Haemophilus parasuis adjuvant vaccine and preparation method thereof

Abstract

The invention belongs to the technical field of vaccine preparation, and particularly relates to a haemophilus parasuis adjuvant vaccine which comprises the following ingredients: serum type 4 haemophilus parasuis liquid, serum type 5 haemophilus parasuis liquid, an alumina gel adjuvant, a composite polysaccharide immunopotentiator, methanol, a solid culture medium and a liquid culture medium. According to the invention, the vaccine stock solution prepared by adding the mixed adjuvant and mixing serum type 4 and type 5 haemophilus parasuis solutions is inactivated and then inoculated to piglets, so that a larger antigen library is formed in the bodies of the piglets, the antigen library is slowly released in the application process, the action time of the antigen is prolonged, and meanwhile, the vaccine has a better promotion effect on the immune effect; the inoculation frequency is reduced, the drug resistance can be alleviated, the cost expenditure of farmers is reduced, and the morbidity and mortality of swinery are reduced.

Description

Technical Field

[0001] The invention belongs to the technical field of vaccine preparation, and particularly relates to a Haemophilus parasuis adjuvant vaccine and a preparation method thereof. Background Art

[0002] Haemophilus parasuis (HPS) is a Gram-negative coccobacillus, an opportunistic pathogen of swine, causing H. parasuis disease. First reported by the German scholar Glasser in 1910, the disease is now widespread worldwide. Currently, infection with H. parasuis is one of the main causes of high piglet mortality in pig farms. The disease currently poses a significant threat to Chinese swine herds, and with the prevalence of porcine reproductive and respiratory syndrome (PRRS, blue ear disease), the toll of H. parasuis disease on swine herds is even more severe than before. H. parasuis infects only pigs and has strong host specificity, affecting pigs aged two to four months, primarily around weaning and in the nursery stage. The incidence rate is generally 10% to 15%, and in severe cases, the mortality rate can reach 50%. In recent years, 15 serotypes of H. parasuis have been identified, with serotypes 4 and 5 being the most prevalent.

[0003] In actual use, the current Haemophilus parasuis vaccine produces low antibodies, has a short duration, and is prone to drug resistance, resulting in poor treatment effects and increasing the cost expenditure of farmers. Summary of the Invention

[0004] In order to solve the problems raised in the above background technology, the present invention provides a Haemophilus parasuis adjuvant vaccine and a preparation method thereof, which has the characteristics of increasing the duration and reducing drug resistance.

[0005] To achieve the above objectives, the present invention provides the following technical solution: an adjuvant vaccine of Haemophilus parasuis, comprising the following ingredients: serotype 4 Haemophilus parasuis liquid, serotype 5 Haemophilus parasuis liquid, aluminum gel adjuvant, complex polysaccharide immunopotentiator, methanol, solid culture medium and liquid culture medium.

[0006] Preferably, the concentration of the serum type 4 Haemophilus parasuis solution is 2.0×10 9 CFU / ml.

[0007] Preferably, the concentration of the serotype 5 Haemophilus parasuis solution is 2.3×10 9 CFU / ml.

[0008] Preferably, the aluminum gel adjuvant is aluminum hydroxide adjuvant.

[0009] Preferably, the complex polysaccharide immunopotentiator is a water-soluble complex of traditional Chinese medicine polysaccharides.

[0010] Preferably, the solid culture medium is TSA culture medium, and the liquid culture medium is TSB culture medium.

[0011] The method for preparing the Haemophilus parasuis adjuvant vaccine comprises the following steps:

[0012] Step 1: prepare a mixed adjuvant by mixing aluminum gel adjuvant and complex polysaccharide immunopotentiator in a ratio of 3:2;

[0013] Step 2: Prepare bacterial antigens by culturing and isolating serotypes 4 and 5 of Haemophilus parasuis in TSA medium, then transferring them to TSB medium for large-scale cultivation. The bacteria are then inactivated by adding 0.3% methanol solution and stored for later use. Inactivate at 35°C for 20 hours, stirring every 4 hours.

[0014] Step 3: Prepare the vaccine stock solution by mixing the serotypes 4 and 5 of Haemophilus parasuis prepared in step 2 in a ratio of 1:1;

[0015] Step 4: Prepare adjuvant vaccine. Use the mixed adjuvant in step 1 and the vaccine stock solution in step 3 to prepare Haemophilus parasuis adjuvant vaccine using conventional methods. The concentration of the mixed adjuvant in the adjuvant vaccine is 3 mg / ml.

[0016] Compared with the prior art, the present invention has the following beneficial effects:

[0017] 1. The present invention prepares a vaccine stock solution by adding a mixed adjuvant and mixing serum types 4 and 5 of Haemophilus parasuis. The vaccine stock solution is inactivated and then inoculated into piglets, thereby forming a large antigen reservoir in the piglets' bodies. The antigen reservoir is slowly released during the application process, thereby prolonging the action time of the antigen and having a better promoting effect on the immune effect. The number of vaccinations is reduced, drug resistance is alleviated, the cost expenditure of farmers is reduced, and the morbidity and mortality of pig herds are reduced. DETAILED DESCRIPTION

[0018] The following is a clear and complete description of the technical solutions in the embodiments of the present invention. Obviously, the embodiments described are only some embodiments of the present invention, not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making any creative efforts are within the scope of protection of the present invention.

[0019] Example 1

[0020] The present invention provides the following technical solution: a Haemophilus parasuis adjuvant vaccine, comprising the following ingredients: serotype 4 Haemophilus parasuis liquid, serotype 5 Haemophilus parasuis liquid, aluminum gel adjuvant, complex polysaccharide immunopotentiator, methanol, solid culture medium and liquid culture medium.

[0021] Specifically, the concentration of Haemophilus parasuis serotype 4 solution was 2.1×10 9 CFU / ml.

[0022] Specifically, the concentration of Haemophilus parasuis serotype 5 solution was 2.3×10 9 CFU / ml.

[0023] Specifically, the aluminum gel adjuvant is aluminum hydroxide adjuvant.

[0024] Specifically, the complex polysaccharide immunopotentiator is a water-soluble complex of traditional Chinese medicine polysaccharides.

[0025] Specifically, the solid culture medium is TSA culture medium, and the liquid culture medium is TSB culture medium.

[0026] A method for preparing a Haemophilus parasuis adjuvant vaccine comprises the following steps:

[0027] Step 1: prepare a mixed adjuvant by mixing aluminum gel adjuvant and complex polysaccharide immunopotentiator at a ratio of 3:1.5;

[0028] Step 2: Prepare bacterial antigens by culturing and isolating serotypes 4 and 5 of Haemophilus parasuis in TSA medium, then transferring them to TSB medium for large-scale cultivation. The cells are then inactivated by adding 0.3% methanol solution and stored for later use. Inactivate at 37°C for 18 hours, stirring every 3 hours.

[0029] Step 3: Prepare the vaccine stock solution by mixing the serotypes 4 and 5 of Haemophilus parasuis prepared in step 2 in a ratio of 1:1;

[0030] Step 4: Prepare adjuvant vaccine. Use the mixed adjuvant in step 1 and the vaccine stock solution in step 3 to prepare Haemophilus parasuis adjuvant vaccine using conventional methods. The concentration of the mixed adjuvant in the adjuvant vaccine is 2.3 mg / ml.

[0031] Example 2

[0032] The present invention provides the following technical solution: a Haemophilus parasuis adjuvant vaccine, comprising the following ingredients: serotype 4 Haemophilus parasuis liquid, serotype 5 Haemophilus parasuis liquid, aluminum gel adjuvant, complex polysaccharide immunopotentiator, methanol, solid culture medium and liquid culture medium.

[0033] Specifically, the concentration of Haemophilus parasuis serotype 4 solution was 2.1×10 9 CFU / ml.

[0034] Specifically, the concentration of Haemophilus parasuis serotype 5 solution was 2.2×10 9 CFU / ml.

[0035] Specifically, the aluminum gel adjuvant is aluminum hydroxide adjuvant.

[0036] Specifically, the complex polysaccharide immunopotentiator is a water-soluble complex of traditional Chinese medicine polysaccharides.

[0037] Specifically, the solid culture medium is TSA culture medium, and the liquid culture medium is TSB culture medium.

[0038] A method for preparing a Haemophilus parasuis adjuvant vaccine comprises the following steps:

[0039] Step 1: prepare a mixed adjuvant by mixing aluminum gel adjuvant and complex polysaccharide immunopotentiator in a ratio of 2:1;

[0040] Step 2: Prepare bacterial antigens by culturing and isolating serotypes 4 and 5 of Haemophilus parasuis in TSA medium, then transferring them to TSB medium for large-scale cultivation. The cells are then inactivated by adding 0.3% methanol solution and stored for later use. Inactivate at 40°C for 15 hours, stirring every 3 hours.

[0041] Step 3: Prepare the vaccine stock solution by mixing the serotypes 4 and 5 of Haemophilus parasuis prepared in step 2 in a ratio of 1:1;

[0042] Step 4: Prepare adjuvant vaccine. Use the mixed adjuvant in step 1 and the vaccine stock solution in step 3 to prepare Haemophilus parasuis adjuvant vaccine using conventional methods. The concentration of the mixed adjuvant in the adjuvant vaccine is 2.8 mg / ml.

[0043] Example 3

[0044] The present invention provides the following technical solution: a Haemophilus parasuis adjuvant vaccine, comprising the following ingredients: serotype 4 Haemophilus parasuis liquid, serotype 5 Haemophilus parasuis liquid, aluminum gel adjuvant, complex polysaccharide immunopotentiator, methanol, solid culture medium and liquid culture medium.

[0045] Specifically, the concentration of Haemophilus parasuis serotype 4 solution was 2.3×10 9 CFU / ml.

[0046] Specifically, the concentration of Haemophilus parasuis serotype 5 solution was 2.1×10 9 CFU / ml.

[0047] Specifically, the aluminum gel adjuvant is aluminum hydroxide adjuvant.

[0048] Specifically, the complex polysaccharide immunopotentiator is a water-soluble complex of traditional Chinese medicine polysaccharides.

[0049] Specifically, the solid culture medium is TSA culture medium, and the liquid culture medium is TSB culture medium.

[0050] Specifically, a method for preparing a Haemophilus parasuis adjuvant vaccine comprises the following steps:

[0051] Step 1: prepare a mixed adjuvant by mixing aluminum gel adjuvant and complex polysaccharide immunopotentiator in a ratio of 2:3;

[0052] Step 2: Prepare bacterial antigens by culturing and isolating serotypes 4 and 5 of Haemophilus parasuis in TSA medium, then transferring them to TSB medium for large-scale cultivation. The cells are then inactivated by adding 0.3% methanol solution and stored for later use. Inactivate at 33°C for 20 hours, stirring every 5 hours.

[0053] Step 3: Prepare the vaccine stock solution by mixing the serotypes 4 and 5 of Haemophilus parasuis prepared in step 2 in a ratio of 1:1;

[0054] Step 4: Prepare adjuvant vaccine. Use the mixed adjuvant in step 1 and the vaccine stock solution in step 3 to prepare Haemophilus parasuis adjuvant vaccine using conventional methods. The concentration of the mixed adjuvant in the adjuvant vaccine is 3.2 mg / ml.

[0055] Finally, it should be noted that the above descriptions are merely preferred embodiments of the present invention and are not intended to limit the present invention. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art will be able to modify the technical solutions described in the aforementioned embodiments or substitute equivalents for some of the technical features. Any modifications, equivalent substitutions, and improvements made within the spirit and principles of the present invention shall be included within the scope of protection of the present invention.

Claims

1. A Haemophilus parasuis adjuvanted vaccine, characterized in that: The invention comprises the following ingredients: serum type 4 Haemophilus parasuis liquid, serum type 5 Haemophilus parasuis liquid, aluminum gel adjuvant, composite polysaccharide immunopotentiator, methanol, solid culture medium and liquid culture medium.

2. The Haemophilus parasuis adjuvanted vaccine according to claim 1, characterized in that: The concentration of the serum type 4 Haemophilus parasuis solution was 2.0×10 9 CFU / ml.

3. The Haemophilus parasuis adjuvanted vaccine according to claim 1, characterized in that: The concentration of the serum type 5 Haemophilus parasuis solution is 2.3×10 9 CFU / ml.

4. The Haemophilus parasuis adjuvanted vaccine according to claim 1, characterized in that: The aluminum gel adjuvant is aluminum hydroxide adjuvant.

5. The Haemophilus parasuis adjuvanted vaccine according to claim 1, characterized in that: The complex polysaccharide immunopotentiator is a water-soluble complex of traditional Chinese medicine polysaccharides.

6. The Haemophilus parasuis adjuvanted vaccine according to claim 1, characterized in that: The solid culture medium is TSA culture medium, and the liquid culture medium is TSB culture medium.

7. The method for preparing a Haemophilus parasuis adjuvant vaccine according to any one of claims 1 to 6, characterized in that: The following steps are involved: Step 1: prepare a mixed adjuvant by mixing aluminum gel adjuvant and complex polysaccharide immunopotentiator in a ratio of 3:2; Step 2: Prepare bacterial antigens by culturing and isolating serotypes 4 and 5 of Haemophilus parasuis in TSA medium, then transferring them to TSB medium for large-scale cultivation. The bacteria are then inactivated by adding 0.3% methanol solution and stored for later use. Inactivate at 35°C for 20 hours, stirring every 4 hours. Step 3: Prepare the vaccine stock solution by mixing the serotypes 4 and 5 of Haemophilus parasuis prepared in step 2 in a ratio of 1:1; Step 4: Prepare adjuvant vaccine. Use the mixed adjuvant in step 1 and the vaccine stock solution in step 3 to prepare Haemophilus parasuis adjuvant vaccine using conventional methods. The concentration of the mixed adjuvant in the adjuvant vaccine is 3 mg / ml.