum hydroxide adjuvant vaccine against Pasteurella multocida was 80%,the protection rate against Mannheimia haemolytica was 40%,and the protection against Escherichia coli was 60%.The autopsy and histomorphological observation of lung,liver and spleen showed that the vaccine had a certain protective effect on the internal organs of mice.This is the first study on the triplex inactivated vaccine against Pasteurella multocida,Mannheimia haemolytica and Escherichia coli in China.After evaluation,the vaccine has good immune effect,and can effectively prevent respiratory diseases.It has the value of popularization and application,and helps to solve the problem that there is no suitable vaccine for respiratory diseases in dairy goat breeding industry.