CN115837073A - Preparation method and application of porcine Serneca valley virus inactivated vaccine - Google Patents

Abstract

The invention discloses a preparation method and application of a swine Seneca valley virus disease inactivated vaccine, wherein the vaccine adopts suspension culture of Seneca valley virus, a chromatography method for purifying antigen and an optimized water adjuvant for preparing the vaccine.

Description

Preparation method and application of porcine Serneca valley virus inactivated vaccine

Technical Field

The invention belongs to the field of biological products for livestock, and particularly relates to a preparation method and application of a swine Seneca valley virus disease inactivated vaccine.

Background

SVV is a newly discovered acute pathogenic microorganism which can cause Swine Idiopathic Vesicular Disease (SIVD) and Epidemic Transient Neonatal Loss (ETNL), and has the advantages of high morbidity, fast disease course, fast propagation speed, low mortality rate and high piglet mortality rate. Clinically, the symptoms of blister, ulceration, lameness, lethargy, diarrhea, acute death and the like of the pig rhinoscope and the coronary area of the hooves are mainly shown. In 2015, the first in-plug valley epidemic situation occurs in Guangdong areas in China, in 2016 and 2017, the in-plug valley epidemic situation continues to spread, outbreaks occur in multiple provinces and markets in China, the epidemic range reaches provinces such as Guangdong, fujian, hunan, henan and Heilongjiang, clinical symptoms are characterized by blister lesions, and occasionally, a large amount of piglets die in low days, so that huge economic loss is brought to the pig raising industry, and the public health safety and the breeding industry development in China are seriously threatened.

Currently, no biological vaccine preparation is available for the prevention and control of the disease. At present, the prevention and control of the swine Sernicke disease depend on the effective management of a pig farm, but the domestic pig raising mode is complex and various, and the management of the pig farm is laggard, so that the development of corresponding biological products for preventing and controlling the outbreak of the epidemic disease is urgent.

Disclosure of Invention

Technical problem to be solved

The invention aims to provide an effective and safe mass preparation method of a seneca valley virus vaccine aiming at the defects of the existing seneca valley virus biological product; the preparation method adopted by the inactivated vaccine is simple and safe, and the vaccine has a good immune effect.

The technical scheme is as follows:

the invention is realized by the following technical scheme:

the invention provides a preparation method of a swine Saikaca valley virus disease inactivated vaccine, which comprises the following specific steps:

1) The specific preparation process of the porcine Seneca valley virus antigen by virus liquidThe following were used: preparing suspended BHK-21 cells by using a bioreactor until the cell density is 3.0-5.0 multiplied by 10 ⁶ cells/ml, inoculating the Sainika virus seeds with MOI of 0.001-0.1, setting the control parameters of a bioreactor after inoculation to be DO of 30-60%, pH7.0-7.4, stirring speed of 80-100 rpm/min, temperature of 36.5-37.5 ℃, sampling periodically during the period, observing the pathological condition of cells by 0.4% trypan blue staining, and harvesting suspension culture virus liquid when the cell viability is less than or equal to 20%.

2) Antigen purification the suspension culture virus liquid obtained above is subjected to repeated freeze thawing for 3 times, a continuous flow centrifugation method is adopted, the virus liquid is centrifuged for 30min at 8000rpm, cell debris precipitate is discarded, and centrifugation supernatant is collected. The supernatant was washed and filtered repeatedly 3 times through a 500KD hollow fiber column, and the crude and pure virus solution was collected. And purifying by molecular sieve chromatography gel to obtain purified virus liquid of the seneca millet.

3) The specific inactivation process is as follows: adding the sterilized and filtered BEI solution into the purified virus solution of the pig Seneca millet according to the volume of 2 percent to ensure that the final concentration of BEI is 0.08 percent, fully and uniformly mixing, pouring the mixture into a tank, timing when the temperature is raised to 30 ℃, inactivating the BEI for 36 hours, and stirring the mixture for 1 time every 120 minutes. And after timing is finished, adding the filtered and sterilized 50% sodium thiosulfate solution into the inactivated virus solution to ensure that the final concentration of the inactivated virus solution is 2%, fully and uniformly mixing, and standing the inactivated antigen at 2-8 ℃ for storage.

Vaccine formulation

Slowly adding the inactivated suinicarava valley antigen into the polymer water-based adjuvant, wherein the amount of the polymer water-based adjuvant is 50% of the mass of the antigen, controlling the stirring speed to be 2000 r/m, controlling the stirring temperature to be below 25 ℃, and performing subpackage marking after the antigen and the adjuvant are uniformly dispersed, thereby obtaining the suinicarava valley adjuvant inactivated vaccine.

Preparation of adjuvants of the invention

The polymer water-based adjuvant comprises the following components in percentage by mass:

preferably, the polymer water-based adjuvant consists of the following components in percentage by mass:

weighing the components according to the proportion, sequentially adding the components into a quantitative water solution for injection, controlling the stirring speed, mixing at the temperature of below 60 ℃, uniformly mixing to obtain a liquid, sterilizing at 121 ℃ for 30 minutes, and cooling to room temperature to obtain the aqueous adjuvant suitable for the swine seneca valley inactivated vaccine.

(III) advantageous effects

1) The swine Seneca valley virus inactivated vaccine prepared by the invention is different from the prior vaccine preparation method, and the antigen prepared by the suspension culture method has high yield and good uniformity, and saves manpower; the virus production virus content is greatly improved, the antigen harvesting time is shortened, and the antigen production cost is saved through optimization of the virus receiving dose, the virus receiving mode and the reactor control parameters. The optimal virus inoculation amount is MOI 0.01-0.05 through optimization parameters, the virus inoculation process is low-rotation speed (60 rpm/min) incubation for 2h at 37 ℃, the virus harvesting time is about 24h, and the virus content is more than or equal to 10 ^9.5 TCID ₅₀ /ml。

2) The invention also optimizes the component combination of the adjuvant, the adjuvant is a polymer water-based adjuvant, the biocompatibility is good, and a large number of screening tests show that the combination of the polyethylene, the propylene glycol and the polyglutamic acid has the characteristics of strong water solubility, biocompatibility, biodegradability, no toxicity and the like, the targeting property of the medicament can be improved, the medicinal effect can be effectively improved, and the performance of the medicament or material can be improved. Moreover, experiments show that the combination of the three components produces a synergistic effect, so that the immune antibody level of the swine cenecarbalt vaccine can be obviously improved, and the vaccine has good stability and safety.

3) The inactivated vaccine has good safety and effectiveness, and can induce animals to generate stronger neutralizing antibodies after immunization. Because no biological products related to the vaccine of the pathogen exist in the markets at home and abroad, the vaccine can effectively prevent the prevalence and spread of SVV in swinery, reduce the economic loss caused by the disease and has wide application prospect.

Detailed Description

The methods in the following examples are conventional methods unless otherwise specified.

The following examples are intended to illustrate the invention but are not intended to limit the scope of the invention. Unless otherwise specified, the technical means used in the examples are conventional means well known to those skilled in the art, and the raw materials used are commercially available products.

Example 1 preparation of inactivated antigen of porcine Seneca valley Virus

1. Preparation of swine seneca valley virus liquid

1. BHK-21 suspension cell resuscitation

The frozen BHK-21 suspension cell strain (BHK-21-BP-2, preservation number is CGMCC NO.17588, cell strain BHK-21-BP-2 can refer to CN 110157659A) is placed in a water bath at 37 ℃ for quick thawing. Adding fresh special culture medium for suspension culture into the thawed cells at initial density of 0.5-0.8 × 10 ⁶ cells/ml were inoculated into a cell flask, 2% newborn bovine serum was added, and the mixture was cultured at 37 ℃ and 120rpm/min in a constant temperature shaker. Samples were taken at regular times and stained with 0.4% trypan blue, and cell viability and cell doubling rate were observed under the microscope. And (4) carrying out normal subculture when the cell culture reaches the normal multiplication rate and the cell survival rate reaches more than 95%.

2. Cell passage

The cell density reaches 3 to 5 multiplied by 10 ⁶ cells/ml, cell viability up to more than 95%. Diluting and passing through suspension culture medium to make cell final concentration be 0.5-0.8X 10 ⁶ cells/ml, placed at 37 ℃,120rpm/min constant temperature oscillator for cell seed expansion.

3. Step-by-step amplification culture of BHK-21 suspension cells in bioreactor

1) Suspension culture of BHK-21 suspension cells in 10L bioreactor

Enlarging cell seeds in a shake flask, inoculating the seeds into a 10L bioreactor, wherein the initial density of the cells is 0.5-0.8 multiplied by 10 ⁶ cell/ml, setting the technological parameters of the reactor at 36.5-37.5 deg.c, pH 7.0-7.2, DO30-60%, stirring at 80-100 rpm, and timing sampling to observe cell activity rate and cell density. When the cell density reaches 3-5 multiplied by 10 ⁶ cells/ml, when the cell viability reached 95% or more, the culture was further expanded.

2) Continuous amplification culture of BHK-21 suspension cells in 10L-50L-100L bioreactor

Transferring and inoculating BHK-21 suspension cells cultured by a 10L bioreactor into a 50L bioreactor, wherein the initial density of the cells is 0.5-0.8 multiplied by 10 ⁶ cell/ml, setting the technological parameters of the reactor at 36.5-37.5 deg.c, pH 7.0-7.2, DO30-60%, stirring at 80-100 rpm, and timing sampling to observe cell activity rate and cell density. When the cell density reaches 3-5 multiplied by 10 ⁶ cells/ml, when the cell viability reaches more than 95%, the culture is further expanded. The cells were cultured in a 100L bioreactor by the same culture process.

3) Production of a 100L reactor for porcine Seneca Valley Virus strains

Adopting 100L of suspension culture cells prepared above, and performing virus inoculation incubation and virus culture until a porcine seneca valley virus suspension culture virus liquid is obtained:

when the cell density reaches 3-5 multiplied by 10 ⁶ cells/ml, when the survival rate reaches more than 95%, inoculating seneca valley virus seed virus (SVV-ZM-201801, the preservation number is CGMCC NO.16396, and the preservation number is CN 109554352A) with MOI 0.001-0.1, incubating the virus seed at 37 ℃ at 60rpm/min for 2h, setting the technological parameters of the reactor at 36.5-37.5 ℃, pH7.0-7.2, DO 30-60%, stirring at 80-100 rpm, and sampling at fixed time to observe the cell survival rate and the cell density. Culturing until the cell survival rate is less than or equal to 20%, harvesting virus liquid, sampling and detecting the virus titer. The virus content is more than or equal to 10 ^9.5 TCID ₅₀ /ml。

2. Purification of antigens

1. Viral fluid cell debris treatment

After 3 times of repeated low-temperature freeze thawing, the suspension culture virus liquid is harvested, a continuous flow centrifugation method is adopted, the virus liquid is centrifuged for 30min at 8000rpm, cell debris precipitate is discarded, and centrifugation supernatant is collected.

2. Crude and pure virus liquid

The virus supernatant is washed and filtered repeatedly for 3 times by 0.01M phosphate buffer solution through a hollow fiber column with 300-500 KD, and the crude pure virus solution is collected.

3. Molecular sieve chromatography purification

And (3) purifying the crude pure virus solution by a 4FF molecular sieve (GE company) and then by molecular sieve chromatography gel, collecting the first peak appearing in the feed liquid, stopping collecting when the light absorption value of a light absorption value curve is reduced to the lowest part, measuring the Protein removal rate of a collected peak sample by using a lowry Protein Assay Kit (Thermo company), and harvesting the purified virus solution of the Seikagaku.

3. Inactivation of antigens

1. Adding the sterilized and filtered BEI solution into the swine Seikaga valley virus solution according to the volume of 2% to ensure that the final concentration of BEI is 0.08%, fully and uniformly mixing, pouring into a tank, timing after the temperature is increased to 30 ℃, inactivating for 36 hours, and stirring for 1 time every 120 minutes.

2. And after timing is finished, adding the filtered and sterilized 50% sodium thiosulfate solution into the inactivated virus solution to ensure that the final concentration of the inactivated virus solution is 2%, fully and uniformly mixing, and standing the inactivated antigen at 2-8 ℃ for storage.

Example 2 optimization of Swine Sernica Valley Virus suspension culture conditions

Aiming at the 100L virus culture process, important process parameters are further optimized, and the influence of the virus inoculation process and the virus inoculation dose on the virus production effect is compared:

1. optimization of virus inoculation process

100L suspension culture of cells until the cell density reaches 3-5 multiplied by 10 ⁶ cells/ml, when the survival rate reaches more than 95%, inoculating the seed virus of the Seneca valley virus, performing inoculation culture by adopting the processes of direct inoculation culture and incubation inoculation culture (37 ℃,60rpm/min,2 h), and comparing the advantages and disadvantages of the two modes for harvesting the virus. The results are given in the following table:

according to the test comparison result, the optimized virus inoculation process can effectively shorten the virus harvesting time and the virus content.

2. Dose optimization of inoculants

100L suspension culture of cells until the cell density reaches 3-5 multiplied by 10 ⁶ cells/ml, when the survival rate reaches more than 95%, inoculating the seed virus of the Selenka valley virus, comparing the antigen production effects of several batches of different inoculation dosages MOI (0.001, 0.01, 0.05 and 0.1) of the seed virus, and the results are as follows:

dose for receiving poison (MOI)	Harvesting of viral lesions time (h)	Viral content (TCID) 50 /ml)
			0.001	35	10 9.0
0.01	30	10 9.8
			0.05	26	10 10.5
0.1	16	10 9.6

According to the test comparison result, the quantity of the virus-receiving agent has obvious influence on the receiving time of the virus production and the virus content, when the quantity of the virus-receiving agent is controlled within the range of 0.01-0.05, the virus harvesting time can be shortened, and the virus content is further improved to 10 ^9.8 TCID ₅₀ More than ml, is beneficial to shortening the time cost and the antigen titer of the regular production.

Example 3 preparation of aqueous adjuvant for inactivated Swine Sernica Valley vaccine

According to the invention, a large number of screening tests show that the combination of polyethylene, propylene glycol and polyglutamic acid can remarkably improve the immune antibody level of the swine seneca valley vaccine, and the vaccine has good stability and safety.

To verify the effect of the adjuvant of the present invention, the following adjuvants were prepared:

polymer water adjuvant combination 1:

according to the technical scheme provided by the invention, the water adjuvant for preparing the polymer comprises the following components in percentage by mass:

82% of an aqueous solution for injection;

polymer combination:

auxiliary agent:

2% of triglycerine/decanoate;

1, 2-propylene glycol 2%.

Polymer water adjuvant combination 2:

according to the technical scheme provided by the invention, the water adjuvant for preparing the polymer comprises the following components in percentage by mass:

84% of an aqueous solution for injection;

polymer combination:

auxiliary agent:

2% of triglycerine/decanoate;

1, 2-propylene glycol 2%.

Polymer water adjuvant combination 3:

according to the technical scheme provided by the invention, the water adjuvant for preparing the polymer comprises the following components in percentage by mass:

84.8 percent of water solution for injection;

polymer combination:

auxiliary agent

1% of triglycerine/decanoate;

5% of 1, 2-propylene glycol;

polymer Water adjuvant combination 4

According to the technical scheme provided by the invention, the water adjuvant for preparing the polymer comprises the following components in percentage by mass:

81% of an aqueous solution for injection;

polymer combination:

auxiliary agent

5% of triglycerol octanes/decanoate;

1, 2-propylene glycol;

weighing the polymer and the auxiliary agent combination according to a proportion, sequentially adding the polymer and the auxiliary agent combination into a quantitative water solution for injection, controlling the stirring speed, mixing, controlling the temperature below 60 ℃, uniformly mixing to obtain a liquid, sterilizing at 121 ℃ for 30 minutes, and cooling to room temperature to obtain the aqueous adjuvant suitable for the swine seneca valley inactivated vaccine.

Comparative example 1:

84% of an aqueous solution for injection;

polymer combination:

3% of glucan;

600% of polypropylene glycol;

8000 percent of polyethylene glycol;

3% of auxiliary agent triglycercine/decanoate;

2% of 1, 2-propylene glycol;

comparative example 2:

90% of water solution for injection;

polymer combination:

3% of glucan;

8000 percent of polyethylene glycol;

3% of auxiliary agent triglycerol octyl/decanoate;

2% of 1, 2-propylene glycol;

comparative example 3:

86% of an aqueous solution for injection;

polymer combination:

3% of glucan;

600% of polypropylene glycol;

3% of auxiliary agent triglycercine/decanoate;

2% of 1, 2-propylene glycol;

comparative example 4

According to the technical scheme provided by the invention, the water adjuvant for preparing the polymer comprises the following components in percentage by mass:

80% of water solution for injection;

polymer combination:

5% of glucan;

600% of polypropylene glycol;

8000% of polyethylene glycol;

auxiliary agent

2% of 1, 2-propylene glycol;

example 3 vaccine preparation

The inactivated suinica valley virus antigen obtained in the embodiment 1 is respectively and slowly added into the polymer water adjuvant described in any one of the polymer water adjuvant combinations 1-4 in the embodiment 2, the amount of the polymer water adjuvant is 50% of the mass of the antigen, the stirring speed is controlled to be 2000 rpm, the stirring temperature is controlled to be below 25 ℃, and after the antigen and the adjuvant are uniformly dispersed, the split charging and marking are carried out, so that the corresponding suinica valley adjuvant inactivated vaccine 1-4 is obtained. The inactivated vaccines of comparative examples 1 to 4 were prepared as adjuvants in the same manner to obtain comparative examples 1 to 4.

Example 4 inactivated vaccine safety test

1-4 parts of the swine Sernica valley water adjuvant inactivated vaccine prepared in the embodiment 3 and 1-4 parts of the comparative example vaccine are used for carrying out antigen-antibody negative screening on 4-week-old piglets, and 10 piglets negative to the antigen-antibody are selected for each group to carry out a safety evaluation test of the Sernica valley virus inactivated vaccine. Grouping condition: as shown in table 1, 10 heads of the polymer water adjuvant combination 1-4 were randomly selected to inoculate the overdose (4 ml/head) of inactivated vaccine 1-4, respectively; comparative examples 1-4 10 random vaccinations of the overdose (4 ml/vaccine) of the comparative vaccines 1-4 and 2 vaccinations of the control group were used as negative controls. Measuring the body temperature every day from 0 to 7 days of immunization; continuously observing the feeding condition of the swinery for 14 days; and observing and touching whether the local part of the injection part has the tumor or not.

TABLE 1 monitoring table for body temperature after inactivated vaccine immunization

The results are shown in table 1, no obvious body temperature rise phenomenon occurs after the inactivated vaccine is used for immunizing piglets, and the piglets are fed normally and have good mental state; the vaccine injection part has no pain feeling and no lump after 5 days of immunization, which indicates that the vaccine has good safety to pigs.

Example 5 monitoring of neutralizing antibody levels after Immunization of inactivated vaccines

In order to evaluate the immune effect of the inactivated vaccine, 43 pigs are selected for an immune test, 25 immunized pigs are respectively subjected to 1-4 inactivated vaccines (groups 1-4) and 1-4 comparative examples, and neck muscles are injected with 2ml of the inactivated vaccine per head; 3 control pigs were not vaccinated as a blank control, a second immunization was performed 21 days after the first immunization, sera were collected at 0d, 7d, 14d, 21d, 28d and 35d after the immunization, and the level of anti-porcine Seikaga virus antibody production in the sera was evaluated using a neutralization test.

The method comprises the following specific steps:

1) Putting all the pig serum to be detected in a 56 ℃ water bath for incubation for 30min, and performing inactivation treatment;

2) Each serum was subjected to 1: 8. 82301, 2048 fold dilution, 4 replicates per dilution, 100 μ L of diluted serum per well, using the SVV neutralizing antibody assay.

3) Diluting the virus solution to 200TCID with cell maintenance solution ₅₀ 0.1mL, the diluted virus solution was added to the serum diluted in the above step at 100ul per well, mixed well, and subjected to 37 ℃ at 5% CO ₂ Incubate in incubator for 1h.

4) BHK21 cells with good growth status were passaged according to 1. And simultaneously setting a positive control group and a negative control group of the virus liquid and the virus liquid which is not added. At 37 ℃ and 5% CO ₂ Culturing in an incubator, observing the pathological condition of cells every day, recording the pathological condition, and calculating the serum neutralizing antibody condition of each group according to a Reed-Muench method. The data were collated after determination of the serum neutralizing antibody titres and the results are shown in table 2.

TABLE 2 level of neutralizing antibodies produced by animals immunized with inactivated vaccine

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As can be seen from Table 2, the content of polypropylene glycol in the adjuvant is different, and whether polyglutamic acid and decanoate are added or not affects the level of antibodies generated after the inactivated vaccine induces piglet immunity. The highest antibody was obtained with the polymer water adjuvant combination 2 at 6% polypropylene glycol content, and the level of antibody production was reduced in the absence of polyglutamic acid and decanoate, see comparative examples 1-4. However, in summary, all vaccines 1-4 achieved vaccine protective antibody levels of 1. Whereas the control group was not vaccinated and no neutralizing antibodies against porcine seneca valley virus were detected. The data lays a certain theoretical and practical foundation for the clinical use of the porcine Seneca valley virus inactivated vaccine.

Example 6 protection test against virulent challenge

The inactivated vaccine group 2 prepared in example 3 was selected to immunize piglets, and 14 days after the second immunization, the immunization dose was 2ml, and all pigs were challenged with GD/01 strain of Sernica valley virus, 5ml per nose drop, and 2.5ml in each of the left and right nostrils, and the results are shown in Table 3.

TABLE 3 protective results of potent and offensive poison

Group of	Quantity (head)	Number of onset of disease
			Immunization group	5	0/5
Control group for counteracting toxic pathogen	3	3/3

As can be seen from Table 3, the virus challenge control group has 3/3 of diseases, the immune group has no disease and 100% of protection, and the inactivated vaccine for porcine Seneca valley virus can resist the virulent attack of Seneca valley virus.

Although the present invention has been described with reference to the preferred embodiments, it should be understood that various changes, substitutions and alterations can be made herein without departing from the spirit and scope of the invention as defined by the appended claims.

Claims (8)

1. A preparation method of an inactivated vaccine against porcine Serneca valley virus disease comprises the following steps:

1) The specific preparation process of the swine Seneca valley virus antigen by virus liquid is as follows: preparing suspended BHK-21 cell strain with bioreactor until cell density is 3.0-5.0 × 10 ⁶ cells/ml, inoculating the seed virus of the Seneca millet with the dose of 0.001-0.1 MOI, setting the control parameters of a bioreactor after inoculation to be DO 30-60%, pH 7.0-7.4, stirring speed of 50-100 rpm/min, temperature of 36.5-37.5 ℃, sampling at regular time during the period, observing the pathological condition of cells by 0.4% trypan blue staining, and harvesting suspension culture virus liquid when the cell survival rate is less than or equal to 20%;

2) Antigen purification: after the suspension culture virus liquid is obtained and repeatedly frozen and thawed for 3 times, a continuous flow centrifugation method is adopted, the virus liquid is centrifuged for 30min at 8000rpm, cell debris precipitate is discarded, and centrifugation supernatant is collected; repeatedly washing and filtering the supernatant for 3 times by using a 500KD hollow fiber column, and collecting crude pure virus liquid; purifying by molecular sieve chromatography gel to obtain purified virus liquid of seneca millet;

3) Virus inactivation: adding the sterilized and filtered BEI solution into the purified virus solution of the porcine cenaca valley according to the volume of 2 percent to ensure that the final concentration of BEI is 0.08 percent, fully and uniformly mixing, pouring into a tank, timing after the temperature is increased to 30 ℃, inactivating for 36 hours, and stirring for 1 time every 120 minutes; after timing is finished, adding a 50% sodium thiosulfate solution for filtration sterilization into the inactivated virus solution to ensure that the final concentration of the inactivated virus solution is 2%, fully and uniformly mixing, and storing the inactivated antigen at 2-8 ℃;

4) Vaccine formulation

Slowly adding the inactivated suinica valley antigen into a polymer water-based adjuvant, wherein the amount of the polymer water-based adjuvant is 50% of the mass of the antigen, controlling the stirring speed to be 2000 r/m, controlling the stirring temperature to be below 25 ℃, and performing subpackage marking after the antigen and the adjuvant are uniformly dispersed to obtain the suinica valley adjuvant inactivated vaccine.

2. The method of claim 1, wherein: in the step 1), the inoculation amount of the virus is 0.01-0.05MOI, the temperature is 37 ℃, the rotating speed is 60rpm/min, and the incubation is carried out for 2h.

3. The method of claim 1, wherein: the suspension BHK-21 cell strain is BHK-21-BP-2; the Sernica valley virus is Sernica valley virus strain SVV-ZM-201801

The polymer water-based adjuvant comprises the following components in percentage by mass:

the balance of water for injection.

4. The method of claim 3, wherein: the polymer water-based adjuvant comprises the following components in percentage by mass:

the balance of water for injection.

5. The method of claim 4, wherein: the preparation method of the polymer water-based adjuvant comprises the following steps:

weighing the components according to the proportion of the components of the polymer aqueous adjuvant according to the claim 3 or 4, adding the components into a quantitative water solution for injection, controlling the stirring speed, mixing, controlling the temperature below 60 ℃, sterilizing for 30 minutes at 121 ℃ after uniformly mixing to form a liquid, and cooling to room temperature to obtain the aqueous adjuvant suitable for the inactivated vaccine of the porcine cenecarbalt.

6. The porcine zernia valley vaccine prepared by the method of claims 1-5.

7. A polymer water adjuvant suitable for a swine seneca grain vaccine is characterized in that: the polymer water-based adjuvant is prepared from the following components in percentage by mass:

the balance of water for injection.

8. The polymeric aqueous adjuvant according to claim 7, characterized in that: the polymer water-based adjuvant is prepared from the following components in percentage by mass:

the balance of water for injection.