RU2403061C1 - Inactivated combined vaccine against infectious rhinotracheitis, paraflu-3, respiratory syncytial disease, viral diarrhea and pasteurellosis of cattle - Google Patents

Abstract

FIELD: medicine, veterinary. ^ SUBSTANCE: invention relates to field of veterinary microbiology. As active substance vaccine contains mixture of inactivated virus suspension of strain of virus of infectious rhynotracheitis of cattle, SCV No 2333, of family Herpesviridae, subfamily Alphaherpesviridae, inactivated virus suspension of strain of virus of cattle paraflu-3 SCV No 2334 subfamily Paramixovirinae, genus Paramixovirus, inactivated virus suspension of strain of virus of cattle respiratory syncytial disease SCV No 2335, subfamily Pneumoviridae, genus Pneumovirus, inactivated virus suspension of strain of virus of cattle diarrhea SCV No 2336, family Flaviviridae, genus Pestivirus, with activity of each virus strain being not less than 106.5 TCD50 in 1 cm3 of vaccine, and inactivated suspension of cells of bacteria Pasteurella multocida serovar A strain 1231, inactivated suspension of cells of bacterium Pasteurella multocida serovar B strain 656, inactivated suspension of cells of bacterium Pasteurella multocida serovar D strain T-80, inactivated suspension of cells of bacterium Mannheimia haemolytica strain 169, in efficient quantity, taken in mixture in equal proportion with concentration not less than 3 billion microbial cells in vaccine dose. ^ EFFECT: vaccine possesses high specificity, stability in preservation of biological properties, creates expressed immunity, is harmless. ^ 3 cl, 10 tbl, 6 ex

Description

The invention relates to the field of virology, biotechnology and veterinary medicine and for the creation of new vaccines that can be used to prevent infectious diseases of cattle (cattle), accompanied by a violation of respiratory functions of varying severity.

Respiratory diseases of cattle are a group of heterogeneous pathologies caused by an extraordinary number of different factors of both infectious and non-infectious nature, including genetic predisposition, violation of sanitary and hygienic norms of keeping and feeding, and exposure to stress factors. In terms of breadth of spread, mortality, forced slaughter, and underweight of respiratory diseases, especially in calves, they significantly prevail over all other diseases and cause enormous economic damage. In some farms, the death of calves in combination with forced slaughter reaches 40-55%, and the return on feed in sick and ill animals is reduced by 2-3 times.

In recent years, massive respiratory diseases of cattle have been manifested as mixed viral and viral-bacterial infections, which are characterized by a particularly malignant course, which is difficult to diagnose, since a secondary bacterial infection “masks” the primary viral disease.

It has been established that the main role in the occurrence of respiratory diseases in young cattle is played by infectious rhinotracheitis viruses (IRT), parainfluenza-3 (PG-3), viral diarrhea (VD) and respiratory syncytial virus (PC), circulating in farms in various combinations. Of the bacterial pathogens, two types of pasteurella are usually involved in the development of respiratory diseases in an animal: Pasteurella multocida and Mannheimia haemolytica (Pasteurella haemolytica), which contribute to complications of bronchopneumonia in susceptible animals and act as a second microflora.

A common drawback of most known vaccines intended for use against massive infectious respiratory diseases in cattle and calves is their low efficiency as a result of the limited antigenic spectrum of action, the lack of a systemic focus, and the absence of bacterial antigens in the composition of the vaccines, which often leads to the development of lethal outcome in sick animals.

Known vaccine "TRIVAC (VIEV)" for the prevention of infectious rhinotracheitis, parainfluenza-3 and viral diarrhea - diseases of the mucous membranes of cattle containing antigens from attenuated strains of the viruses of infectious rhinotracheitis, parainfluenza-3 and diarrhea - mucosal diseases. As the virus strains, the vaccine contains the Herpes virus bovis strain "TK-A (VIEV) B-2" with an activity of 10 ^4.5 -10 ^6.5 TCD50 / ml, the Virus diarrhea strain - mucosal disease bovinum "BK-1 (B- 1) N 28 "with an activity of 10 ^4.0 -10 ^6.0 TCD50 / ml, Paramyxovirus strain" PTK-45/86 "with an activity of 10 ^5.0 -10 ^7.0 TCD50 / ml (Patent RU No. 21111011, A61K 39/295, publ. 05.20.1998).

The disadvantage of this vaccine is the low protection of cattle from pneumogastroenteritis due to the inferiority of the antigenic composition of its constituent components and the complex interactions of the viruses from which it is made.

Also known is a combined vaccine against viral pneumogastroenteritis of cattle and calves containing adjuvant and inactivated suspensions of infectious rhinotracheitis viruses, viral diarrhea, rotavirus and coronavirus infection in a titer of 10 ^7.5 -10 ^8.5 TCD _{50 /} cm ³ , inactivated suspensions of parainfluenza viruses -3 and respiratory syncytial infections - in the titer of 10 ^6.5 -10 ^7.5 TCD _{50 /} cm ³ taken in the vaccine in a volume ratio of 1: 1: 1: 1: 0.5: 0.5, respectively (Patent RU No. 2261111, A61K 39/295, publ. 09/27/2005).

However, the strains of viruses included in the known vaccine do not provide immunity to different subtypes and main antigenic variants of epizootic strains of pathogens circulating on farms, and, as a result, the known vaccine is not effective enough. In addition, the known vaccine does not allow to achieve a high level of antibody titers in the blood of cows and in colostrum, fed to newborn calves.

A known vaccine against animal pasteurellosis containing an inactivated bacterial suspension of Pasteurella multocida serological variants A, B, D from strains No. 1231, 681, T-80 with activity of soluble surface antigens in the reaction of passive hemagglutination as 1: 256-4096, 1: 512-8192 , 1: 256-4096, and the adjuvant emulsifier 139 and mineral oil Markol 52, taken in a mass ratio of 1: 10-12 (Patent RU No. 2311199, A61K 39/102, publ. 27.11. 2007).

The disadvantage of the animal pasteurellosis vaccine is that it does not provide immunological protection for animals against infectious bronchopneumonia of viral etiology due to the absence of viral antigens in the vaccine. In addition, the vaccine has a relatively low immunogenic activity with a high content of ballast substances and reactogenicity, which causes sensitization of the animal organism.

Known vaccine against viral diseases of cattle, characterized by respiratory syndrome, containing attenuated viruses of infectious rhinotracheitis, viral diarrhea; parainfluenza-3 and inactivated bacterial cells Pasteurella multocida and Pasteurella haemolytica (US Patent No. 3634587, C12K 5/00, publ. 11.01.1972).

However, this vaccine composition is characterized by low antiepizootic efficacy due to the fact that the strains used for its manufacture possess antigenic and immunobiological differences compared with similar characteristics of epizootic pathogens circulating in Russia and the CIS countries.

In addition, the use of live vaccines containing uninactivated antigens of the viruses of infectious rhinotracheitis and viral diarrhea is categorically contraindicated for pregnant cows and newborn calves because of the possibility of homologous recombination of vaccine and field strains of the virus, leading to exacerbation of infections, abortions and even death of individual animals.

The closest analogue is the inactivated combined vaccine against infectious rhinotracheitis, parainfluenza-3, respiratory syncytial disease, viral diarrhea and calf pasteurellosis KOMBOVAK-R, which is made from inactivated strains of infectious rhinotracheitis viruses, parainfluenza-3, viral diarrhea, respiratory respiratory diseases of cattle and two types of pasteurella: Pasteurella multocida (serovars A, B, D) and Pasteurella haemolitica (Instruction for the use of inactivated combined vaccine against ektsionnogo rhinotracheitis, parainfluenza-3, respiratory syncytial disease, viral diarrhea and pasteurellosis calves KOMBOVAK-R, approved by the Department of Veterinary 30.03.2007).

A disadvantage of the known combination vaccine is the relatively low titer of infectious activity of the viral suspensions included in its composition, as a result of which the necessary amount of protective antigens against these diseases is not provided in the vaccine dose.

The objective of the invention is to develop a safe combined (multivalent) vaccine containing antigenically and immunogenically balanced viral and bacterial antigens, which will create intense immunity against the most important pathogens associated with damage to the respiratory tract, and thus ensure the safety of young animals and productivity adult animals.

Another objective is to obtain new production strains of the viruses IRT, PG-3, PC and VD, which have a wide antigenic spectrum, high antigenic and immunogenic activity, not only in their native form, but also after inactivation, stably preserving their properties and suitable for creating inactivated combined vaccines different valencies.

The technical result of the invention is to expand the arsenal of combined vaccines against infectious diseases, accompanied by damage to the respiratory tract, as well as to increase the stability, antigenic and immunogenic activity of the combined vaccine and create intense and lasting immunity in vaccinated animals and born calves against infectious rhinotracheitis, parainfluenza-3, respiratory syncytial disease, viral diarrhea and cattle pasteurellosis.

The technical result of the invention is achieved through the use of new vaccine strains of infectious rhinotracheitis viruses, parainfluenza-3, respiratory syncytial disease and cattle viral diarrhea with a wide antigenic spectrum, as well as by creating a synergistic composition of a multivalent vaccine when combining new production strains of viruses with known vaccine strains of pasteurell.

The invention consists in the following:

The proposed inactivated combination vaccine against infectious rhinotracheitis, parainfluenza-3, respiratory syncytial disease, viral diarrhea and cattle pasteurellosis, containing the active substance and target additives, is characterized in that it contains a mixture of an inactivated suspension of a large strain of infectious rhinotracheitis virus STB cattle No. 2333, family Herpesviridae, subfamily Alphaherpesviridae; inactivated suspension of the strain of parainfluenza virus 3 of bovine bovine bovine virus No. 2334, subfamily Paramixovirinae, genus Paramixovirus; inactivated suspension of a strain of the virus of respiratory syncytial disease of cattle STB No. 2335, subfamily Pneumoviridae, genus Pneumovirus; inactivated suspension of the strain of bovine diarrhea virus GKV No. 2336, family Flaviviridae, genus Pestivirus, inactivated suspension of bacterial cells of Pasteurella multocida serovar A strain 1231, inactivated suspension of bacterial cells of Pasteurella multocida serovar B strain 656, inactivated suspension of bacterial cells of Pasteurera multacida D multurec -80, an inactivated suspension of bacterial cells of Mannheimia haemolytica (Pasteurella haemolytica) strain 169, in an effective amount.

The term “effective amount” means an amount of an active substance which, when administered to cows and / or calves, provides the induction of a specific immune response of a sufficient level in animals, followed by a warning against the disease and the symptoms of the disease to be prevented.

The vaccine rhinotracheitis virus strain of bovine bovine animals No. 2333, the Herpesviridae family, the Alphaherpesviridae subfamily strain, the bovine parainfluenza-3 virus strain No. 2,334, the Paramixovirinae subfamily of the bovine genus Paramixovirus, and the virus No. 2335, subfamilies of Pneumoviridae, genus Pneumovirus, and a strain of bovine diarrhea virus of bovine bacilli No. 2336, of the family Flaviviridae, genus Pestivirus, are new production strains and are used for the first time as part of vaccines preparations.

The term "production strain" means a vaccine strain, studied by its biological properties, stable during storage and reseeding, harmless after inactivation or attenuation for the animal, well adapted to the cultivation system, with high antigenic and immunogenic activity both for a sensitive laboratory model and for naturally susceptible animals, and also characterized by a wide antigenic spectrum that dominates epizootic strains of the pathogen.

The production strain of the infectious rhinotracheitis virus of cattle, STB No. 2333, was deposited in the State Virus Collection of the Institute of Virology of the Russian Academy of Medical Sciences, has the registration number of the STB participant No. 2333 and is characterized by the following features and properties.

Source of Origin:

The source of the production of a new production strain of the cattle infectious rhinotracheitis virus is the B-25 virus (author's name), isolated in 1991 from a nasal flush of a calf with respiratory syndrome, which passed 35 consecutive passages in cattle transplant cultures.

Morphological properties:

GKV strain No. 2333 belongs to the family Herpesviridae, subfamily Alphaherpesviridae. An electron microscopic examination using the method of negative contrasting of the virus-containing culture material revealed virions characteristic of the cattle infectious rhinotracheitis virus, 150-200 nm in size and consisting of a capsid of cubic symmetry and an outer shell covering it. Capsid has 162 prismatic hollow capsomeres. A nucleocapsid is formed inside the nucleus, where type A inclusions are formed.

Antigenic properties:

GKV strain No. 2333 shows high antigenic and immunogenic activity both in the native form and after inactivation, has a wide antigenic spectrum and dominates in relation to field strains.

The introduction of guinea pigs strain GKV No. 2333 of the IRT virus leads to the formation of a humoral immune response to various strains and field isolates of the IRT virus (table 1).

Table 1 Cross antigenic activity of strain GKV No. 2333 virus IRT RTI virus strain / isolate for pH The titer of virus-neutralizing antibodies in the blood serum of guinea pigs immunized with strain ГКВ №2333 of the virus ИРТ *, М ± m ГКВ №2333 142.1 ± 24.2 B-25 84.4 ± 13.8 Isolate No. 1702 76.1 ± 23.9 Isolate No. 2105 87.3 ± 12.5 Isolate No. 1320 111.4 ± 26.5 * hereinafter in the tables the reverse values of antibody titer are given

Biotechnological characteristics:

GKV strain No. 2333 proliferates well in primary and transplanted cell cultures (kidneys, testicles) obtained from tissues and organs of cattle. In these cell cultures, the virus accumulates in a titer of 7.5-8.5 lg TCD ₅₀ / cm ³ .

Recommended storage method:

In a lyophilized state at a temperature not exceeding minus 60 ° С.

Frequency of refreshment:

Once every three years in a transplanted culture of MDVC calf kidney cells (hereinafter referred to as MDVC cell culture).

Additional information about the strain:

GKV strain No. 2333 is not contaminated by extraneous microflora. Tested with reference serum (GB).

Reactogenicity - strain GKV No. 2333 does not have reactogenic properties.

Stability - retains the original biological (antigenic) properties during successive passages in the culture of MDVC cells for 10 passages (observation period).

Pathogenicity - expressed.

Contagiousness - expressed.

Oncogenicity is absent.

The bovine parainfluenza virus-3 virus strain No. 2,334 was deposited with the State Virus Collection of the Institute of Virology of the Russian Academy of Medical Sciences, has the registration number of No. 2,334, and it is characterized by the following features and properties.

Source of Origin:

The source of the production of a new strain of the production virus of parainfluenza-3 cattle is the B-19 virus strain (author's name), isolated in 1990 from a nasal washout of a calf with signs of rhinitis in a calf kidney cell culture, which passed 67 consecutive passages in the MDVK cell culture

Morphological properties:

GKV strain No. 2334 relates to the order Paramixoviridae, subfamily Paramixovirinae, genus Paramixovirus. An electron microscopic examination using the method of negative contrasting of the virus-containing cultural material revealed virions characteristic of the PG-3 virus: a polymorphic slightly oval shape, the size of the virion varies from 150 to 250 nm. The virion consists of a shell and an internal component, which is represented by a ribonucleoprotein helix.

Antigenic properties:

GKV strain No. 2334 shows high antigenic and immunogenic activity both natively and after inactivation, possesses a wide antigenic spectrum and dominates against field strains.

The introduction of guinea pigs strain GKV No. 2334 virus PG-3 leads to the formation of a humoral immune response to various strains and field isolates of the virus PG-3 (table 2).

Table 2 Cross antigenic activity of strain GKV No. 2334 virus PG-3 PG-3 virus strain / isolate for pH The titer of virus-neutralizing antibodies in the blood serum of guinea pigs immunized with the strain of STB No. 2334 of the virus PG-3, M ± m ГКВ №2334 322.4 ± 18.5 B-19 276.2 ± 36.7 Isolate No. 543 111.4 ± 26.5 Isolate No. 486 161.1 ± 25.6

Biotechnological characteristics:

GKV strain No. 2334 proliferates well in primary and transplantable cell cultures (kidneys, lungs) obtained from tissues and organs of cattle. In these cell cultures, the virus accumulates in a titer of 7.5-9.0 lg TCD ₅₀ / cm ³ .

Recommended storage method:

In a lyophilized state at a temperature not exceeding minus 60 ° С.

Frequency of refreshment:

Once every three years in a culture of MDVC cells.

Additional information about the strain:

GKV strain No. 2334 is not contaminated by extraneous microflora.

Tested with reference serum (GB).

Reactogenicity - strain GKV No. 2334 does not have reactogenic properties.

Stability - retains the original biological (antigenic) properties during successive passages in the culture of MDVC cells for 10 passages (observation period).

Pathogenicity - expressed.

Contagiousness - expressed.

Oncogenicity is absent.

The bovine bovine diarrhea virus production strain No. 2,336 was deposited with the State Virus Collection Institute of the Institute of Virology of the Russian Academy of Medical Sciences, has the registration number of No. 2,336 bills and is characterized by the following features and properties.

Source of Origin:

The source of the new production strain of STB diarrhea virus No. 2336 is the ST-89 virus strain (author's name), isolated in 1989 from the intestines of calves with signs of diarrhea in the cell culture of the kidney of a newborn calf, which passed 82 consecutive passages in a transplanted cell culture of MDVC .

Morphological properties:

GKV strain No. 2336 belongs to the family Flaviviridae, genus Pestivirus. Electron microscopic examination using the method of negative contrasting of virus-containing culture material revealed virions characteristic of the cattle diarrhea virus, 40-60 nm in size and consisting of icosahedral symmetric nucleocapsid and external lipoprotein membrane.

Antigenic properties:

GKV strain No. 2336 shows high antigenic and immunogenic activity both natively and after inactivation, possesses a wide antigenic spectrum and dominates against field strains.

The introduction of Guinea pigs strain No. 2,336 of the cattle diarrhea virus leads to the formation of a humoral immune response to various strains and field isolates of the VD virus (Table 3).

Table 3 Cross antigenic activity of the strain of ST-Bills No. 2336 virus VD VD strain / isolate for pH The titer of virus-neutralizing antibodies in the blood serum of guinea pigs immunized with the strain of STB No. 2336 of the VD virus, M ± m ГКВ №2336 374.8 ± 31.6 ST-89 107.6 ± 24.5 Isolate No. 865 168.9 ± 21.3 Isolate No. 1204 337.7 ± 26.8 Isolate No. 663 174.8 ± 22.4

It has a close antigenic relationship with the viruses of classical swine fever and border disease of sheep.

Biotechnological characteristics:

GKV strain No. 2336 multiplies well in the primary culture of calf kidney cells (cow embryos) and in the MDVK cell culture. In these cell cultures, the virus accumulates in a titer of 10 ^7.5 -10 ^8.5 MCC ₅₀ / cm ³ .

Recommended storage method:

In a lyophilized state at a temperature not exceeding minus 60 ° С.

Frequency of refreshment:

Once every three years in a culture of MDVC cells.

Additional information about the strain:

GKV strain No. 2336 is not contaminated by extraneous microflora. It is tested with reference serum (GB) and in the PCR test system.

Reactogenicity - the strain of ST-Bills No. 2336 does not have reactogenic properties.

Stability - retains the original biological (antigenic) properties during successive passages in the culture of MDVC cells for 10 passages (observation period).

Pathogenicity - expressed.

Contagiousness - expressed.

Oncogenicity is absent.

The production strain of the virus of respiratory syncytial disease in cattle, STB No. 2335, was deposited in the State Virus Collection of the Institute of Virology RAMS, has the registration number of the STB participant No. 2335 and is characterized by the following features and properties.

Source of Origin:

The source of the production of a new strain of the production cattle PC virus is virus strain 89 (the author’s name), isolated in 1989 from nasal swabs of calves with signs of rhinitis and pneumonia in a cell culture of a kidney of a cow embryo, which passed 72 consecutive passages in a culture of MDVC cells.

Morphological properties:

GKV strain No. 2335 relates to the order Paramixoviridae, subfamily Pneumoviridae, genus Pneumovirus. Electron microscopic examination using the method of negative contrasting of virus-containing culture material revealed virions characteristic of respiratory syncytial cattle virus, 200-300 nm in size and consisting of a nucleocapsid of spiral symmetry and an external lipoprotein membrane, on the surface of which there are protrusions 8-12 nm long.

Antigenic properties:

GKV strain No. 2335 shows high antigenic and immunogenic activity both natively and after inactivation, possesses a wide antigenic spectrum and dominates against field strains.

The introduction of Guinea pigs strain GKV No. 2335 of the PC virus leads to the formation of a humoral immune response to various strains and field isolates of the PC virus (table 4).

Table 4 Cross antigenic activity of a strain of ST-Bills No. 2335 of the PC virus PC virus strain / isolate for pH The titer of virus-neutralizing antibodies in the blood serum of guinea pigs immunized with the STB strain No. 2335 of the PC virus, M ± m ГКВ №2335 162.5 ± 22.8 PC-89 81.6 ± 22.5 Isolate No. 124 62.3 ± 16.4

Antigenically linked to the human PC virus.

Biotechnological characteristics:

GKV strain No. 2335 proliferates well in the primary culture of calf kidney cells (cow embryos) and in the MDVK cell culture. In these cell cultures, the virus accumulates in the titer of 6.5-7.0 lg TCD _{50 /} cm ³

Recommended storage method:

In a lyophilized state at a temperature not exceeding minus 60 ° С.

Frequency of refreshment:

Once every three years in a culture of MDVC cells.

Additional information about the strain:

The strain is not contaminated by extraneous microflora. Tested with reference serum (GB).

Reactogenicity - the strain of ST-Bills No. 2335 does not have reactogenic properties.

Stability - retains the original biological (antigenic) properties during successive passages in the culture of MDVC cells for 10 passages (observation period).

Pathogenicity - expressed.

Contagiousness - expressed.

Oncogenicity is absent.

As a part of the combined vaccine against infectious rhinotracheitis, parainfluenza-3, respiratory syncytial disease, viral diarrhea and cattle pasteurellosis, inactivated viral suspensions of the strains of GKV No. 2333, GKV No. 2334, GKV No. 2335 and GKV No. 2336 are contained with the titer of the infectious activity of each virus not less than 10 ^6.5 TCD ₅₀ in 1 cm ^{3 of the} vaccine.

Inactivation of viral suspensions is carried out by any known method, for example, by adding formalin or ethyleneimine dimer (DEI).

In the composition of the proposed vaccine, viral suspensions of the strains can be used in any combination by volume, for example, with the titer of the infectious activity of the strains STB No. 2333, STB No. 2336, STB No. 2334 - 10 ^8.5-9.0 MTC ₅₀ / cm ³ and strain STB No. 2335 - 10 ^7.0-7.5 TCD ₅₀ / cm ³ viral suspensions are mixed in a ratio by volume of 1: 1: 1: 1 (mixture No. 1); or titer of infectious activity strains STB №2333, STB №2334 - 10 ^7.5-8.5 TCD ₅₀ / cm ^3, strain STB №2336 - 10 ^7.5-8.5 TCD ₅₀ / cm ^3, strain STB № 2335 - 10 ^7.0-7.5 TCD ₅₀ / cm ³ viral suspensions are mixed in a ratio by volume of 2.5: 2.5: 1: 1 (mixture No. 2); or with the titer of infectious activity of strains of STB No. 2333, STB No. 2336, STB No. 2334 - 10 ^7.5-8.5 TCD ₅₀ / cm ³ and strain of STB No. 2335 - 10 ^6.5-7.0 TCD ₅₀ / cm ³ viral suspensions are mixed in a ratio by volume of 2: 1.5: 1.5: 1.5 (mixture No. 3). It should be noted that a double increase or decrease in volume of one of the components does not significantly affect the antigenic activity of the finished vaccine.

The strains of Pasteurella multocida (strain 1231 (serovar A), strain 656 (serovar B), strain T-80 (serovar D)) and Mannheimia haemolytica (Pasteurella haemolytica) (strain 169) are known and used in various combinations in the manufacture of vaccines and sera against animal pasteurellosis (see, for example, copyright certificate SU No. 1792708, RF patents No. 2118169, 2191598, 2246316, 2304442, etc.). However, we found that when combining the known strains of pasteurella in the combination according to the invention with new production strains of viruses, the formation of a synergistic composition is observed, which manifests itself in an increase in tension and duration of immunity to the viral and pasteurellosis components of the vaccine.

The composition of the proposed vaccine inactivated cell suspension of bacteria Pasteurella multocida (strain 1231 (serovar A), strain 656 (serovar B), strain T-80 (serovar D)) and Mannheimia haemolytica (Pasteurella haemolytica) (strain 169) are contained in equal proportions with a final concentration of at least 3 billion microbial cells in 1 cm of the vaccine.

Inactivation of pasteurell cell suspensions is carried out by any known method, for example, by adding formalin containing at least 37% formaldehyde.

According to the invention, the combination vaccine also contains one or more veterinary acceptable target additives.

The term “target additive” means any pharmaceutical components acceptable in veterinary medicine that are traditionally used to ensure the preservation of antigens during processing, preparation of the final prescription form or subsequent storage and the like.

As a targeted additive, the combination vaccine may contain a solvent, an adjuvant, a solubilizing agent, a diluent, a preservative, a stabilizer, an antibacterial and antifungal agent, an isotonic agent, an absorption delaying agent, and the like. Diluents may include distilled water, saline, saline, dextrose, ethanol, glycerol and the like. Isotonic agents may include, among others, sodium chloride, dextrose, mannitol, sorbitol, and lactose. Stabilizers include albumin, sucrose, gelatose, skim milk, etc.

Adjuvants include substances that non-specifically enhance the immune response to antigens and can be of mineral origin - aluminum oxide hydrate, aluminum phosphate, alum-potassium alum; plant origin - saponin (Quil-A); adjuvants based on mineral and non-mineral oils; synthetic adjuvant - muramyl dipeptide, etc.

As a target additive, the proposed vaccine mainly contains an adjuvant, for example, alumina hydrate or a mixture of alumina hydrate with saponin.

If necessary, as a target additive, the proposed vaccine may contain a preservative sodium thiolate in an amount of 1: 10000.

Although a person skilled in the art can easily determine the amount and concentration of adjuvants and additives that can be used in the context of the present invention, the present invention relates to compositions comprising preferably from 20 to 30 vol.% Adjuvant.

In a preferred embodiment, the vaccine contains 6% alumina hydrate gel in an amount of 20 vol.% And saponin in an amount of 0.05 vol.%, Which corresponds to 12 g of alumina hydrate and 500 mg of saponin in 1 liter of vaccine.

The following formulation examples are illustrative only and do not limit the scope of the present invention.

The inactivated combined vaccine against infectious rhinotracheitis, parainfluenza-3, respiratory syncytial disease, viral diarrhea and cattle pasteurellosis is a light red liquid, during storage, a loose grayish precipitate forms, which easily breaks up into a homogeneous suspension with shaking.

The vaccine is stable and retains its original activity at a temperature of 2-8 ° C for at least 18 months (observation period).

In order to prevent respiratory diseases, the resulting vaccine is administered intramuscularly to pregnant animals twice: the first time - 50-60 days before calving, the second time - 14-21 days after the first immunization (no later than 30 days before calving) at a dose of 3 cm ³ . Calves aged 30 days and older are vaccinated twice with an interval of 20-25 days at a dose of 2 cm ³ . Animals older than 4 months of age are vaccinated twice intramuscularly at a dose of 3 cm ³ with an interval of 14-21 days. Revaccination is carried out once in the same dose, every 6 months.

The vaccine was tested on cattle in dysfunctional farms. This testing confirms the harmlessness of the drug and its antiepizootic efficacy, manifested in enhancing the immunizing activity of the components of the inactivated vaccine. Vaccination increases immunity in mothers and protects offspring during embryonic development and in the postnatal period. Moreover, the induced immune response was characterized by high specificity with respect to different subtypes and main antigenic variants of epizootic strains of pathogens circulating on farms.

The protective level of antibodies in animals vaccinated with this vaccine was maintained for at least 8 months in adult animals and 6 months in young animals under the age of 1 year, colostral immunity - up to 1.5 months.

The invention is illustrated, but not limited to the following examples.

Example 1. Obtaining an inactivated combination vaccine against infectious rhinotracheitis, parainfluenza-3, respiratory syncytial disease, viral diarrhea and cattle pasteurellosis.

The vaccine is prepared from new production strains of the infectious rhinotracheitis virus GKB No. 2333, parainfluenza virus 3 GKV No. 2334, respiratory syncytial virus GKV No. 2335, diarrhea virus GKV No. 2336 (viral component) and known vaccine strains of pasteurellum serum Pasteurella multocida (12 pasteurella multocida A) strain 656 (serovar B), strain T-80 (serovar D)), Mannheimia haemolytica (Pasteurella haemolytica) (strain 169) (pasteurellose component).

The technological process consists of the following main stages: the stage of obtaining the viral component, the stage of obtaining the pasteurellosis component, combining the obtained viral and pasteurellosis components, adding an adjuvant, mixing all components of the vaccine with subsequent packaging of the finished product.

A. Stage of obtaining the viral component of the combination vaccine.

The manufacture of the viral component of the combination vaccine includes separate steps for the preparation of viral suspensions from seeds of the strains of the infectious rhinotracheitis virus GKV No. 2333, parainfluenza virus-3 GKV No. 2334, respiratory syncytial virus GKV No. 2335 and diarrhea virus GKV No. 2333; determination of the infectious activity of each viral suspension, inactivation of virus-containing culture fluids of each strain and their combination.

The step for producing viral suspensions according to the invention is based on a seed lot system, in which successive series of products are obtained from the main seed (master seed lot) for a certain number of transfers (passages). In the current production, the working seed lot is prepared from the main seed. The finished product is made from working seed, and the number of transfers from the main seed should not exceed the value established during clinical trials of vaccines, based on safety and efficacy requirements.

The process of obtaining a seed system of each strain of the virus includes several stages:

- preparation of cell culture;

- carrying out passages of the virus;

- obtaining a viral suspension;

- intermediate control of a strain of the virus;

- lyophilization, freezing or inactivation of the virus strain;

- control of the strain of the virus.

The strains of infectious rhinotracheitis viruses ГКВ №2333, parainfluenza-3 ГКВ №2334, respiratory syncytial ГКВ №2335 and viral diarrhea ГКВ №2336 are grown separately in the transplanted cell culture of MDVC calf kidney.

The accumulation of viruses in cell culture is assessed by the cytopathic effect. The virus titer is calculated by the method of Reed and Mench and expressed in lg TCD ₅₀ / cm ³ .

The accumulation of viruses in cell culture is assessed by the cytopathic effect, the accumulation of PG-3 virus also by hemagglutinating activity using guinea pig erythrocytes. The virus titer is calculated by the method of Reed and Mench and expressed in lg TCD ₅₀ / cm ³ .

Upon reaching the titer of infectious rhinotracheitis viruses, viral diarrhea and parainfluenza-3 to values of 10 ^7.5 -10 ^9.0 TCD ₅₀ / cm ³ , and the virus respiratory syncytial infections - 10 ^6.5 -10 ^7.5 TCD ₅₀ / cm ³ , the resulting virus-containing material of each strain is purified from ballast impurities by centrifugation or any other known method.

Further, virus suspensions are inactivated with a formalin solution containing at least 37% formaldehyde. The completeness of inactivation is assessed by the absence of virus propagation in a sensitive cell culture for four passages.

Inactivated virus suspensions are combined so that in each series the titer of the infectious activity of each of the strains is at least 10 ^6.5 TCD ₅₀ / cm ³ .

Moreover, viral suspensions of the strains can be used in any combination by volume, for example, with the titer of the infectious activity of strains of STB No. 2333, STB No. 2336, STB No. 2334 - 10 ^8.5-9.0 TCD ₅₀ / cm ³ and strain STB No. 2335 - 10 ^7.0-7.5 TCD ₅₀ / cm ³ viral suspensions are mixed in a ratio by volume of 1: 1: 1: 1 (mixture No. 1); or titer of infectious activity strains STB №2333, STB №2334 - 10 ^7.5-8.5 TCD ₅₀ / cm ^3, strain STB №2336 - 10 ^7.5-8.5 TCD ₅₀ / cm ^3, strain STB № 2335 - 10 ^7.0-7.5 TCD ₅₀ / cm ³ viral suspensions are mixed in a ratio by volume of 2.5: 2.5: 1: 1 (mixture No. 2); or with the titer of infectious activity of strains of STB No. 2333, STB No. 2336, STB No. 2334 - 10 ^7.5-8.5 TCD ₅₀ / cm ³ and strain of STB No. 2335 - 10 ^6.5-7.0 TCD ₅₀ / cm ³ viral suspensions are mixed in a ratio by volume of 2: 1.5: 1.5: 1.5 (mixture No. 3).

B. Stage receiving pasteurellosis component of the combination vaccine.

Pasteurella cultures of Pasteurella multocida (strain 1231 (serovar A), strain 656 (serovar B), strain T-80 (serovar D)), Mannheimia haemotytica (Pasteurella haemolytica) (strain 169) were grown separately using nutrient media based on the Hottinger c pH 7.9-8.0 and an amino nitrogen content of at least 178-180 mg%. Cultivation is carried out at a temperature of 37 ° C ± 1 ° C for 8-10 hours. Cultivation is carried out until the maximum accumulation of surface soluble antigens in the growth medium, determined by the level of titers in the reaction of passive hemagglutination (RPHA).

Next, the resulting culture will be combined in a ratio of 1: 1: 1: 1, formalin is added to a concentration of 0.3%. The resulting mixture of inactivated cell suspensions is concentrated by ultrafiltration to obtain a back mass with a content of 100 billion microbial cells in 1 cm ³ or more.

B. The step of combining the viral and pasteurellose components of the combination vaccine.

Option 1.

Inactivated virus suspensions prepared in stage A are mixed in the prescribed ratio depending on the titer of the infectious activity of the viral suspension of each strain (mixture No. 1), mixed thoroughly. Then add 6% gel of aluminum hydroxide (GOA) in an amount of 20% of the total volume, add saponin at a concentration of 500 mg / l and preservative sodium thiolate at a concentration of 1: 10000, mix thoroughly.

Sodium merthiolate at a concentration of 1: 10000 as a preservative is added to the inactivated bacterial cell suspension of the pasteurellum strain mixture prepared in stage B. Then it is mixed with 6% GOA (20% of the total volume), mixed thoroughly.

The viral and bacterial parts of the drug are mixed in such a way that the concentration of viral antigens in the finished vaccine is at least 6.5 lg TCD ₅₀ , pasteurell - at least 3 billion cubic meters. in 1 cm ³ .

Option 2

Inactivated virus suspensions prepared in stage A are mixed in the prescribed ratio depending on the titer of the infectious activity of the viral suspension of each strain (mixture No. 2), mixed thoroughly. Then add 6% gel of aluminum hydroxide (GOA) in an amount of 20% of the total volume, add saponin at a concentration of 500 mg / l and preservative sodium thiolate at a concentration of 1: 10000, mix thoroughly.

To an inactivated bacterial cell suspension of a mixture of pasteurellum strains prepared in stage B., 6% GOA (20% of the total volume) is added and mixed thoroughly.

The viral and bacterial parts of the drug are mixed in such a way that the concentration of viral antigens in the finished vaccine is at least 6.5 lg TCD ₅₀ , pasteurell - at least 3 billion cubic meters. in 1 cm ³ .

Option 3

Inactivated virus suspensions prepared in stage A are mixed in the prescribed ratio, depending on the titer of the infectious activity of the viral suspension of each strain (mixture No. 3), mixed thoroughly. Then add 6% gel of aluminum hydroxide (GOA) in an amount of 30% of the total volume, mix thoroughly.

To inactivated bacterial cell suspension of a mixture of leptospira strains prepared in stage B., add 6% GOA (30% of the total volume) and mix thoroughly.

The viral and bacterial parts of the drug are mixed in such a way that the concentration of viral antigens in the finished vaccine is at least 6.5 lg TCD ₅₀ , pasteurell - at least 3 billion cubic meters. in 1 cm ³ .

The viral and bacterial parts of the drug are mixed in such a way that the concentration of viral antigens in the finished vaccine is at least 6.5 lg TCD ₅₀ , pasteurell - at least 3 billion cubic meters. in 1 cm ³ .

The obtained variants of the inactivated combined vaccine against infectious rhinotracheitis, parainfluenza-3, respiratory syncytial disease, viral diarrhea, and cattle pasteurellosis have the component composition shown in Table 5

Table 5 The content of ingredients in the liquid mixture, vol.% Ingredient Titer, lg TCD ₅₀ / cm ³ The content in the liquid mixture, vol.% Option 1 Option 2 Option 3 IRT virus 7.5-8.5 - 25 twenty 8.5-9.0 17.5 - - PG-3 virus 7.5-8.5 - 25 fifteen 8.5-9.0 17.5 - - PC virus 6.5-7.0 - - fifteen 7.0-7.5 17.5 10 - VD virus 7.5-8.5 - - fifteen 8.5-9.0 17.5 10 - Pasteurellum cell suspension - 9.94 9.94 5,0 GOA (6% gel) - twenty twenty thirty Saponin - 0.05 0.05 - Mertiolate Sodium - 0.01 0.01 -

Example 2. Evaluation of the antigenic activity of a combination vaccine in experimental models.

To determine the antigenic activity of the combination vaccine according to the invention, laboratory animals are used as an experimental model, while for assessing the antigenic activity of the viral components of the vaccine, guinea pigs weighing 350-400 g, pasteurellosis component are used, rabbits weighing 3-3.5 kg and white mice weighing 16 -18 g

Groups of 10 animals are formed in each: 3 experimental groups and 1 control (guinea pigs), 3 experimental groups and 1 control (rabbits).

Guinea pigs of the experimental groups are immunized with the test vaccine once at a dose of 0.25 cm ³ ; 0.5 cm ³ and 1.0 cm ^3, respectively, for rabbits twice with an interval of 30-35 days for the first time at a dose of 1.5 cm ³ , the second time at a dose of 3.0 cm ³ . In this case, the vaccine is administered intramuscularly to guinea pigs and rabbits. Animal control groups are not immunized.

On the 21st day after the injection of the vaccine, the blood is taken from the guinea pigs and the serum is separated, which is then examined for the presence of virus-specific antibodies.

The antigenic activity of the viral component of the vaccine is assessed by the level of virus-specific antibodies to the viruses IRT, PG-3, PC and VD. The antibody titer is determined in the neutralization reaction (PH), which is carried out in 96-well plates according to the standard method by double dilutions of the test serum against a working dose of each virus of 100 TCD ₅₀ / well.

The virus neutralizing titer of the test serum is expressed as its limiting dilution, inhibiting the development of the virus's CPD in 50% of infected cell cultures.

The immunogenic activity of the pasteurellosis component is determined by the preventive activity of the blood serum of vaccinated rabbits for white mice.

To do this, 14 days after the second injection of the vaccine, blood samples are taken from rabbits, the obtained blood serum is mixed in equal volumes, and the collection sample is injected subcutaneously to 10 white mongrel mice at a dose of 0.5 cm ³ . After 24 hours, 10 immunized and 10 control mice were infected with a virulent strain of P. multocida No. 1231 (Serovar A) at a dose of 5 LD ₅₀ . The observation period is 10 days.

The results of the study are presented in table 6, which indicate that the combined vaccine against IRT, PG-3, PC, VD and cattle pasteurellosis is superior to existing analogues in antigenic and immunogenic activity. So, the proposed vaccine already 21 days after immunization provides the production of specific antibodies to all components of the drug in high titer: to the IRT virus - 1: 154.0; to the PG-3 virus - 1: 180.0; PC virus - 196.5; to the VD virus - 1: 165.3, the preventive activity of rabbit blood serum was at least 90%.

Animals of the control group throughout the experiment were seronegative to the viruses IRT, PG-3, PC, VD and pasteurell.

Table 6 Antigenic activity of the combined vaccine against infectious rhinotracheitis, parainfluenza-3, respiratory syncytial disease, viral diarrhea and cattle pasteurellosis in experimental models Live group Guinea pig serum antibody titer Batch number Preventive activity of rabbit blood serum to pasteurell% The dose of the drug, cm ³ The titer of neutralizing antibodies, M ± m IRT PG-3 PC VD I 0.25 0 42.2 ± 7.1 59.7 ± 7.2 55.7 ± 16.4 one 90 II 0.5 157.6 ± 28.6 , 274.4 ± 25.3 194.0 ± 26.7 168.9 ± 24.4 2 one hundred III 1,0 78.8 ± 19.6 119.4 ± 15.7 207.9 ± 28.1 157.6 ± 28.6 3 one hundred the control - 0 0 0 0 0 0

Example 3. Evaluation of the stability of the combination vaccine during storage.

The stability test of the combination vaccine obtained in example 1 is carried out on experimental animals - guinea pigs and rabbits. Moreover, to assess the stability of the antigenic activity of the viral components of the vaccine, guinea pigs and the pasteurellosis component are used in rabbits.

For this, the vaccine is administered after 6, 12 and 18 months of storage at a temperature of 4 ° C for guinea pigs once in a dose of 0.5 cm ³ . Twenty-one days after vaccination, the blood serum of the animals was taken for examination. Rabbits are vaccinated twice with an interval of 30-35 days at a dose of 1.5 and 3.0 cm ^3, respectively. 14 days after the second injection of the vaccine, blood samples were taken from rabbits, the obtained blood serum was mixed in equal volumes and the preventive activity of blood serum was determined for white mice.

The stability of the viral component of the vaccine is evaluated in pH by the absence of a decrease in antibody titer below the established level (1:16), pasteurellosis - by the preservation of preventive activity (must be at least 90% of surviving white mice) when stored at + 4 ° C for 18 months .

The test results are presented in table.7.

Table 7 Change in antigenic activity of the combined vaccine during storage Storage time Antigenic activity of a vaccine against viral antigens Immunogenic activity of the pasteurella vaccine IRT PG-3 PC VD Preventive activity of blood serum of vaccinated rabbits for white mice,% surviving animals In the manufacture 157.6 ± 28.6 274.4 ± 25.3 194.0 ± 26.7 168.9 ± 24.4 90 6 months 137.2 ± 22.8 256.0 ± 32.4 168.9 ± 21.8 157.6 ± 28.6 one hundred 12 months 103.9 ± 18.1 207.9 ± 28.1 119.4 ± 15.7 137.2 ± 22.8 80 18 months 78.8 ± 19.6 168.9 ± 24.4 78.9 ± 19.6 119.4 ± 15.7 70

Example 4. Production test of experimental series of combination vaccines in experiments on cows.

The test is carried out on cows and heifers of a random age in farms with a high level of respiratory diseases of calves that are unsuccessful for infectious rhinotracheitis, parainfluenza-3, respiratory syncytial disease, viral diarrhea and cattle pasteurellosis.

In this case, animals are divided into 2 groups: experimental and control, 20 goals in each group.

For the animals of the experimental group, the combined vaccine obtained according to Example 1 is administered intramuscularly to pregnant animals at a dose of 3.0 cm ³ twice with an interval of 14-21 days, a second time no later than 30 days before calving.

In the control group, animals are not vaccinated.

To determine the tension and duration of immunity in all animals, blood is taken before vaccination and after 1, 2, 4, 6, 8 and 12 months after vaccination.

Vaccination efficiency is assessed by the level of virus-neutralizing antibodies in the blood serum of cows and pasteurellas by the preventive activity of the blood serum of vaccinated animals for white mice infected with 5 LD ₅₀ virulent strain of pasteurellas.

The test results are presented in table.8.

Table 8 Evaluation of the activity of the vaccine against infectious rhinotracheitis, parainfluenza-3, respiratory syncytial disease, viral diarrhea and cattle pasteurellosis in an experiment on cows Research Dates Antigenic activity of a vaccine against viral antigens Immunogenic activity of the pasteurella vaccine IRT PG-3 PC VD Preventive activity of blood serum of vaccinated rabbits for white mice,% surviving animals Experienced group Before vaccination 57.3 ± 6.6 54.9 ± 5.2 15.2 ± 5.3 28.6 ± 2.2 twenty 1 month p / vacc. 234.7 ± 31.1 241.8 ± 18.8 201.1 ± 18.7 412.4 ± 26.9 80 2 months p / vacc. 209.1 ± 16.2 233.2 ± 25.3 233.4 ± 42.3 412.5 ± 18.2 one hundred 4 months p / vacc. 1b8.9 ± 24.4 194.0 ± 26.7 168.9 ± 21.8 337.8 ± 24.9 one hundred 6 months p / vacc. 157.6 ± 28.6 119.4 ± 15.7 137.2 ± 22.8 274.4 ± 25.3 80 8 months p / vacc. 84.4 ± 12.5 103.9 ± 18.1 97.0 ± 18.6 256.0 ± 32.4 60 12 months vaccine 55.7 ± 10.9 90.5 ± 20.3 45.3 ± 11.5 78.9 ± 19.6 40 Control group - 52.0 ± 13.8 48.5 ± 12.4 18.4 ± 10.2 35.1 ± 8.6 10

The results of the study show that immunization of cows with the combination vaccine according to the invention causes the formation of a pronounced immune response to all antigens that make up the vaccine for at least 8 months.

Example 5. Production test of experimental series of combined vaccines in experiments on calves.

To determine the prophylactic effectiveness of the combination vaccine obtained in Example 1, 60 calves aged 30-40 days born from unvaccinated mothers were selected.

3 groups of 20 goals are formed in each: 2 experimental and control.

The calves of the experimental groups studied combination vaccine is administered intramuscularly at a dose of 1.0 and 2.0 cm ³ . Revaccination is carried out 20-25 days after the first immunization in the same doses.

The calves of the control group are not given a vaccine.

Before each administration of the drug and 21 days after the 2nd vaccination, blood was taken from all animals and the level of virus-specific antibodies in the blood serum in the PH was determined, as well as the preventive activity of the blood serum of vaccinated calves for white mice infected with 5 LD ₅₀ virulent pasteurella strain. The test results are presented in table.9.

Table 9 The level of specific antibodies in the blood serum of calves immunized with the combination vaccine of the invention Group of animals The dose of the vaccine, cm ³ The titer of neutralizing antibodies, M ± m To the pathogen Before vaccination 21 days after the 1st vaccination 21 days after the 2nd vaccination Experienced group I 1,0 IRT 6.1 ± 3.0 12.5 ± 1.1 85.7 ± 3.1 PG-3 5.7 ± 6.9 18.3 ± 5.2 117.1 ± 17.9 PC 5.9 ± 2.9 28.3 ± 4.2 123.1 ± 24.9 VD 2.6 ± 2.1 53.7 ± 15.1 401.7 ± 38.0 Pasteurella * 0 40 60 II 2.0 IRT 7.0 ± 8.8 76.0 ± 9.4 205.0 ± 33.7 PG-3 5.0 ± 9.5 36.5 ± 8.7 294.0 ± 24.5 PC 5.4 ± 2.5 42.5 ± 5.7 184.0 ± 23.5 VD 5.0 ± 7.3 96.0 ± 21.9 664.0 ± 52.9 Pasteurella * 0 70 one hundred Control group III - IRT 5.7 ± 6.9 0 0 PG-3 5.4 ± 2.5 0 0 PC 2.6 ± 2.1 0 0 VD 5.9 ± 2.9 0 0 Pasteurella * 0 0 0 * Preventive activity of serum of vaccinated calves for white mice infected with 5 LD _{50 is} determined for pasteurells

The data in Table 9 indicate that immunization of the calves of both experimental groups with the vaccine of the invention leads to the formation of a pronounced immune response and a statistically significant increase in antibodies and the preventive activity of the blood serum of vaccinated animals (P <0.01). The maximum antibody content for all viruses is recorded on the 21st day after the second vaccination of each of the two test doses of the vaccine. The highest titer of neutralizing antibodies is detected in calves with the introduction of a vaccine in a dose of 2 cm ³ . Maximum preventive activity is also expressed in calves of the same experimental group.

Example 6. Evaluation of colostral immunity in newborn calves.

In the experiments, 7-14-day-old calves born to pregnant cows immunized with the combination vaccine of the invention are used, calves of a similar age obtained from unvaccinated cows are used as controls.

All experimental and control calves receive colostrum in a timely manner from their mothers during the first two hours after birth and are kept under the same conditions.

Assessment of colostral immunity is carried out by determining the titer of specific antibodies in the blood serum of all experimental and control animals and to pasteurella by the preventive activity of the blood serum of vaccinated calves for white mice infected with 5 LD ₅₀ virulent pasteurella strain.

The test results are presented in table 10.

Table 10 Evaluation of colostral immunity of a vaccine against infectious rhinotracheitis, parainfluenza-3, respiratory syncytial disease, viral diarrhea and cattle pasteurellosis in an experiment on calves Age of newborn calves, weeks Antigenic activity of a vaccine against viral antigens Immunogenic activity of the pasteurella vaccine IRT PG-3 PC VD Preventive activity of blood serum of vaccinated rabbits for white mice,% surviving animals Experienced group one 169.5 ± 25.6 155.3 ± 32.1 125.3 ± 31.1 340.8 ± 15.6 one hundred 2 125.6 ± 26.0 123.1 ± 24.9 115.3 ± 26.3 205.5 ± 29.7 70 four 85.7 ± 3.1 96.0 ± 21.9 76.0 ± 9.4 185.5 ± 20.7 fifty 6 42.5 ± 5.7 78.9 ± 19.6 53.7 ± 15.1 117.1 ± 17.9 40 Control group one 35.6 ± 9.8 42.9 ± 3.3 8.9 ± 2.6 16.9 ± 5.3 10 2 28.3 ± 4.2 18.3 ± 5.2 0 12.5 ± 1.1 0 four 0 0 0 0 0 6 0 0 0 0 0

As a result of the study, it was found that vaccination of pregnant cows with the studied vaccine provides the creation of colostral immunity in calves obtained from vaccinated animals and received colostrum in time from their mothers. The formation of colostral immunity is confirmed by high titers of specific antibodies to viral components and preventive activity against pasteurellae, which persisted throughout the observation period.

Thus, the experiments indicate that the inactivated combination vaccine against infectious rhinotracheitis, parainfluenza-3, respiratory syncytial disease, viral diarrhea and cattle pasteurellosis in accordance with the invention has a decisive advantage over other known vaccines in terms of specific protection and efficacy for susceptible group of animals, duration of intense immunity, harmlessness to pregnant animals and newborn calves, one hundred conservation properties of biological properties, including antigenic and immunogenic activity for 18 months (observation period). The vaccine is environmentally friendly, does not cause dissemination of pathogenic agents into the external environment.

Claims (3)

1. Inactivated combination vaccine against infectious rhinotracheitis, parainfluenza-3, respiratory syncytial disease, viral diarrhea and cattle pasteurellosis, containing the active substance and target additives, characterized in that it contains a mixture of an inactivated virus suspension of an infectious virus strain bovine rhinotracheitis STB No. 2333, family Herpesviridae, subfamily Alphaherpesviridae, inactivated viral suspension of the strain of parainfluenza virus-3 cattle about cattle bills ГВВ №2,334 of the subfamily Paramixovirinae, genus Paramixovims, inactivated viral suspension of the virus strain of respiratory syncytial disease of cattle ГКВ №2,335, subfamily Pneumoviridae, genus Pneumovirus, inactivated virus suspension of the strain of bovine diarrhea virus No. 6, cattle family Pestivirus, the activity of each of the viral strains of at least 10 ^6,5 TCD ₅₀ per 1 cm ³ of the vaccine and inactivated suspensions Pasteurella multocida serotype a strain of bacteria cells 1231 inactivated Pasteurella multocida gray suspension of bacterial cells ara B strain 656 cells inactivated suspensions of bacteria Pasteurella multocida serovar D strain T-80 strain suspension inactivated Mannheimia haemolytica bacteria cells 169, in an effective amount in the mixture taken in equal proportions at a concentration of not less than 3 billion microbial cells in the vaccine dose. 2. The vaccine according to claim 1, characterized in that it contains an adjuvant as a target additive. 3. The vaccine according to claim 2, characterized in that it contains saponin and / or aluminum hydroxide as an adjuvant.