Porcine Japanese encephalitis virus and porcine parvovirus dual inactivated vaccine and preparation method thereof

Abstract

The invention provides a preparation method of a binary inactivated vaccine against a Japanese encephalitis virus and a porcine parvovirus, which comprises the following steps of: (1) inoculating the animal cells to a microcarrier for performing the absorption culture and multiplication culture processes of the cells in a tidal biological reactor; (2) inoculating the Japanese encephalitis virus and the porcine parvovirus on the cells being subjected to multiplication culture; (3) when the inoculated animal cell CPE (Cytopathic Effect) reaches above 75%, harvesting a viral solution; and (4) freezing and thawing the harvested viral solution twice at minus 20 DEG C, inactivating after the concentration, and adding an emulsifier for emulsification to obtain the binary inactivated vaccine against Japanese encephalitis virus and porcine parvovirus. The binary inactivated vaccine provided by the invention has the advantages of good immunogenicity, stable growth and high titer of the preserved seed virus; and the tidal biological reactor is adopted to respectively culture and harvest two viruses with high titers, and the two viruses are directly emulsified and prepared after the concentration and the inactivation, so that the binary inactivated vaccine can be continuously produced in a large scale.

Description

technical field [0001] The invention relates to a porcine dual inactivated vaccine, in particular to a dual inactivated vaccine for treating and preventing porcine Japanese encephalitis and porcine parvovirus infection, belonging to the field of veterinary vaccines. Background technique [0002] Porcine encephalitis virus (Swine Japanese Encephalitis Virus, JEV) and porcine parvovirus (Porcine Parvovirus Virus, PPV) are one of the important pathogens that cause reproductive disorders in pigs, mainly manifested as abortion, infertility, stillbirth and mummification in sows Fetal and weak piglets, boar testicular acute inflammation or infertility, so that the pig industry has suffered huge losses. Except for single infection, JEV and PPV are often co-infected. The survey of mixed infection in large-scale pig farms in Sichuan, Chongqing, Hubei, and Henan found that the positive rate of mixed infection was ≥ 60%. [0003] In "Porcine Parvovirus Infection-Japanese Encephalitis D...

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Problems solved by technology

However, there are some disadvantages common to the traditional spinner bottle culture in this technical scheme of spinner bottle culture: (1) spinner bottle cell culture cannot adjust the culture conditions such as nutrient, pH and dissolved oxygen of the culture medium in real time, so it cannot Ensure that the cells are in the best culture state; (2) spinner bottle cell culture, the operation procedures are cumbersome, there are many exposure points with pollution risks, and the production process is difficult to scale up; (3) spinner bottle cell culture, the batch-to-batch variation is large, and the production quality is difficult to control , the product quality is unstable

[0005] The Chinese patents "Method for industrial production of porcine Japanese encephalitis vaccine using bioreactor" (CN 102038943A) and "Method for industrial production of porcine parvovirus vaccine using bioreactor" (CN 102038945A) use microcarrier bioreactors to cultivate pigs Japanese encephalitis virus and porcine parvovirus have further improved the scale of production, but because most of the subcultured cells used to proliferate viruses, such as BHK-21 or ST cells, are all animal cells, the outer layer of the cell is a plasma membrane, Highly brittle, so shear stress must be minimized in the reactor

However, the virus culture method in the above-mentioned patent uses a stirred bioreactor. The high shear force generated by this stirred culture method will lead to a high cell loss rate, which in turn affects the production efficiency and cell yield of the bioreactor. In particular, unnecessary cell fragments are formed, which can easily cause allergic reactions in animals and affect the quality of vaccines

[0006]Therefore, due to the above-mentioned difficulties, there is currently no production and sales of dual inactivated vaccines for porcine encephalitis virus and porcine parvovirus on the market to prevent these two For diseases caused by viruses, JEV and PPV single vaccines need to be vaccinated separately, the immunization procedure is complicated and the cost is high

Moreover, because the diseases caused by the two viruses often occur at the same time, the immune effect of a single vaccine is not satisfactory.

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