RU2199342C1 - Method for preparing inactivated vaccine against hen's hydropericarditis syndrome - Google Patents

Abstract

FIELD: veterinary virology, biotechnology. SUBSTANCE: method deals with preparing antigenic material as 10-20% virus-containing suspension made using either the liver of 5-50-d-aged chickens infected with HHPS virus from KR95N114-DEP strain (VGNKI Institute collection) or dead ones with HHPS signs. Virus is cultivated in chicken's body for 48- 120 h. To prepare vaccine one should use virus at its biological activity being 3,25-5,0 lgLD₅₀/0,2cm³. After purification necessary to remove ballast admixtures due to well-known technique one should add ethylenimine dimer into virus-containing suspension up to 0.15-0.4% for virus inactivation. Purified and avirulent antigenic material should be combined with adjuvants due to aluminum hydroxide and saponin. Ready-to-use product undergoes control in accordance to Technical Conditions. The method provides higher antigenic and immunogenic activity and harmlessness of the target product. EFFECT: higher efficiency. 13 cl, 4 ex, 6 tbl

Description

 The invention relates to the field of veterinary virology and biotechnology and can be used in the manufacture of agents for the specific prophylaxis of chicken hydropericarditis syndrome (HHPC).

 CGPC (hepatitis with inclusion bodies, Angara disease, Lychee disease) is an acute infectious disease of hens of viral etiology, causing significant economic damage to the developed poultry industry in many countries of the world.

 The hepatitis C virus virus causes a disease of chickens mainly aged 10 to 50 days, which is characterized by sudden death of the bird, accumulation of excess fluid in the pericardial cavity, damage to the liver and kidneys, depression, diarrhea, and in some cases anemia.

 In August 1987, in Pakistan, a disease with signs of hepatitis with inclusion bodies and heart damage (hydropericarditis) was recorded at a broiler farm in Angara Goth near Karachi in Pakistan. The disease was noted, as a rule, in birds of 3-6 weeks of age, but sometimes it was found in younger or older birds, mortality reached 50-70%. Initially, the disease was given the name "Angara disease" in accordance with the geographical area where the outbreak was first recorded. Later, the disease was given the name "hydropericarditis syndrome" due to the fact that when autopsies of birds were observed accumulation of fluid (up to 20 ml) in the cardiac chemise due to impaired blood circulation in the cardiovascular system. A similar disease has been reported in Iraq, India, Mexico, Kuwait, Australia, Japan, and Chile (1).

 Since the early 90's. outbreaks of high-mortality SGPKs are recorded in Russia (2). The lack of diagnostic tools and specific prophylaxis of SGPK in domestic veterinary practice has caused significant economic damage to poultry farms. The economic losses incurred by poultry farms from outbreaks of agro-agricultural complex are made up of the death of birds (from 10 to 60%), the death of developing chicken embryos during incubation, reduced productivity and payment of feed, as well as funds spent on unscheduled veterinary and sanitary measures et al. (3). Vaccine prophylaxis of CGD takes a leading place in the fight against this disease. Both live and inactivated vaccines are used to prevent CGD. In recent years, inactivated vaccines have been most commonly used (4). However, the technology of their manufacture has drawbacks, the elimination of which remains an urgent task and serves as the main prerequisite for research on improving the technology of manufacturing an inactivated vaccine against SGPK and solving the issue of protecting the poultry industry of our country from this dangerous disease.

The closest to the invention according to the totality of features is a method of manufacturing an inactivated vaccine against CHPA, comprising preparing an antigenic material in the form of a virus-containing suspension from the liver of chickens infected with the pathogen and falling with signs of CHPP, cleaning the antigenic material from ballast impurities by centrifugation at 2500-3000 rpm within 15-20 minutes, inactivation of the virus with formaldehyde for 24 hours at 4 ^o With periodic stirring and subsequent monitoring of the target product ( 5, 6).

The disadvantages of the prototype method are that
- to obtain antigenic material, the liver of chickens is used, in which the antigen content often varies greatly, and it could also be contaminated with pathogens of other infectious diseases of birds (Gamboro disease, Marek's disease, infectious bronchitis, infectious anemia of hens, etc.);
- to clean the virus-containing suspension from cell detritus, the method of low-speed centrifugation is used, which does not sufficiently purify the virus-containing suspension from ballast impurities;
- formaldehyde is used as an inactivant, causing destructive changes in viral proteins and reducing the immunogenic activity of the vaccine. The probability of incomplete inactivation of the virus during formaldehyde treatment of a viral suspension containing a significant amount of ballast protein is also high;
- this method does not provide for the introduction of an adjuvant into the preparation, the use of which improves storage and increases the antigenic activity of the vaccine.

 The task of creating the present invention was the development of a method of manufacturing an inactivated vaccine against SGPK with high immunogenic activity and harmlessness and protecting the poultry industry of the Russian Federation from epizootic virus SGPK, causing significant economic damage.

 The technical result from the use of the invention is to increase the antigenic and immunogenic activity and harmlessness of the inactivated vaccine against SGPK.

The specified technical result from the use of the invention was achieved by creating a method for the manufacture of an inactivated vaccine against SGPK, characterized by the following set of features:
1. A method of manufacturing an inactivated vaccine against SGPK.

 2. Preparation of antigenic material in the form of a 10-20% virus-containing suspension from the liver of chickens infected with an infectious agent and who died with signs of CGD.

 3. As the causative agent of the infection, strain KP95 114 of the SGPV virus is used in an effective amount.

 4. To obtain antigenic material use the liver of 5-50-day-old chickens.

 5. The virus is cultured in the body of 5-50-day-old chickens for 48-120 hours

6. To prepare the vaccine using antigenic material with biological activity of 3.25-5.0 Ig LD ₅₀ / 0.2 cm ² .

 7. The resulting antigenic material is subjected to purification from ballast impurities.

 8. The purified antigenic material is inactivated.

 9. As an inactivating agent, an ethyleneimine dimer (DEI) is used.

 10. DEI is used as a 10% aqueous solution.

 11. DEI contribute to the antigenic material to a concentration of 0.15-0.4%.

12. The inactivation of the HCGV virus is carried out for 23-25 hours at 35-36 ^o C and a pH of 7.4-7.6.

 13. The combination of purified and avirulent antigenic material with adjuvant.

 14. Of the adjuvants, aluminum hydroxide (GOA) is used.

 15. GOA is used in the form of a 3-4% colloidal solution.

 16. GOA contribute to the antigenic material to a concentration of 0.1%.

 17. Of the adjuvants, saponin is additionally used.

 18. Saponin is used as a 10% aqueous solution.

 19. Saponin is introduced into antigenic material to a concentration of 0.5-2%.

 20. Control of the target product.

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. A method of manufacturing an inactivated vaccine against SGPK.

 2. Preparation of antigenic material in the form of a virus-containing suspension from the liver of chickens infected with an infectious agent and who died with signs of CGD.

 3. In the sway of the causative agent of the infection, the strain KP95 114 of the HCGP virus is used in an effective amount.

 4. Purification of antigenic material from ballast impurities.

 5. Inactivation of antigenic material.

 6. The connection of purified and avirulent antigenic material with adjuvant.

 7. Control of the target product.

The present invention is also characterized by other features expressing specific forms of execution or special conditions for its use:
1. To obtain antigenic material use the liver of 5-50-day-old chickens.

 2. The strain KP95 114 virus SGPK cultured in the body of 5-50-day-old chickens for 48-120 hours

3. For the manufacture of the vaccine using antigenic material from strain KP95 114 with biological activity of 3.25-5.0 Ig LD ₅₀ / 0.2 cm ² .

 4. Purified antigenic material inactivate DEI.

 5. DEI is used as a 10% aqueous solution.

 6. DEI contribute to the antigenic material to a concentration of 0.15-0.4%.

7. Inactivation of antigenic material is carried out for 23-25 hours at 35-36 ^o C and a pH of 7.4-7.6.

 8. Of the adjuvants, GOA is used.

 9. GOA is used in the form of a 3-4% colloidal solution.

 10. GOA contribute to the antigenic material to a concentration of 0.1%.

 11. Of the adjuvants, saponin is additionally used.

 12. Saponin is used as a 10% aqueous solution.

 13. Saponin is introduced into the antigenic material to a concentration of 0.5-2%.

The signs of the invention, characterizing the proposed method and coinciding with the signs of the prototype, including the generic concept, reflecting the purpose, are:
1. A method of manufacturing an inactivated vaccine against SGPK.

 2. Obtaining antigenic material in the form of a virus-containing suspension of their liver in chickens infected with an infectious agent and who died with signs of CGD.

 3. Purification of antigenic material from ballast impurities.

 4. Inactivation of the virus.

 5. Control of the target product.

Compared with the prototype method, the salient features of the invention are:
1. As the causative agent of the infection using strain KP95 114-DEPT virus SGPK.

 2. The strain KP95 114 virus CGPC used in an effective amount.

 3. The connection of the purified avirulent antigenic material with adjuvant.

The present invention is also characterized by other distinctive features that express specific forms of execution or special conditions for its use:
1. To obtain antigenic material use the liver of 5-50-day-old chickens.

 2. The strain KP95 114 virus SGPK cultured in the body of 5-50-day-old chickens for 48-120 hours

3. For the manufacture of the vaccine using antigenic material from strain KP95 114 with biological activity of 3.25-5.0 Ig LD ₅₀ / 0.2 cm ² .

 4. Purified antigenic material inactivates DEI.

 5. DEI is used as a 10% aqueous solution.

 6. DEI contribute to the antigenic material to a concentration of 0.15-0.4%.

7. Inactivation of antigenic material is carried out for 23-25 hours at 35-36 ^o C and a pH of 7.4-7.6.

 8. Of the adjuvants, GOA is used.

 9. GOA is used in the form of a 3-4% colloidal solution.

 10. GOA contribute to the antigenic material to a concentration of 0.1%.

 11. Of the adjuvants, saponin is additionally used.

 12. Saponin is used as a 10% aqueous solution.

 13. Saponin is introduced into the antigenic material to a concentration of 0.5-2%.

 The vaccine obtained by the proposed method, in comparison with the vaccine obtained by the prototype method, has a higher antigenic and immunogenic activity and is harmless, provides protection against the causative agent of SGPK circulating on the livestock of birds in the Russian Federation.

 The achievement of the technical result from the use of the proposed method is explained by the fact that in the manufacture of the vaccine using a new strain KP95 114 virus SGPK.

 An additional technical result from the use of the proposed method is achieved due to the fact that the inactivation of antigenic material is carried out by DEI, which can significantly reduce labor and energy costs for the manufacture of vaccines and improve the quality of antigenic material.

 An additional technical result from the use of the proposed method is achieved through the use of adjuvants GOA and saponin.

 The original virus for obtaining strain KP95 114 was isolated from the liver of sick broilers at the Krasnoyarskaya poultry farm in the Krasnoyarsk Territory in 1995. Production strain KP95 114 was obtained by repeated successive passages on chickens.

 Strain KP95 114 was deposited in the State Collection of Microorganisms of the All-Russian Scientific Research Institute for Control, Standardization and Certification of Veterinary Medicines (VGNKI) on December 14, 2000 under registration KP95 114-DEP.

 The strain is virulent and has high antigenic and immunogenic activity after inactivation. The possibility of its use for the manufacture of inactivated vaccines has been experimentally confirmed.

 Strain KP95 114 of the HCVV virus is characterized by the following features and properties.

Morphological properties
The strain KP95 114 of the SGPV virus belongs to the family Adenoviridae, genus Aviadenovirus, group 1, serotype 4, has morphological characteristics characteristic of adenoviruses: like all other viruses of this family lacking a supercapsid coat, the diameter of the virion of strain KP95 114 and nucleocapsid are equal. The size of the virion is 75-80 nm. Virion has an icosahedral type of symmetry. The virion is represented by a nucleoid consisting of nucleic acid and core proteins and a capsid protein coat. The capsid contains 252 capsomeres, which are located in 20 facets having the shape of equilateral triangles. On the edges of equilateral triangular faces common to neighboring triangles, there are six capsomeres; the rib length is about 42 nm. Each of the 240 capsomeres of the capsid shell forming equilateral triangles is surrounded by six of the same capsomeres and is called hexon. Each of the 12 capsomeres located at the tops of the icosahedron is surrounded by only five neighboring capsomeres and is called peptone. Each capsomer (except for a complex peptone) is a prismatic particle with a diameter of 6-7 nm with a central cavity of 1.5-2 nm in size. The method of laying capsomeres in a capsid is unique for viruses with an icosahedral type of symmetry and differs in geometric ordering. Hexons are the main soluble component of the pathogen during its reproduction in sensitive cells.

 Peptones are associated with their ability to agglutinate red blood cells, due to the presence of complete hemagglutin in their composition.

 The core of the virion contains a double-stranded DNA molecule and two internal proteins. DNA contains about 60 genes that carry information that is sufficient to encode from 30 to 50 proteins.

Antigenic properties
By its antigenic properties, strain KP95 114 belongs to group 1 of avian adenoviruses. The virus is stably neutralized by homologous antiserum. The virus does not exhibit hemagglutinating activity. When vaccinated, the viral antigen induces the formation of specific antibodies detected in the RDP at a dilution of 1: 2-1: 64.

Biotechnological characteristics
Strain KP95 114 is virulent and has high biological activity in its native form, and also exhibits high antigenic and immunogenic properties after inactivation.

Strain KP95 114 is intended for use as a raw material for the manufacture of inactivated vaccines, as well as diagnostic biologics. Strain KP95 114 is reproduced in chickens 5-90 days old. Within 48-120 hours of incubation, the virus accumulates in the liver of chickens in a titer of 3.25-5.0 Ig LD ₅₀ / 0.2 ml. Strain KP95 114 is stable.

Chemo- and genotaxonomic characterization
The core of the virion contains a double-stranded DNA molecule and two internal proteins. The nucleic acid mass reaches 17.3% of the mass of the virion.

The bulk of the proteins of the HCV virus are concentrated in the capsid shell of the virion. Core proteins account for up to 20% of all virus proteins. Core protein "1" with a molecular weight of 46 • 10 ³ g / mol is easily separated from the internal nucleoprotein containing DNA in complex with arginine-rich core protein "2".

Hexon, peptone base and its filament (fiber) of the K95 114 virus are constructed from polypeptides with molecular weights of 120 • 10 ³ , 70 • 10 ³ , 62 • 10 ³ g / mol, respectively.

 The protein structures of the virus strain KP95 114 form several antigens. In the structure of the structural viral antigen, type-specific, group-specific, subgroup-specific antigens of hexon polypeptides, peptone bases and subgroup-specific and type-specific antigens of peptone filament were found in them.

The structure of the structural viral antigen revealed three main components, designated A, B and C. Antigen A is structurally associated with hexon, antigen B with peptone, antigen C with virion filaments (fiber). Hexon-linked component A contains a group-specific antigen (α) and a type-specific antigen (ε). The antigenic factor associated with peptone is called the antigen (β), and with the filament of the virion (fiber) - (γ).
Physical properties
The mass of the virion is 252 • 10 ³ g / mol. Floating density 1.32-1.35 g / ml in a gradient of C _s Cl. The sedimentation constant of the mature virion is 810 • 10 ^-13 -910 • 10 ^-13 s.

Resistance to external factors
Strain KP95 114 is resistant to ether, chloroform, saponin and sodium deoxycholate, is sensitive to formaldehyde, β-propiolactone, hydroxylamine, ethyleneimine derivatives, UV radiation, and γ-radiation.

 Strain KP95 114 is quite resistant to heat and to a change in the pH of the medium over a wide range, resistant to 0.25% trypsin solution, 2% phenol solution and 50% ethyl alcohol solution. Absolute ethanol, like its iodine solution, causes complete inactivation of the virus.

 Strain KP95 114 is resistant to repeated cycles of freezing and thawing, does not lose infectious properties during freeze-drying.

Additional features and properties
Immunogenic activity -100%
Pathogenicity is expressed
Virulence expressed
Contagiousness expressed
No oncogenicity
Based on the data obtained, it can be argued that strain KP95 114 in terms of antigenic and immunological spectra is an original, taxonomically new, previously unknown variant of the CGD virus. To reduce its epizootic danger, timely prevention of newly emerging foci of the disease is necessary, and for this a highly active and harmless vaccine is needed.

 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 existing signs of the invention. The definition from the list of identified analogues of the prototype as the closest in the totality of the features of the analogue made it possible to establish a set of essential distinguishing features of the proposed method with respect to the technical result perceived by the applicant, as set forth in the independent claim.

 Therefore, the proposed method meets the condition of patentability "novelty."

 To verify the conformity of the proposed solution to the patentability condition "inventive step", an additional search was carried out for known solutions 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 features matching the distinguishing features of the present invention have been identified), and the effect of the transformations provided for by the essential features of the proposed invention has not been identified to achieve a technical result.

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

 The essence of the invention is illustrated by examples of its implementation.

 Example 1

To obtain a vaccine from strain КР95, 5–50-day-old chickens obtained from farms that are successful in infectious diseases are infected intramuscularly in a volume of 1 cm ³ with 114 matrovirus. Chickens contain 120 hours after infection. Livers are taken from sick and dead chickens. The liver is subjected to grinding in a colloidal mill for 10-12 minutes, then the virus-containing mass is mixed with phosphate buffer solution, resulting in a 10-20% virus-containing suspension. The virus-containing suspension is purified from ballast impurities in a known manner. To do this, you can use low-speed centrifugation, processing the suspension with various detergents (Freon-113, chloroform) and a high molecular weight polyhexamethylene guanidine hydrochloride flocculant (HSV) in combination with low-speed centrifugation or separation. Purification of the virus-containing suspension from ballast proteins is carried out at 6-12 ^o C. the Purified suspension is served in a sterile reactor. Ethyleneimine dimer (DEI) is used to inactivate the virus. Pre-prepared 10% solution of DEI in demineralized water, pH 7.9-8.0. Then, a 10% aqueous solution of DEI is added to a virus-containing suspension heated to 28-34 ^° C to a concentration of 0.15-0.4%. The mixture is thoroughly mixed and maintained at 35-36 ^o C for 24 hours. Upon completion of inactivation, the antigenic material is cooled to 2-8 ^o C. As a adjuvant sorbent in the vaccine use a sterile 3-4% colloidal GOA solution. The pH of GOA is adjusted to between 7.5-7.6 and cooled to 2-8 ^° C. GOA is added to the cooled antigen with constant stirring to a concentration of 0.1%. Then, a 10% aqueous solution of saponin adjuvant is added to the mixture at the rate of 0.5 mg per dose of the vaccine, bringing the concentration of saponin in the vaccine to 0.5-2%. After mixing the mixture for one hour, determine the pH of the drug and, if necessary, bring its value to 7.4-7.6. The resulting vaccine is monitored for avirulence, safety and sterility, and then packaged in sterile vials.

 Sterility of the vaccine is determined in accordance with GOST 28085-89.

 Avirulence and harmlessness of the drug is determined by introducing a 10-15-day-old chickens vaccine in a triple dose.

 The resulting vaccine is a pink-brown liquid with a loose precipitate of sorbent, which, when shaken, easily breaks into a homogeneous suspension.

 The obtained vaccine against CHP has the optimal component composition (formulation), wt.% (See table.A).

 The content of antigen in the vaccine within the above limits is its effective amount in the drug, ensuring the achievement of a technical result.

A vaccine series is produced for practical use, if the maximum vaccination volume for chickens equal to 0.5 cm ³ contains at least 7 protective doses (PD ₅₀ ).

The vaccine is used in clinically healthy birds to create a high level of immunity. Broiler chickens are vaccinated once at the age of 1-18 days. The chickens are given the vaccine intramuscularly in the thigh or chest or subcutaneously in the middle third of the neck in a volume of 0.3 cm ³ . A single inoculation with an inactivated sorbed anti-hepatitis C vaccine provides 90-100% protection of immunized chickens against a disease of hepatitis C virus in the period starting from the third day after the vaccine is administered. Repair young animals are immunized twice: the first time at 1-18 days of age at a dose of 0.3-0.5 cm ³ and the second time at 90-140 days of age at a dose of 0.5-0.7 cm ³ . Immunity begins on the third day after vaccination and lasts up to 8 months. Twice use of the vaccine in the parent herd provides reliable passive maternal immunity in young animals up to 8-12 days of age, which allows it to be protected from the field virulent virus of the CGPS in early life.

 Example 2

 Studies were carried out to determine the antigenic activity of each formulation of the inactivated vaccine against CHPA obtained as described in example 1.

The antigenic activity of the vaccine samples was determined on chickens of 15 days of age, free of antibodies to the virus CGPC. Samples of the vaccine were administered at a dose of 0.3 cm ³ into the thigh muscle. 21 days after vaccination, the blood was taken from the chickens, serum was obtained, and they were examined in the RDP with a specific serum hyperimmune to the CGPC virus. The research results are presented in table. 1, 2 and 3, respectively.

 The results presented in table. 1, 2 and 3, indicate a high antigenic activity of the inactivated vaccine against SGPK from strain KP95 114.

 Example 3

Studies were carried out to determine the immunogenic activity of an inactivated vaccine against hepatitis C, obtained as described in Example 1. For this, a series of experiments were performed on chickens of 10 days old that did not have antibodies to the hepatitis C virus, which served as a biological model to test the bird's immune response to the introduction of a vaccine. The experience was taken 350 chickens, which were divided into nine groups of 50 goals each. Eight groups were test vaccine subjects, and one group served as a control. The chickens of the experimental groups were vaccinated once intramuscularly in doses of 0.15; 0.3 and 0.5 cm ³ inactivated sorbed vaccine 2 and 5 series. On the 3rd, 5th, 7th, 10th and 14th day after vaccination, 10 chickens were selected from each group, which were placed in a separate box and intramuscularly infected with SGPV virus from strain KP95 at a dose of 3.78 ± 0.23 Ig LD ₅₀ / 0.2 cm ³ . After infection, the birds were monitored for 10 days. Fallen chickens were recorded and the percentage of protection was counted. The results of the studies are presented in table.4. As can be seen from the data given in table 4, a single vaccination of chickens at a dose of 0.5 cm ³ provided 100% protection against control infection after 3 days after administration of the drug. In groups of chickens that were injected with an inactivated sorbed vaccine at doses of 0.15 and 0.3 cm ³ , 100% protection against control infection occurred 5-7 days after vaccination. Mortality in the control group of chickens was at the level of 80-100%.

 Example 4

Studies were carried out to determine the actual immunogenic activity (ImD ₅₀ / cm ³ ) and the protective activity of the inoculated dose of the inactivated sorbed anti-hepatitis C vaccine, made as described in Example 1. For this, experiments were carried out on 8-day-old egg-cross chickens without antibodies to the virus CGPC. In the experiments used chickens with a total of 250 goals. The chickens were inoculated with an inactivated sorbed vaccine against SGPK 2, 3 and 5 series. The immunogenic activity of vaccine samples was determined by the volumetric method using calculations according to the Kerber-Ashmarin formula. The protective activity of the vaccine (PD ₅₀ ) was calculated by dividing the vaccine dose of the vaccine by the value of IMD ₅₀ . The research results are given in table. 5. The experiments found that the immunogenic activity of the inactivated vaccine against CHPK is not significantly affected by the method of administration of the drug. So, the ImD ₅₀ when the vaccine was administered subcutaneously was 0.0107 cm ³ , and intramuscularly - 0.012 cm ³ . From the data of Table 5 it is seen that one vaccine dose of the vaccine in the volume of 0.3 cm ³ for subcutaneous administration of the drug contains (28 ± 1.15) PD ₅₀ , and one vaccine dose of the vaccine in the volume of 0.3 cm ³ for intramuscular injection contains (25 ± 1.15) PD ₅₀ .

Thus, the above information indicates the fulfillment when using the invention of the following set of conditions:
- a method of manufacturing an inactivated vaccine against SGPK embodying the invention is intended for use in agriculture, namely in veterinary virology and biotechnology;
- for the proposed invention in the form described in the independent claim, the possibility of its implementation using the means and methods given in the description or known prior to the priority date is confirmed;
- when using the proposed method for the manufacture of inactivated vaccines against SGPK achieved the technical result provided for by the task of creating the invention.

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

SOURCES OF INFORMATION
1. Cowen BS, Lu H. et al. Characterization studies of fowl adenoviruces associated with inclusion body hepatitis / hydropericardium syndrome in chickens. Presented at the 68-th Northeastern Conference in Avian Diseases. June 10-12 1996, p.14.

 2. Lagutkin N.A., Arsentiev A.R. et al. Massive hydropericarditis in young meat chickens. Questions of veterinary virology, microbiology and epizootology: Materials scientific. conf. VNIIVViM. -Pokrov, 1992, part 2, 248-251.

3. Borisov VV, Borisov AV et. al. Hydropericardium syndrome in chickens in Russia // Proc. of the 11 ^th Int. Congress of the World Vet. Polt. Ass. August 18-22, Budapest, Hungary.-1997.-P.258.

 4. Winterfield R.W., Fadly A.M. et al. Immunization of chicken against adenovirus infection. Poult. Sci., 1977, V. 56, N5, 1481-1486.

 5. Ahmad I., Malik M.I. et. al. Efficacy of formalinized level-organ-vaccine against Angara disease in broilers // Veterinarski Arhiv.-1990.-V. 60.-N3.-P.131-138 (prototype).

 6. Ahmad I., Ahmad K. et al. Comparative efficacy of different Angara disease vaccine in broilers. Zootecnica International, 1991, V.30, N6, 67-69.

Claims (14)

 1. A method of manufacturing an inactivated vaccine against chicken hydropericarditis syndrome, comprising preparing antigenic material in the form of a virus-containing suspension from the liver of chickens infected with an infectious agent and killed with signs of chicken hydropericarditis syndrome, purification of antigenic material from ballast impurities, virus inactivation and control of the target product, characterized in that as a causative agent of infection using a strain of the virus of the syndrome of hydropericarditis of hens. Adenoviridae, genus Aviadenovirus, group 1, serotype 4, collection VGNKI KR95 114-DEPT, in an effective amount, and purified and avirulent antigenic material from this virus is combined with an adjuvant.  2. The method according to claim 1, characterized in that to obtain antigenic material using the liver of 5-50-day-old chickens.  3. The method according to claim 1, characterized in that the strain KP95 114 of the chicken hydropericarditis syndrome virus is cultivated in the body of 5-50-day-old chickens for 48-120 hours 4. The method according to claim 1, characterized in that the use of antigenic material with biological activity of 3.25-5.0 lgLD ₅₀ / 0.2 cm ³ .  5. The method according to claim 1, characterized in that the purified antigenic material is inactivated by an ethyleneimine dimer.  6. The method according to claim 5, characterized in that the ethyleneimine dimer is used in the form of a 10% aqueous solution.  7. The method according to any one of claims 5 and 6, characterized in that the ethyleneimine dimer is introduced into the antigenic material to a concentration of 0.15-0.4%. 8. The method according to any one of claims 5 to 7, characterized in that the inactivation of the antigenic material by ethyleneimine dimer is carried out for 23-25 hours at 35-36 ^° C and a pH of 7.4-7.6.  9. The method according to claim 1, characterized in that of the adjuvants use aluminum hydroxide.  10. The method according to claim 9, characterized in that the aluminum hydroxide is used in the form of a 3-4% colloidal solution.  11. The method according to any one of claims 9 and 10, characterized in that the aluminum hydroxide is introduced into the antigenic material to a concentration of at least 0.1%.  12. The method according to claim 1, characterized in that of the adjuvants use additional saponin.  13. The method according to p. 12, characterized in that the saponin is used in the form of a 10% aqueous solution.  14. The method according to any one of paragraphs.12 and 13, characterized in that saponin is introduced into the antigenic material to a concentration of 0.5-2%.

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