DE1767959A1 - Process for the production of a lyophilized combination vaccine against swine fever and bacterial diseases in pigs - Google Patents

Description

1767959 FARBENFABRIKEN BAYER AG

LEVERKUSEN-myenverk July 4th 1968 Pitent-Abwilunj Si / HM \

Process for the production of a lyophilized combination vaccine against swine fever and bacterial diseases

on pigs

The invention relates to the production of a vaccine, with pigs of any age, but at the earliest one week after weaning, as well as pregnant sows, with off -, __ would take the first and last weeks of gestation the classical swine fever and infections with pathogeneii ^. Bacteria can be immunized as a preventive measure.

It is known that secondary diseases caused by Salmonella and Pasteurella bacteria very frequently occur as a result of an infection of M pigs with the swine fever virus. Likewise, one often finds disease caused by one of these pathogens alone z.ii. Piglet paratyphoid fever or hemorrhagic septicemia. These infections cause very severe disease syndromes that occur in

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in many cases to the death of the infested iüiere after a short time to lead. These diseases are particularly severe in young animals, which are also affected relatively more often, It is therefore largely customary to give piglets a few weeks

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to immunize against these pathogens with vaccines after birth. However, the usual practice so far has in addition to the higher costs the disadvantage that there is a period of time between the individual vaccinations

* must be several weeks for the vaccinations to be

Protective training can lead. With the majority of those so far The usual bacterines (vaccines against bacterial pathogens) do not concentrate the bacteria, so that the vaccination usually has to be repeated after a few weeks. ^ By the protein content of the nutrient broth, which is necessary for the Breeding of the bacteria is required, allergic reactions may occur. The dispatch of these vaccines and theirs Use must take into account the instability of the solutions against higher temperatures at temperature ranges between + 4. and + 8. -

The invention relates to a method for producing a grain ' bination vaccine that eliminates these disadvantages. The vaccine is in lyophilized i'orm, so that it can be stored ™ is achieved by at least 1 1/2 years, with storage temperatures below + 18 ° are sufficient. Occasional short term Exceeding these conditions up to 35 ° does not harm. After dissolving the lyophilized material in the enclosed solvent that is an adjuvant to strengthen immunizing Effect of the bacterial component contains, the antigens are administered in one injection, so that-in leu vaccinee the protection trained against the various pathogens at the same time

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will be. Due to the special manufacturing process, the concentration of the bacterial antigen can be adjusted to the optimum Value can be set,; so that in combination with · the attached Adjuvant with a single injection is a very good one Protective effect is achieved. That to inactivate the bacteria used acetone is completely removed before .Lyophilization. This eliminates after-effects of the inactivating agent, which is a gradual diminution of the immunizing Effect properties. .

The vaccine claimed according to the invention is produced in three steps: 1.) Production of the antigens, 2.) Mixing of the antigens and lyophilization with the addition of a Stabilizer solution, 3 «) Preparation of the solvent with Adjuvant content. -

1.) Serves as an antigen to combat classic swine fever a specially atenuated swine fever virus strain, its production for use as a live vaccine by the Spanish patents.Hr. 547294 and 347295 is protected. The bacterial strains are isolated, especially in disease outbreaks pathogenic germs that are propagated in the usual Uährbouillon and after reaching the optimal bacterial concentration with Acetone can be precipitated at + 4. The bacteria will be up for complete inactivation at + 4 ° in the acetone suspension stirred and then dried.

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2.) Solutions of the swine fever virus obtained in cell cultures C

are, by titration of the protective effect on pigs j

examined for their "virus content and by mixing with a ν

Stabilizer solution that contains carbohydrates and proteins, so, '' diluted, that in every production batch per vaccination dose the fer- * term vaccines contain the same virus content of 800 UMP.

A UMP is the smallest amount of virus that ι when inoculated on pigs

still protects all animals against a test infection with virulent swine pest virus « \

φ In this solution, so much of the dry bacterial powder becomes uniform distributed that in the finished vaccine the desired number j

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of bacteria obtained per vaccination dose. z. B. 20 χ 1O ³ bacteria per vaccination dos is. The ratio of the bacterial species to each other can be set as required, e.g. 60 $ Salmonella choleraesuis and 40 % Pasteurella multocida. The mixture is filled into the usual lyophilization bottles and lyophilized.

3.) Hatriumalginat whose molecular weight was chosen so _Ä that a 3 $ strength by weight solution thereof in distilled water without difficulty in a syringe with needle no. Can be absorbed 18, 3 $ IG together with 0.4 f ° of calcium gluconate in ■ dissolved in distilled water, the pH value adjusted to pH 7.6 by adding buffer solution and sterilized by heating in an autoclave.

The ready-to-use vaccine is prepared in which with a suitable injection syringe the amount of solution added in each case

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agent is added to the lyophilized material. By Shake the substances evenly. Of this suspension, 2 ml subcutaneously or per pig injected intramuscularly. '··' _ ■ *

The following examples are intended to explain the invention in more detail:

The attenuated virus strain of the classical swine fever virus is multiplied according to the method of the Spanish patent No. 34 72 94. The virus content of the solution obtained in this way is determined by titration in the pig. In the 1:10 000 dilution, all 4 pigs that were vaccinated with this dilution are still protected. Based on this result, the virus is diluted with two parts of stabilizer solution. 1.0 gram of dry bacterial powder is added to 5 liters of this suspension. This product is made up of 0.6 gram Salmonella choleraesuis and 0.4 gram Pasteurella multocida. The bacteria are mixed evenly with the virus solution, the mixture is distributed on lyophilization bottles and, after freezing at -40 °, dried in vacuum W. In 40 liters of distilled water, 1,200 grams of sodium alginate and 200 grams of calcium gluconate are dissolved and, by adding 0 , 5 M phosphate buffer adjusted to pH 7.6. the 'mixture is heated for 15 minutes 115 ° in an autoclave. After cooling the solution y is centrifuged off from the precipitate .and the slightly opalescierende supernatant to 10 ml according to the solvent for five Vaccine doses filled with vaccines and sterilized after the bottles have been closed.

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The contents of a bottle with lyophilized vaccine, corresponding to ¹ five vaccine doses, are added. 10 ml of solvent mixed. Of this vaccine, each 2 ml undiluted, 1: 100 and 1: 400 diluted with physiological saline solution are injected intramus - * - into 4 pigs each. Three weeks later, the pigs receive an injection of 2 ml of blood from pigs that had been bled 5 days after infection with a highly pathogenic strain of sinewine fever. The animals vaccinated undiluted and diluted 1: 100 remained completely healthy. The animals vaccinated in a dilution of 1: 400 showed a temperature increase of up to 40.9 without further reactions from the 4th to the 7th day pi.

Another part of the vaccine is 0.2 ml each in 2 × 10 mice injected intraperitonea ^. 21 days later, 10 of the mice received one injection intraperitoneally of 5 x 10 salmonella, the other

10 an injection of 5x10 Pasteurella. With the same number of bacteria 10 mice that were not vaccinated are injected simultaneously per bacterial strain. All non-vaccinated animals died within 3 days with the typical signs of Salmonello se or Pasteurellosis, of the vaccinated animals, 9 of 10 survived the infection with Salmonella and 8 of the infection with Pasteurella.

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Claims (4)

Claims; 1) Process for the preparation of a combination vaccine against Swine fever and bacterial diseases of pigs, dadureh marked that a solution of swine fever virus that has been made suitable for vaccinating pigs by attenuation, is mixed with a stabilizer solution and with a dry powder of inactivated bacteria, so that per vaccine dose of the finished 'vaccine at least 800 UMP Swine fever virus and 20 χ 10 ^ bacteria come that mix is preserved by freeze-drying and is dissolved in a solvent before inoculation, which is an adjuvant to strengthen the immunizing effect. 2) A process for producing a Koabinationsvakzine against swine fever and bacterial diseases of pigs according to claim 1), characterized dadureh that as bacteria Salmonella choleraesuis'und Pasteurella multocida, preferably in the ratio 60 Jt S.choleraesuis and 40 i »comparable P.multocida 'be applied. 3) Process for the production of a coabination vaccine against swine fever and bacterial diseases in pigs according to Claim 1), characterized in that the bacteria are precipitated from the nutrient broth by adding acetone at + 4 ° and digested with acetone until they are infectious Animals and their ability to circulate in culture media disappeared and dried after removing the acetone. le a 11 608 -7- 109840/1651 8AD ORfGJNAL 4) Process for the production of a combination vaccine against swine fever and bacterial diseases in pigs, characterized in that sodium alginate is used as the adjuvant, the molecular weight of which is so high that a 3% solution of it in 0.4 i <> calcium gluconate solution is passed through an injection cartridge No. 18 can be easily vacuumed. Le A 11 608. - 8th BATH ORIGINAL 109840/1651