RU2259845C1 - Inactivated fixed vaccine against infectious bird laryngotracheitis - Google Patents

Abstract

FIELD: veterinary medicine.

SUBSTANCE: vaccine has avirulent antigen material produced from O№112-DEP strain homological virus, protection medium and adjuvant taken in effective proportions. The strain is deposited in micro-organism VGNKI collection under registry number of O№112-DEP. The O№112-DEP strain virus is reproduceable in 9-11 days old SPF-hen embryos having infectious activity of at least 6.9 lg EID₅₀/cm³. The vaccine comprises ethylene imine dimer in 0.1% concentration as inactivation agent. The antigen material is mixed with protection medium after activation and subjected to lyophilization. Before being used, the dried antigen material is solved in adjuvant containing aerosyl, saponin, glycerol and phosphate buffer solution taken in effective proportions. The vaccine is intramuscularly introduced for immunizing clinically healthy 25-30 days old birds in the amount of 0.5 cm³. Immunity comes to a vaccinated bird at the fourteenth-twenty first day after vaccination.

EFFECT: high antigenic and immunogenic activity; no adverse side effects.

8 cl, 5 tbl

Description

The invention relates to the field of veterinary virology and biotechnology and can be used in the development and production of means of specific prophylaxis of infectious laryngotracheitis (ILT) of birds.

ILT is an acute contagious disease of birds, which causes a high percentage of mortality (usual mortality is 10-20%, there are cases of high mortality, which is 50-70%), in cases of ill individuals the daily gain and egg production are reduced. ILT is caused by herpes virus. One of the main means of combating ILT is vaccination with live and inactivated vaccines.

In world practice, more than 20 strains of the ILT virus are known that differ in their antigenic properties and are used in the manufacture of virus vaccines against bird ILT (1-7). In Russia, the VNIIBP strains (8) and the NT clone of the TsNIIPP strain (9) were widely used in the manufacture of virus vaccines against ILT.

Although the advantage of live vaccines due to the technological effectiveness of production and ease of use is not in doubt, the immune response in birds when they are administered is not always stable and in some cases causes complications after vaccination. In addition, with ILT, the administration of live vaccines to an adult bird does not form a strained immunity.

Inactivated emulsion vaccines, for the manufacture of which virulent strains of the homologous virus have also been used, have also been used for the prevention of bird ILT.

Known inactivated emulsion vaccine against bird ILT containing ethyleneimine inactivated antigenic material from the SZ-81-ILT strain of a homologous virus reproduced in 9-11-day-old SPF chicken embryos and Freund's incomplete adjuvant in the following ratio of ingredients, wt.%:

Antigenic material 50,0 Freund's Incomplete Adjuvant 50.0 (10).

The applicant did not find information on the use of inactivated adsorbed vaccines for the prevention of IFI in birds in accessible sources of information.

According to the applicant, the closest to the present invention in terms of essential features is an inactivated vaccine against bird ILT containing avirulent antigenic material from a strain of a homologous virus reproduced in a sensitive biological system and an adjuvant in an effective ratio (11).

The disadvantages of the known inactivated vaccines against ILT birds, including the vaccine prototype, are that for the preparation of antigenic material using virulent strains of the causative agent of the infection, which always create problems when observing veterinary and sanitary safety.

In this regard, the problem of creating an inactivated sorbed vaccine against bird ILT, which has both high antigenic and immunogenic activity and is harmless, remains urgent.

The objective of the present invention was to develop an inactivated sorbed vaccine against bird ILT based on a new strain of a homologous virus that has high biological, antigenic and immunogenic activity both natively and after inactivation, and is suitable for the manufacture of a highly immunogenic and harmless inactivated sorbed vaccine.

The technical result from the use of the present invention is to expand the arsenal of inactivated vaccines against ILT birds with high antigenic and immunogenic activity and harmlessness.

The specified technical result was achieved by creating an inactivated sorbed vaccine against bird ILT, characterized by the following set of features:

1) inactivated adsorbed vaccine against bird ILT;

2) avirulent antigenic material from a strain of a homologous virus reproduced in a sensitive biological system;

3) 9-11-day-old SPF-chicken embryos are used as a sensitive biological system;

4) from vaccine strains, the vaccine contains the strain "About No. 112-DEPT" of the bird ILT virus;

5) avirulent antigenic material from strain "About No. 112-DEPT" of a homologous virus reproduced in 9-11-day-old embryos of SPF chickens with infectious activity of at least 6.0 lg EID ₅₀ / cm ³ ;

6) avirulent antigenic material from strain "About No. 112-DEPT" of a homologous virus reproduced in 9-11-day-old embryos of SPF-chickens with infectious activity of at least 6.0 lg EID ₅₀ / cm ³ , in an amount of 13, 6-15.2 wt.%;

7) inactivant;

8) of inactivants - ethyleneimine dimer (DEI);

9) DEI at a concentration of 0.1%;

10) upon completion of inactivation, the DEI is neutralized with sodium thiosulfate;

11) protective environment;

12) a protective medium containing lactose and / or sucrose, and / or gelatose, in an amount of 0.8-2.4 wt.%;

13) adjuvant;

14) the adjuvant contains aerosil, saponin, glycerin and the FBI in the ratio, wt.%:

Aerosil 1.5-2.0 Saponin 0.007-0.009 Glycerol 23.0-27.0 FBI up to 100.0;

15) an adjuvant containing aerosil, saponin, glycerin and the FBI, in the amount of 84.0 wt.%;

16) avirulent antigenic material from strain "About No. 112-DEPT" of a homologous virus reproduced in 9-11-day-old embryos of SPF-chickens with infectious activity of at least 6.0 lg EID ₅₀ / cm ³ , a protective medium containing lactose and / or sucrose, and / or gelatose, and an adjuvant containing aerosil, saponin, glycerin and the FBI, in the ratio, wt.%:

Antigenic material 13.6-15.2 Protective environment 0.8-2.4 Adjuvant up to 100.0.

The present invention includes the following set of essential features that provide a technical result in all cases for which legal protection is sought:

1) inactivated adsorbed vaccine against bird ILT;

2) avirulent antigenic material from the strain "About No. 112-DEPT" of a homologous virus reproduced in a sensitive biological system in an effective amount;

3) Protective environment;

4) Adjuvant.

The present invention is also characterized by other features expressing specific forms of its implementation or special conditions of use:

1) avirulent antigenic material from strain "About No. 112-DEPT" of a homologous virus reproduced in 9-11-day-old embryos of SPF chickens with infectious activity of at least 6.0 lg EID ₅₀ / cm ³ ;

2) avirulent antigenic material from strain "About No. 112-DEPT" of a homologous virus reproduced in 9-11 day old embryos of SPF chickens with infectious activity of at least 6.0 lg EID ₅₀ / cm ³ , in the amount of 13.6 -15.2 wt.%;

3) a protective medium containing lactose and / or sucrose, and / or gelatose, in an amount of 0.8-2.4 wt.%;

4) the adjuvant contains aerosil, saponin, glycerin and the FBI in the ratio, wt.%:

Aerosil 1.5-2.0 Saponin 0.007-0.009 Glycerol 23.0-27.0 FBI up to 100.0.

5) an adjuvant containing aerosil, saponin, glycerin and the FBI, in an amount of 84.0 wt.%;

6) avirulent antigenic material from strain "No. 112-DEPT" of a homologous virus reproduced in 9-11-day-old embryos of SPF chickens with infectious activity of at least 6.0 lg EID ₅₀ / cm ³ , a protective medium containing lactose and / or sucrose, and / or gelatose, and an adjuvant containing aerosil, saponin, glycerin and the FBI, in the ratio, wt.%:

Antigenic material 13.6-15.2 Protective environment 0.8-2.4 Adjuvant up to 100.0.

The signs of the invention, characterizing the proposed vaccine and coinciding with the signs of the prototype, including the generic concept, reflecting the purpose, are:

1) inactivated adsorbed vaccine against bird ILT;

2) avirulent antigenic material from strain "About No. 112-DEPT" of a homologous virus reproduced in a sensitive biological system;

3) adjuvant.

Compared with the prototype vaccine, the salient features of the invention are:

1) avirulent antigenic material from a strain of homologous avian ILT virus;

2) avirulent antigenic material from the strain "About No. 112-DEPT" of the bird ILT virus in an effective amount.

The present invention is also characterized by other distinctive features expressing specific forms of its implementation or special conditions of use:

1) avirulent antigenic material from strain "About No. 112-DEPT" of a homologous virus reproduced in 9-11-day-old embryos of SPF chickens with infectious activity of at least 6.0 lg EID ₅₀ / cm ³ ;

2) avirulent antigenic material from strain "About No. 112-DEPT" of a homologous virus reproduced in 9-11-day-old embryos of SPF chickens with infectious activity of at least 6.0 lg EID ₅₀ / cm ³ , in an amount of 13, 6-15.2 wt.%;

3) a protective medium containing lactose and / or sucrose, and / or gelatose, in an amount of 0.8-2.4 wt.%;

4) the adjuvant contains aerosil, saponin, glycerin and the FBI in the ratio, wt.%:

Aerosil 1.5-2.0 Saponin 0.007-0.009 Glycerol 23.0-27.0 FBI up to 100.0;

5) the adjuvant containing aerosil, saponin, glycerin and the FBI, in the amount of 84.0 wt.%;

6) avirulent antigenic material from strain "About No. 112-DEPT" of a homologous virus reproduced in 9-11-day-old SPF chicken embryos with infectious activity of at least 6.0 lg EID ₅₀ / cm ³ , a protective medium containing lactose and / or sucrose, and / or gelatose, and an adjuvant containing aerosil, saponin, glycerin and the FBI, in the ratio, wt.%:

Antigenic material 13.6-15.2 Protective environment 0.8-2.4 Adjuvant up to 100.0.

The proposed vaccine has high antigenic and immunogenic activity, is harmless and expands the arsenal of inactivated vaccines against bird ILT.

The achievement of the technical result from the use of the proposed vaccine is explained by the use of avirulent antigenic material from the strain “About No. 112-DEP” of the bird ILT virus, which has higher antigenic and immunogenic activity and harmlessness compared to the strain VNIIBP and TsNIIPP as in native form, and after inactivation.

Strain "About No. 112-DEPT" is a new, previously unknown. The original virus (the author’s name is strain “O”) for obtaining strain “About No. 112-DEPT” was isolated from the larynx and trachea of chicken with signs of a latent form of the disease in Kazakhstan in 1992. The production strain "About No. 112-DEPT" was obtained by successive repeated passages on 9-11-day-old embryos of SPF hens.

The resulting strain was deposited in the All-Russian state collection of strains of microorganisms used in veterinary medicine and animal husbandry, the All-Russian State Research Institute of Control, Standardization and Certification of Veterinary Medicines (VGNKI) 05.06.2000, under the registration name "About No. 112-DEPT".

The possibility of its use for the manufacture of an inactivated sorbed vaccine against bird ILT has been experimentally confirmed.

The strain "About No. 112-DEPT" virus ILT birds is characterized by the following features and properties.

Morphological properties

The strain "About No. 112-DEPT" of the ILT virus belongs to the family Herpesviridae. Electron microscopy (instrumental magnification of 4.5 × 10 ⁵ and 5 × 10 ⁵ ) in the field of view of the microscope shows clearly visible viral particles of ILT with their characteristic morphology. Virions have a spherical shape, their sizes vary from 200 to 250 nm (some up to 330 nm). ILT virions consist of a spherical nucleocapsid surrounded by a complex-shaped supercapsid membrane. Virion has an icosahedral type of symmetry. The icosahedral capsid is coated with a lipoprotein membrane, which has on its surface morphological units-peplomeres. The diameter of the nucleocapsid is 113 ± 2 nm, the average diameter of the nucleocapsid shell is about 350 nm.

Nucleocapsids 100-110 nm in size are located in the viral particle, usually eccentric, less often in the center of the particles. The genome of the ILT virus has a length of 155,000 nucleotide pairs and consists of a long unique sequence U _L (120,000 bp) and a short unique sequence U _S (17,000 bp). The virus contains glycoproteins (205, 160, 115, 90, 85, 74, 60 and 50 kD). Glycoprotein B (gB) is the first structural protein of the ILT virus.

Antigenic properties

The strain "No. 112-DEPT" of the ILT virus is stably neutralized by homologous serum, is identified in the blood serum of chickens containing antibodies to this virus, in RDP, PH and ELISA. In RDP, it is identified by homologous hyperimmune sera in a titer of 1: 8. In pH, the neutralization index of homologous hyperimmune sera at dilutions of 1:10 and 1:20 averages 3.8 lg EID ₅₀ / cm ^3, respectively. In ELISA is identified by field and laboratory hyperimmune sera in the titer up to 1: 24579. Antigenic differences of the strain "About No. 112-DEPT" in relation to other strains of the ILT virus have not been established. The strain does not have hemagglutinating properties. Biotechnological characteristics

Strain "About No. 112-DEPT" shows high biological, immunogenic and antigenic activity both in the native form and after inactivation. Strain "About No. 112-DEPT" is used for the manufacture of live and inactivated vaccines against bird ILT and diagnostic preparations. The virus has a pronounced tropism to the epithelial cells of the mucous membrane of the larynx, trachea and cloaca of birds. The strain is cultured in 9-11 day old embryos of SPF hens. Reproduction of the virus occurs in the tissue of the chorioallantoic membrane (HAO). The infectious titer of the virus in a 10-20% suspension of HAO on the 5th day of cultivation under the condition of generalized distribution throughout the membrane is at least 6.2 lg EID ₅₀ / cm ³ .

Chemo- and genotaxonomic characterization

The strain "About No. 112-DEPT" is a DNA-containing virus.

Resistance to external factors

The virus is sensitive to lipolytic agents, heat and various disinfectants. A temperature of 55 ° C destroys the virus within 10-15 minutes, and 38 ° C in 48 hours. The virus remained active for more than 19 months in the frozen carcasses of sick and forcibly killed chickens stored at minus 10, 18 and 28 ° C, on the surface of the shell in thermostat it is inactivated after 12 hours, and in infected poultry houses it lasts no more than 9 days. Completely inactivated in a 1% solution of creole for 30 seconds. It retains its activity for a long time in a lyophilized state or at minus 20-60 ° C. In the lyophilized HAO homogenate, the virus remains infectious for at least 1.5 years. In the frozen HAO tissue, the infectivity of the virus persists for at least 4 years.

Immunogenicity of the strain

When vaccinating chickens older than 15 days of age, the ocular method at a dose of 1000 EID ₅₀ and the oral method (drink) at a dose of 4000 EID ₅₀ induces the production of specific antibodies and creates immunity against ILT in 100% of vaccinated chickens. In the lyophilized HAO homogenate, the virus retains immunogenicity for at least 1.5 years.

Epizooticity of the strain

When the vaccinated and intact birds are kept together, the transmission of the vaccine virus of strain “No. 112-DEPT” by contact has not been established.

Strain Maintenance Conditions

Lyophilized material from strain "About No. 112-DEPT" is stored at a temperature of 4 ° C, virus-containing HAO - at a temperature not exceeding minus 40 ° C. For lyophilization use a protective environment "ARRIAH". For resuspension of the lyophilized virus, an isotonic phosphate-buffered solution of pH 7.4-7.6 is used.

Genetic purity

After 20 passages on the Khao 9-11-day-old embryos of SPF-hens, no changes in the immunobiological properties of the strain "About No. 112-DEPT" were found.

Additional features and properties

There is no reactogenicity, no pathogenicity, no virulence, no oncogenicity, no contagiousness.

The attenuation stability was observed during 4 passages on sensitive chickens.

Based on the data obtained, it can be argued that the strain "About No. 112-DEPT" of the ILT virus according to antigenic and immunological spectra is an original, taxonomically new, previously unknown variant of the ILT virus.

The analysis of the prior art by the applicant, including a search by patent and scientific and technical sources of information and identification of sources containing information about analogues of the invention, allowed to establish that the applicant did not find sources characterized by signs that are identical (identical) to all the essential features of the invention. The definition from the list of identified analogues of the prototype, as the closest in the set of essential features of the analogue, made it possible to establish a set of essential distinctive features of the proposed vaccine relative to the applicant’s technical result, which are listed in the independent claim.

Therefore, the present invention meets the condition of patentability "novelty."

To verify the conformity of the proposed vaccine with the condition of patentability "inventive step", an additional search for known solutions was carried out to identify features included in the distinctive part of the independent claim. The search results showed that the proposed solution does not follow explicitly for the specialist from the prior art set forth in the corresponding section of the description (no solutions having signs matching the distinctive features of the proposed vaccine were identified), and the effect of the transformations provided for by the essential features of the proposed vaccine was not identified to achieve a technical result.

Therefore, the proposed vaccine meets the condition of patentability "inventive step".

The essence of the invention is illustrated by examples of its implementation, which do not limit its scope.

Example 1

An inactivated adsorbed vaccine against bird ILT is prepared as follows.

Well-developed 9-11-day-old SPF-chicken embryos are infected with the mother virus from strain “About No. 112-DEPT”. For this, a hole with a diameter of 8-10 mm is formed in the shell of the chicken embryo from the side of the air chamber. Four punctures in the corners of the imaginary square with a diagonal of about 5 mm are made in the shell-shell and chorioallantoic membrane (CAO) with an injection needle. After that, virus-containing material in a volume of 0.1 cm ³ is applied to the HAO. The hole in the shell is sealed with adhesive tape. Infected embryos are incubated for 3-6 days at 37.7 ° C and a humidity of 60%. According to the results of ovoscopy and subsequent opening of the embryos, the presence of HAO lesions characteristic of the ILT virus is determined. Affected HAO is used as a virus-containing material. A 15-30% suspension is prepared from the affected HAO, which is frozen and freed from coarse particles by low-speed centrifugation. The virus-containing supernatant is carefully poured under sterile conditions into a common container and subjected to control for infectious activity. For the manufacture of the vaccine using material with a titer of infectious activity of at least 6.0 lg EID ₅₀ / cm ³ . Then the resulting virus is subjected to inactivation. For inactivation using DEI. A 10% aqueous solution of DEI is added to a virus-containing suspension heated to 27 ° C to a concentration of 0.1%. The mixture is thoroughly mixed and kept at 27 ° C for 24 hours. At the end of the inactivation, the DEI is neutralized with sodium thiosulfate for 120 minutes and the antigenic material is cooled to a temperature of 2 ÷ 8 ° C. A protective medium is added to the resulting antigen, which can be used alone or in combination with lactose and / or sucrose and / or gelatose. The antigen and protective environment are mixed in the ratio, wt.%: 95: 5 ÷ 85: 15. The resulting mixture is packaged in bottles of 4 cm ³ , frozen at a temperature of minus 48 ÷ 50 ° C and subjected to lyophilization. The drying time is 59-68 hours. The resulting semi-finished product is a homogeneous finely porous mass in the form of a beige cylindrical tablet. The drug is stored in lyophilized form at a temperature of 2 ÷ 6 ° C until use.

An adjuvant is prepared by mixing Aerosil, saponin, glycerin and the FBI in the following ratio of these ingredients, wt.%:

Aerosil 1.5-2.0 Saponin 0.007-0.009 Glycerol 23.0-27.0 FBI up to 100.0.

Ready adjuvant has the form of an opalescent liquid with a loose precipitate of white color (about 1/5 of the volume). When the bottle is shaken, a homogeneous suspension is formed with slight foaming.

To obtain the vaccine, the lyophilized prefabricated product (initial volume of 4 cm ³ ) is rehydrated in 25 cm ^{3 of} adjuvant. After mixing with adjuvant, the vaccine becomes pink. The resulting vaccine is monitored for safety, antigenic and immunogenic activity. The vaccine is used in clinically healthy birds aged 25-30 days to create a high level of immunity. The drug in a volume of 0.5 cm ³ is administered intramuscularly. Immunity in a vaccinated bird occurs on 14-21 days after vaccination. The vaccine induces a high level of antibodies against the virulent avian ILT virus.

The optimal component composition of the obtained vaccine against bird ILT is shown in table 1.

According to table 1, the antigen content in the vaccine within the specified limits is its effective amount in the drug, ensuring the achievement of a technical result.

Example 2

Inactivated sorbed vaccine against bird ILT, made as described in example 1, and containing, wt.%:

Avirulent Antigenic material from the strain "About No. 112-DEPT" homologous virus 14,4 Protective environment 1,6 Adjuvant up to 100.0

subjected to reactogenicity control. For this, 6 cm ³ vaccines were taken from 3 vials, forming a common sample. The vaccine was administered to 10 chickens intramuscularly in the chest area of 1.5 cm ³ . The bird was monitored daily for 10 days. The effect of the drug did not entail a death or clinical signs of ILT. During the observation period (10 days), no toxic effect or manifestation of clinical signs of the disease was detected.

Example 3

Inactivated sorbed vaccine against bird ILT, made as described in example 1, and containing, wt.%:

Avirulent Antigenic material from the strain "About No. 112-DEPT" homologous virus 14,4 Protective environment 1,6 Adjuvant up to 100.0

subjected to control for antigenic and immunogenic activity. For this, from 6 vials of a vaccine containing antigenic material at a concentration of 14.4%, 6 cm ^{3 of the} drug were taken, forming a single sample. In addition, samples were prepared comprising 1/4 and 1/16 of the above antigen concentration. Samples were indicated: "C" - whole, "1/4" - and "1/16", respectively. To test these drugs, 3 groups of 15 experimental birds were formed. The vaccine was administered intramuscularly into the chest area at 0.5 cm ³ . In parallel, a control group was formed, consisting of 10 intact birds, which were kept under conditions similar to the experimental bird during the entire experiment.

On the 21st day after vaccination in vaccinated chickens in ELISA, serum was examined for the presence of antibodies to the ILT virus. For ELISA, KPL diagnostic kits were used (USA, Catalog No. 54-93-01; Serial No. F 032). In accordance with the instructions for using the kits, the reaction was considered positive if the antibody titer was equal to or greater than 1: 336.

21 days after vaccination, the experimental and control birds were infected with the pathogenic strain “Bogatishchevsky” of the ILT virus, which was administered intratracheally at a dose of 1000 EID ₅₀ in a volume of 0.5 cm ³ . An infected bird was monitored for 10 days. Clinical signs of ILT were detected. Pathological studies were performed (specific lesions for ILT were established: exudative-hemorrhagic inflammation of the mucosa of the trachea and pharynx, as well as the presence of fibrinous plugs in the upper part of the trachea). In the experimental and control groups, the proportion of birds that fell and died from ILT was determined.

The results of the study of the antigenic and protective effect of the sorbed vaccine in the form of titer of serum antibodies, as well as the proportion of sick and dead birds after infection with a pathogenic strain of the ILT virus, respectively, the tested antigen concentrations in the preparations are presented in table 2.

The results presented in table 2 suggest that the sorbed vaccine showed a pronounced antigenic and protective effect, which depended on the concentration of antigen in the drug.

It was found that the incidence rate of vaccinated birds (C) during control of infection with a pathogenic strain of the virus was stably associated with the proportion of positively reacting sera in the study group. The value of the correlation coefficient of these variables was 0.908 for the sorbed vaccine.

Analysis of the results showed that the relationship between the estimates of D and T can be described by an empirical equation that allows you to determine the expected value of antibody titer (T) for a given concentration of antigen in the vaccine. The equation for the sorbed vaccine was:

logT '= 3.584 + (- 0.722) × log D

The established values of the empirical coefficients had no static differences. After using the sorbed vaccine, the experimental and expected antibody titers were high.

An estimate of the protective effect of vaccines for a given antigen concentration was considered to be P = 1-C (the proportion of unwell birds). Moreover, focusing on obtaining more cautious resulting estimates, the experimental values of C = 0 were taken as C = 0.01. To describe the relationship between the concentration of antigen (D) and the expected protective effect of the vaccine, the established estimates of P were expressed as their corresponding normal equivalent deviations (Y is the tabular value). Thus, the equation for the sorbed vaccine was constructed

logY '= 2,192 + (- 2,146) × logD,

where Y 'is the predicted value of the normal equivalent deviation of the proportion of unwell birds for a given value D.

This equation made it possible to determine that, to protect 50% of the vaccinated poultry head, the expected antigen concentration (dilution value of the whole preparation) should be D ' _sb = 10.76 in the sorbed vaccine. Moreover, for the protection of 95% of birds, this value was D ' _95copb = 1.82. The whole preparation of this vaccine (D = 1) had an expected protection rating of about 99%.

Thus, the sorbed vaccine showed a pronounced antigenic and protective effect. The relationship between the concentration of antigen and the proportion of protected birds, as well as the average titer of serum antibodies, was established and quantified. In an experiment, a vaccine containing a whole antigen determined the protection of 100% of the birds. The mean antibody titers in these groups were 1: 3981 for the sorbed vaccine.

Example 4

Studies of changes in poultry muscle tissue at the injection site of an inactivated sorbed vaccine against bird ILT from strain O No. 112-DEPT (in the amount of the immunizing dose) 30, 40, 60 and 90 days after vaccination were studied. For this, a group of 32 animals was formed from the number of birds vaccinated but not sick after the control infection. After vaccination of an experimental vaccine in an experimental bird, no visible changes were detected at the injection site after 30 days.

Example 5

Tests of the immunogenic activity of an inactivated sorbed vaccine against bird ILT, made as described in example 1, and containing, wt.%:

Avirulent Antigenic material from the strain "About No. 112-DEPT" bird ILT virus 13.6 Protective environment 2,4 Adjuvant up to 100.0.

The immunogenic activity of the vaccine was determined on chickens of 30 days of age, free of antibodies to the ILT virus of birds. The vaccine was administered at 0.5 cm ³ into the thigh muscle. After 21 days, the birds took blood, received serum and examined them in the ELISA. The research results are presented in table 3.

The results of table 3 indicate the high immunogenic activity of the inactivated sorbed vaccine against bird ILT, made from antigenic material from strain "About No. 112-DEPT".

Example 6

Tests of the immunogenic activity of an inactivated sorbed vaccine against bird ILT, made as described in example 1, and containing, wt.%:

Avirulent Antigenic material from the strain "About No. 112-DEPT" bird ILT virus 14,4 Protective environment 1,6 Adjuvant up to 100.0.

The immunogenic activity of the vaccine was determined on chickens 25-30 days old, free of antibodies to the ILT virus of birds. The vaccine was administered at 0.5 cm ³ into the thigh muscle. 21 days after vaccination, blood was collected from birds, sera were obtained and examined in ELISA for the presence of antibodies. The research results are presented in table 4.

Example 7

Tests of the immunogenic activity of an inactivated sorbed vaccine against bird ILT, made as described in example 1, and containing, wt.%:

Avirulent Antigenic material from the strain "About No. 112-DEPT" bird ILT virus 15,2 Protective environment 0.8 Adjuvant until 100.0

The immunogenic activity of the vaccine was determined on chickens 25-30 days old, free of antibodies to the ILT virus of birds. The vaccine was administered at 0.5 cm ³ into the thigh muscle. 21 days after vaccination, blood was collected from birds, sera were obtained and examined in ELISA for the presence of antibodies. The research results are presented in table 5.

The results of table 5 indicate the high immunogenic activity of the inactivated sorbed vaccine from strain "About No. 112-DEPT" of the bird ILT virus.

Thus, the above information indicates the fulfillment when using the invention of the following set of conditions:

- inactivated sorbed vaccine against bird ILT, embodying the invention, is intended for use in agriculture, namely in veterinary virology and biotechnology;

- for the proposed invention in the form as described in the independent claim, the possibility of its implementation using the means and methods described in the application or known prior to the priority date is confirmed;

- inactivated sorbed vaccine against bird ILT, prepared from strain "About No. 112-DEPT" in accordance with the invention, has a high antigenic and immunogenic activity and is harmless.

Therefore, the present invention meets the condition of patentability "industrial applicability".

Sources of information

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4. Auth. testimonial. CCCH No. 1750683, A 61 K 39/245, 07/30/92.

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6. Syurin V.N., Samuilenko A.Ya. and other viral diseases of animals // M. - VNITIBP, - 1998. - S.672-683.

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8. Malushko V.V., Sokolov V.D. et al. Experience in combating infectious laryngotracheitis // Poultry farming. - 1983. - 12. - S.21-23.

9. Dutko Yu.S., Shevchenko A.A. and other Immunological efficacy of a vaccine against infectious laryngotracheitis of hens // Veterinary medicine. - 1991. - 3. - P.32-34.

10. Barhoom S., Forgacs A. and Solyom F. Development of an inactivated vaccine against infectious laryngotracheitis (ILT) - serological and protection studies // Avian Pathology. - 1986. - V.15, No. 2. - P.213-221.

11. Babkin V.F. Infectious laryngotracheitis of birds (Development of inactivated vaccines, diagnostic methods and systems of antiepizootic measures): Author. dis. Doct. vet. sciences. - Kharkiv. - 1996. - 33 pp. (In Ukrainian) (prototype).

Table 1
The optimal component composition of the inactivated sorbed vaccine against bird ILT from strain "About No. 112-DEPT" Ingredients Minimum, wt.% Average, wt.% Maximum, wt.% Avirulent antigenic material from strain "About No. 112-DEPT" of the bird ILT virus 13.6 14,4 15,2 Protective environment 2,4 1,6 0.8 Adjuvant 84.0 84.0 84.0

table 2
Antigenic and protective effect of an inactivated sorbed vaccine against bird ILT from strain "No. 112-DEPT" containing various concentrations of antigen Values of titers of serum antibodies, their average scores (T), as well as the proportion of sick (C) and fallen (L) birds after control infection with ILT, respectively, antigen concentration Antigen Concentration (D) The control C (D = 1) 1/4 (D = 4) 1/16 (D = 16) 4476 1321 509 109 {-} 2548 826 98 {-} - 8128 1346 - - 6427 241 {-} 359 54 {-} 3040 2705 896 46 {-} 5296 991 - - 2301 557 134 {-} 17 {-} 4645 2450 b4 {-} - 2051 75 {-} - - 3097 826 872 - 6950 2301 - 203 {-} 4295 - - - 3372 306 {-} 304 {-} 74 {-} 2523 1953 457 32 {-} 5887 636 - - T = 3981 T = 1278 T = 580 T = 0 C = 0/15 C = 4/15 C = 9/15 C = 15/15 L = 0 L = 1/4 L = 3/9 L = 8/15

Table 3
Immunogenicity of inactivated sorbed vaccine against bird ILT from strain "O No. 112-DEPT" №№ Name of material ELISA antibody titer 1. Vaccinated Bird Serum 2600 2. Vaccinated Bird Serum 3850 3. Vaccinated Bird Serum 4073 4. Vaccinated Bird Serum 4645 5. Vaccinated Bird Serum 2380 6. Vaccinated Bird Serum 3020 7. Vaccinated Bird Serum 2700 8. Vaccinated Bird Serum 3200 9. Vaccinated Bird Serum 2950 10. Vaccinated Bird Serum 4500 eleven. The control 0

Table 4
Immunogenic activity of inactivated sorbed vaccine against bird ILT from strain "O No. 112-DEPT" №№ Name of material ELISA antibody titer 1. Serum vaccines. birds 3550 2. Serum vaccines. birds 3400 3. Serum vaccines. birds 2950 4. Serum vaccines. birds 4350 5. Serum vaccines. birds 3730 6. Serum vaccines. birds 2900 7. Serum vaccines. birds 3200 8. Serum vaccines. birds 3650 9. Serum vaccines. birds 4050 10. Serum vaccines. birds 3800 eleven. The control 0

Table 5
Immunogenic activity of inactivated sorbed vaccine against bird ILT from strain "O No. 112-DEPT" №№ Name of material ELISA antibody titer 1. Vaccine Serum 4200 2. Vaccine Serum 3850 3. Vaccine Serum 4050 4. Vaccine Serum 3600 5. Vaccine Serum 2890 6. Vaccine Serum 3630 7. Vaccine Serum 3850 8. Vaccine Serum 2990 9. Vaccine Serum 4100 10. Vaccine Serum 3100 eleven. The control 0

Claims (8)

1. Inactivated sorbed vaccine against infectious laryngotracheitis of birds, containing avirulent antigenic material from a strain of a homologous virus reproduced in a sensitive biological system, and an adjuvant, characterized in that as avirulent antigenic material it contains avirulent antigenic material from a strain of the virus of infectious laryngitis, . Herpesviridae, genus Herpesvirus, collection VGNKI "About No. 112-DEPT", in an effective amount. 2. The vaccine according to claim 1, characterized in that it contains avirulent antigenic material from strain "About No. 112-DEPT" homologous virus reproduced in 9-11-day-old embryos of SPF-chickens with infectious activity of at least 6, 0 lg EID ₅₀ / cm ³ . 3. The vaccine according to claim 2, characterized in that it contains avirulent antigenic material from strain "About No. 112-DEPT" homologous virus in an amount of 13.6-15.2 wt.%. 4. The vaccine according to claim 1, characterized in that it additionally contains a protective environment. 5. The vaccine according to claim 4, characterized in that it contains a protective medium containing lactose and / or sucrose, and / or gelatose in an amount of 0.8-2.4 wt.%. 6. The vaccine according to claim 1, characterized in that it contains an adjuvant containing aerosil, saponin, glycerin and phosphate-buffer solution in the ratio, wt.%: Aerosil 1.5-2.0 Saponin 0.007-0.009 Glycerol 23.0-27.0 Phosphate buffered saline Up to 100.0 7. The vaccine according to claim 1, characterized in that it contains an adjuvant in an amount of 84.0 wt.%. 8. The vaccine according to any one of claims 1 to 4, characterized in that it contains avirulent antigenic material from strain "About No. 112-DEPT" of a homologous virus, a protective environment and an adjuvant in the ratio, wt.%: Antigenic material 13.6-15.2 Protective environment 0.8-2.4 Adjuvant Up to 100.0