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Production method of living vaccine of Brucella meliteusis

A Brucella and production method technology, applied in the directions of antibacterial drugs, bacterial antigen components, etc., can solve the problems of low freeze-drying survival rate, difficult storage, transportation and use in remote areas, poor protection function, etc.

Active Publication Date: 2015-08-19
CHINA INST OF VETERINARY DRUG CONTROL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

But there are following outstanding problems: the vaccine adopts 1%~2% gelatin, 5%~10% sucrose, 1%~3% thiourea as protective agent, the prescription is simple, and the freeze-drying survival rate is low (generally 60%~70%) ), the protection function is poor at 37°C for 7 days (the reduction rate of viable bacteria is more than 80%), and it is not suitable for long-term storage at 2-8°C (storage at 2-8°C for 6 months, the reduction rate of viable bacteria is more than 40%)
In addition, Brucella is sensitive to heat and requires a better cold chain system for preservation, which brings difficulties to storage, transportation and use in remote areas

Method used

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  • Production method of living vaccine of Brucella meliteusis
  • Production method of living vaccine of Brucella meliteusis
  • Production method of living vaccine of Brucella meliteusis

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0067] - vaccine preparation

[0068] Brucella suis (CVCC70502 strain) seed liquid is inoculated in the liquid culture medium provided by the present invention by 1%~2% of culture medium total amount, 37 ℃ of fermentations or aerated culture 30~36h, in the cultivation process according to It is necessary to add an appropriate amount of antifoaming agent, and add an appropriate amount of sterilized 40% glucose to control the pH value according to the increase in pH. After the cultivation, centrifuge at 4500r / min for 20min, remove the supernatant, immediately add the sterilized and preheated to 37°C heat-resistant freeze-drying protective agent provided by the present invention, mix well, and dispense according to the regulations. Should pay attention to insulated and shaken evenly in the subpackaging process, and carry out freeze vacuum drying rapidly, press the method that " Chinese Veterinary Pharmacopoeia " stipulates, use freeze-drying curve 2 that the present invention pro...

Embodiment 2

[0070] --Product testing

[0071] Appearance Off-white spongy loose mass, easy to separate from the bottle wall, dissolve quickly after adding diluent.

[0072] The pure test was carried out according to the method stipulated in the "Chinese Veterinary Pharmacopoeia", all of which were pure.

[0073] Sampling for variation test, dilute the vaccine appropriately and inoculate with trypsin On the agar plate, culture at 37°C for at least 72 hours, and check with Brucella colony crystal violet staining method, and the rough colonies do not exceed 5%.

[0074] Viable bacteria counts are marked according to the bottle label, with trypsin Count the live bacteria on the agar plate (according to the method stipulated in the "Chinese Veterinary Pharmacopoeia"), and the number of live bacteria in each head should be greater than 1×10 10 CFU.

[0075] In the safety inspection, the head portion should be marked according to the bottle label, and the vaccine should be filled with tryp...

Embodiment 3

[0080] ——Screening of heat-resistant protective agent formula and freeze-drying curve

[0081] Through the freeze-thaw test and freeze-drying test of 10 kinds of protective agent matrices, the applicable concentrations of various protective agent matrices were preliminarily clarified, ascorbic acid (0.2%, 0.1%), vitamin C (0.2%, 0.1%), trehalose (5 %, 7.5%), BSA (0.5%) had the highest bacterial survival rate in the freeze-thaw test; 1.5% PVP + 5% trehalose + 5% mannitol + 1% thiourea could significantly improve the freeze-dried bacterial storage rate, in addition, 1.5% Adding 0.2% vitamin C, 0.2% ascorbic acid, 0.5% BSA, and 0.5% 199 medium to PVP and 5% trehalose can obviously improve the anti-aging effect of bacteria. Based on this, four heat-resistant protective agent formulas 1, 2, 3 and 4 were designed, mixed with Brucella liquid in equal amounts, and freeze-dried by freeze-drying curves 1 and 2 respectively. The results showed that heat-resistant protective agent formula...

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Abstract

The invention relates to a production method of a living vaccine of Brucella meliteusis. A whole set of technology including a heat-resisting freeze-drying protective additive and a freeze-drying curve adapted to the heat-resisting freeze-drying protective additive is adopted in the production method provided by the invention, so that the survival rate of freeze drying strains of Brucella meliteusis CVCC70502 of pigs is increased, the loss ratio of the freeze drying process is reduced, the anaphylactic reaction caused by strain death is reduced, the storage temperature of a finished product of the vaccine is raised to 2-8 DEG C, the valid period is up to 15-18 months, and relatively-high temperature and short-term storage can be realized, wherein the vaccine is stored at the temperature of 37 DEG C for 7 days, and the reduction rate of living strains is lower than 20%.

Description

technical field [0001] The invention relates to a production method of live Brucella vaccine. It belongs to the field of veterinary biological products. technical background [0002] Brucellosis, referred to as brucellosis, is a zoonotic infectious disease mainly infecting domestic animals caused by bacteria of the genus Brucella. The World Organization for Animal Health (OIE) lists it as a variety of animal diseases, and my country lists it as a second-class animal disease. [0003] In recent years, the brucellosis epidemic situation in my country has been severe, which has caused serious harm to human health and the development of animal husbandry. At present, my country mainly adopts prevention-based epidemic prevention measures. Therefore, the quality of brucellosis vaccine products is directly related to the effective control of the national brucellosis epidemic. [0004] The source of infection related to human brucellosis in my country is mainly sick sheep and catt...

Claims

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Application Information

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IPC IPC(8): A61K39/10A61P31/04
Inventor 朱良全丁家波毛开荣蒋卉宁宜宝王楠
Owner CHINA INST OF VETERINARY DRUG CONTROL
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