RU2247577C2 - Vaccine for specific prophylaxis of porcine chlamydiosis - Google Patents

Abstract

FIELD: veterinary microbiology, biotechnology.

SUBSTANCE: the suggested vaccine contains, as a source of specific antigens, an equal mixture of inactivated purified chlamydial antigens of strains: "PC-85" - an agent of porcine chlamydial abortion and "LC-87" - an agent of porcine chlamydial pneumonia simultaneously grown in vitelline and allantoid membranes of hen's embryos, at concentration of Chlamydia being 10-15 mg/ml adsorbed at potassium alum and emulsified in oil-lanolin adjuvant at the content of 0.1%-glutaric aldehyde. Application of vaccine in porcine clamydiosis-unfavorable porcine farms decreases the number of abortions and stillbirths in sows, decreases sickness rate and mortality level in piglets as a result of chlamydial infection.

EFFECT: higher efficiency of prophylaxis.

4 ex, 10 tbl

Description

The invention relates to the field of veterinary microbiology and biotechnology, in particular to the production of a vaccine for the specific prophylaxis of pig chlamydia, the etiological agent of which is Chlamydia psittaci.

Chlamydia of pigs is an infectious disease that occurs in the form of abortion, bronchopneumonia, urethritis, polyarthritis, encephalomyelitis and other syndromes. Chlamydia of pigs in recent years has become widespread in pig farms of the country (G.P. Shcherban and others, G.D. Firsova, T.G. Voskresenskaya. Chlamydia of pigs. // J. Veterinary Medicine. - 1978. - No. 8. - S. 55-58; V. A. Bortnichuk. Chlamydia of pigs // Kiev: Harvest, 1991; S. I. Prudnikov, P. I. Mitrofanov, T. M. Prudnikova. Chlamydia of pigs // Chron. Inf. Animals . - M .: Kolos, 1984. - S. 71-76). Chlamydial infection as a whole is characterized by a weak immunological reaction, a tendency to a chronic and latent course with the preservation of the pathogen in the body even after acute illness and clinical recovery (Chlamydia of farm animals. M .: Kolos, 1984. - S. 71-76).

Until recently, measures to combat chlamydia of pigs in our country and abroad were based on the use of medications, veterinary-sanitary and restrictive measures. While effective measures of control and specific prophylaxis have been developed and introduced in the case of chlamydia of sheep and cattle, in case of chlamydia of pigs these issues are under development and testing. In the United States in 1979, A.S. Seawell proposed a comprehensive vaccine for the prevention of chlamydia in sheep, cattle and pigs. The drug is a live embryo vaccine - made from a strain of chlamydia pneumoniae cats, safe for agricultural. animals (Seawell A.S. Methods of wsing chlamydia vaccine for preventing and treating bovine and diseases, USA). When taking into account the effectiveness of the use of live vaccines, the researchers concluded that they are more promising compared to inactivated drugs, but due to circumstances these drugs did not find widespread use in veterinary medicine, primarily due to the fact that live vaccines had a limited shelf life (N .Z. Khazipov et al. Immunity and specific prophylaxis // Chlamydia of agricultural animals. - M .: Kolos, - 1984. - S.193-200).

In our country, employees of the North Caucasus Zonal Scientific Research Veterinary Institute (G.P. Shcherban, G.D. Firsova, T.G. Voskresenskaya) proposed a polyvalent inactivated vaccine against chlamydia of pigs, the authors used chicken fibroblast culture as a substrate for chlamydia (Chlamydia of pigs. J. Veterinary medicine. - 1978. - No. 8. - S.55-58). According to the report, this vaccine has been shown to be highly effective in the prevention of chlamydia in pigs, horses and other animal species.

The well-known "Inactivated emulsin vaccine against chlamydial abortion of sheep" produced by the biological industry, the authors Yu.D. Karavaev, I. I. Naletov (USSR Author's Certificate No. 692128) recommended this vaccine for specific prevention of pig chlamydia, however, its use did not lead to tangible results (Tr. Institute VIEV. - 1981. - No. 53. - S.95-102). The multivalent vaccine "Plach" (against parvovirus infection, leptospirosis, Aujeszky's disease and chlamydia) has positively proven itself for immunization on large pig-breeding complexes. Currently, the vaccine is still being tested in pig farms in the country (V.A. Bortnichuk. Chlamydia pigs // Kiev: Urozhay, 1991. - P.150).

The closest analogue in essence to the present invention (prototype) is a “vaccine against chlamydia of farm animals”, containing as an antigen the causative agent of chlamydial abortion in cattle, grown in the yolk shell of chicken embryos, inactivated with merthiolate, and a solution of trihydrooxymethiamine tetamine , a solution of acrylamide and methylbisacrylamide, a solution of ammonium persulfate and a solution of glutaraldehyde (see Auth. certificate. No. 826744 “Vacci but against chlamydia farm animals ", cl. C 12 N 5/00 for 1984).

The composition of the vaccine, taken as a prototype, the following, ml:

Antigenic liquid of a 20-40% suspension of yolk sacs infected with chlamydia (ELD 50 10-6.5-7.0) 8-10

A mixture of solutions of trihydroxymethylaminomethane (0.23%) 1-2

A mixture of solutions of acrylamide (30%) and methylbisacrylamide (0.8%) 3-4

1.4% solution of ammonium persulfate 1-2

25% glutaraldehyde solution to a final concentration of 0.05 0.038

A disadvantage of the known vaccine is its low efficiency against chlamydia of pigs due to the existing antigenic differences between the strains of chlamydia of cattle and pigs, as well as the purity of the components of the vitelline membrane of chicken embryos of the original chlamydial biomass, which is used as a 20-40% suspension of the pathogen chlamydial abortion of cows (strain “250”) and, as a result of this, weak immunogenic activity of the vaccine, which does not ensure the formation of long-term and intense immunity against rubbish the dose of pigs. Inside the species Chl. psittaci, isolated from pigs, strains differ in biochemical properties, antigenic structure, as well as virulence in relation to experimental and natural organisms. This gives grounds to assert the existence inside the species Chl. psittaci serotypes and even variants. It was established that the strains isolated from pigs during abortion and bronchopneumonia have significant antigenic differences, which must be taken into account when designing antichlamydia vaccines. But some researchers, on the basis of the presence of a general group of thermostable thermostable complement-binding antigen in all chlamydia, speak about the possibility of creating a single vaccine that provides protection against chlamydia of animals of all kinds (V.A. Bortnichuk. Chlamydia of pigs // Kiev: Urozhay, 1991. - S. 152).

Therefore, the problem of creating an effective vaccine against chlamydia of pigs continues to be relevant.

The objective of the invention is to obtain a vaccine with high immunogenic activity and a wide spectrum of antigenicity in various clinical forms of the disease of pigs.

The essence of the invention is to obtain an inactivated vaccine against chlamydia of pigs containing two purified antigens from chlamydia strains of the species Chlamydia psittaci: "PC-85" - the causative agent of chlamydial abortion in pigs and "LS-87" - the causative agent of chlamydial pneumonia of pigs, grown simultaneously in vitelline and allois shells of chicken embryos, with a concentration of 10-15 mg / ml, adsorbed on potassium alum and emulsified in an oil-lanolin adjuvant, containing 0.1% glutaraldehyde, in the following ratio ntov, vol.%:

1. Inactivated chlamydial antigens from strains of the species Chl. psittaci isolated from pigs.

"PC-85" with a titer of ELD ₅₀ 10 ^-5.5 -10 ^-7.7 in 0.3 mm 24-25

"HP-87" with a titer ELD ₅₀ 10 ^-6.5 -10 ^-7.7 0.3 24-25 mm

2. Oil-lanolin adjuvant 48-49

3. Potassium alum 0.4-0.5

10% glutaraldehyde solution 0.1-0.15

The strains of chlamydia "LS-87" and "PC-85" included in the vaccine are exquisite and are industrial.

The strain "LS-87" the causative agent of chlamydial pneumonia of pigs was isolated in 1987 from light pigs, assigned to the order Chlamydiales, fam. Chlamydiaseae, genus Chlamydia, species Chlamydia psittaci. The strain of the causative agent of chlamydial pneumonia of pigs was assigned registration number LS-87 DEPT in the All-Russian State Collection of Microorganism Strains Used in Veterinary and Animal Husbandry (123022, Moscow, 5 Zvenigorodskoye Shosse, VGNKI). The strain was deposited on April 26, 2000.

Strain "PC-85" the causative agent of chlamydial abortion in pigs was isolated in 1986 from an aborted pig fetus, assigned to the order Chlamydiales, fam. Chlamydiaceae, a genus of Chlamydia, a species of Chlamydia psittaci. The strain of the causative agent of chlamydial abortion in pigs was assigned registration number PC-87 DEPT in the All-Russian State Collection of Microorganism Strains Used in Veterinary and Animal Husbandry (123022, Moscow, 5 Zvenigorodskoye Shosse, VGNKI). The strain was deposited on April 26, 2000.

Strain "LS-87" has the following properties:

Morphological signs. The strain culture is small elementary bodies of chlamydia, having a spherical shape of 200-400 nm in size, which well perceive Tsilya carbol-fuchsin in a 1: 5 dilution and are stained in bright red color according to the modified Stamp method.

Cultural properties. The strain of chlamydia "LS-87" is well cultured in the vitelline membrane of 6-7-day old and the allantoic membrane of 8-9-day-old developing chicken embryos.

The specific death of chicken embryos infected in the vitelline cavity is observed on the 5-9th day after infection; while the pathogen accumulates in the titer of 10 ^-5.25 -10 ^-6.5 ELD ₅₀ in 0.3 ml.

Pathogenic properties. The strain of chlamydia "LS-87" has high virulence for laboratory and susceptible animals. When infecting white mice and guinea pigs intranasally, subcutaneously and intraperitoneally with a 10% freshly obtained chlamydia suspension, 100% disease and the death of 31-73% of mice are noted. Guinea pigs are ill, with and / n infection, about 25% of the animals die. In pigs, the strain of chlamydia "LS-87" causes pneumonia, the birth of dead piglets after intravenous infection in a dose of 5 ml of 10 ^-6.5 ELD ₅₀ in 0.3 ml.

Antigenic properties. Evaluation of the antigenic properties of the strain was carried out in CSC, ELISA by cross-sectional studies of antigen made from chlamydia strain "LS-87" according to the "Instructions for the manufacture and control of a set of antigens and sera for serological diagnosis of chlamydia of farm animals, approved. GUV Gosagroprom of the USSR in 1988 with sheep’s immune serum obtained by hyperimmunization with this strain, as well as standard diagnostic kits made in the conditions of the virology laboratory of VNIVI from the Rostin-70 strains of the sheep chlamydia pathogen, the Lori ornithosis strain, the PC-85 strain - the causative agent of chlamydial abortion in pigs and strain "LS-87" - the causative agent of chlamydial pneumonia of pigs (see table 1).

Table 1
The study of the antigenic properties of the strain of chlamydia "LS-87" No. p / p Hyperimmune sera obtained for chlamydia strains Antigens from strains LS-87 RS-85 Rostinovo Laurie 1. "LS-87" 160 ^x 40 80 twenty 2. Rostinovo-70 40 80 160 40 3. "Lori" 10 twenty 80 twenty 4. "PC-85" 40 160 80 twenty Designation: "x" is the reciprocal of the antigen titers.

The data in table 1 indicate the affiliation of the strain "LS-87" to chlamydia antigenically different from the strain "PC-85", hyperimmune sera and antigens from the strain "PC-85" react with the antigen and sera of the strain "LS-87" titers 1:40, i.e. two orders of magnitude lower than with a homologous 1: 160.

Thus, the strains "LS-87" and "PC-85" have distinctive antigenic properties.

Immunogenic properties. The antigen made from the LS-87 strain in an inactivated adsorbed emulsified vaccine provides the formation of intense and prolonged immunity in vaccinated animals.

In experimental experiments on white mice, it was found that double immunization with a vaccine based on the strain LS-87 prevents white mice from developing chlamydia infection after artificial infection with 87% strain LS-87 and 30% strain PC-85 (table 2).

Thus, in experiments on white mice, the high immunogenic activity of the strain "LS-87" was established, the vaccine made on its basis provides the formation in vaccinated animals of intense immunity from the homologous strain "LS-87", and when infected with the strain "PC-85" immunity weak.

The strain "Chlamydia psittaci" "PC-85" has the following properties:

Morphological signs. The strain culture is small elementary bodies of chlamydia, having a spherical shape of 200-400 nm in size, which well perceive Tsilya carbol-fuchsin in a 1: 5 dilution and are stained in bright red color according to the modified Stamp method.

Cultural properties. Chlamydia strain "PC-85" is well cultivated in the vitelline membrane of 6-7-day old and allantoic membrane of 8-9-day-old developing chicken embryos.

The specific death of chicken embryos infected in the vitelline cavity is observed 5–9 days after infection; while the pathogen accumulates in the titer of 10 ^-6.25 - 10 ^-6.5 ELD ₅₀ in 0.3 ml.

Pathogenic properties. The strain of chlamydia "PC-85" has a high virulence for laboratory and susceptible animals. When infecting white mice and guinea pigs intranasally, intraperitoneally and subcutaneously with a 10% freshly obtained chlamydia suspension, 100% disease and death of 54-73% of infected white mice is noted. Guinea pigs are ill. In pigs, the PC-85 chlamydia strain causes abortion, the birth of non-viable piglets after intravenous infection at a dose of 5.0 ml 10 ^-6.5 ELD ₅₀ in 0.3 ml.

Antigenic properties. Evaluation of the antigenic properties of the PC-85 strain was carried out in (CSC, ELISA) by cross-sectional studies of an antigen made from the PC-85 chlamydia strain according to the Instructions for the Production and Control of a Set of Antigens and Serums for Serological Diagnosis of S. ch. animals ”, approved by the GUV Gosagroprom of the USSR in 1988 with sheep’s immune serum obtained by hyperimmunization with this strain, as well as standard diagnostic kits made in the laboratory of virology VNIVI from strains“ Rostinovo-70 "- the causative agent of sheep chlamydia strain of ornithosis - "Lori", strain "LS-87" - the causative agent of chlamydial pneumonia in pigs and strain "PC-85" - the causative agent of chlamydia pig abortion.

The data in table 3 data indicate the affiliation of the strain "PC-85" to chlamydia, antigenically different from the strain "LS-87". So hyperimmune sera and antigens from the strain "LS-87" reacted with the antigen and sera from the strain "PC-85" in titers of 1:40, and with homologous 1: 160.

Table 3
The study of the antigenic properties of the strain of chlamydia "PC-85" No. p / p Hyperimmune sera obtained for chlamydia strains Antigens from strains PC-85 LS-87 Rostinovo-70 Laurie 1. "PC-85" 160 ^x 40 80 twenty 2. Rostinovo-70 80 40 160 40 3. "Lori" 10 twenty 80 twenty 4. "LS-87" 40 160 80 twenty Designation: "x" is the reciprocal of the antigen titers.

Thus, the antigenic properties of the strain "PC-85" - the causative agent of chlamydial abortion in pigs differ from the antigenic properties of the strain "LS-87" - the causative agent of chlamydial pneumonia of pigs.

Immunogenic properties. The use of biomass as an antigen prepared from the PC-85 strain in an inactivated adsorbed-emulsified vaccine ensures the formation of intense and prolonged immunity in 80% of vaccinated animals.

In experimental experiments (table 4), double immunization of white mice with a vaccine based on the PC-85 strain found that it protected white mice from developing chlamydial infection after artificial infection with 80% virulent strains of the chlamydia pathogen of pigs PC-85 and 36% of the causative agent of chlamydial pneumonia of pigs (strain "LS-87").

Thus, in experiments on white mice, a high immunogenic activity of the PC-85 strain was established. A vaccine made on its basis ensures the formation of intense immunity in vaccinated animals.

In a comparative assessment of the two strains "PC-85" and "LS-87" belonging to the same species of Chl. psittaci, significant differences in virulence, immunogenicity, and antigenicity were identified (Table 5). So, with the intraperitoneal method of infection of white mice with the PC-85 strain, death of white mice was more than 73%, and with subcutaneous and intranasal methods it was 54-57%. The strain "LS-87" the greatest death of white mice was observed with the intranasal method of infection - 73%, and with subcutaneous and intraperitoneal - 31-34%, respectively. When guinea pigs were infected with the PC-85 strain, all animals were ill, while when infected with the LS-87 strain, more than 25% of the guinea pigs died from pneumonia.

Significant differences in clinical signs were observed when pigs were infected with PC-85 and LS-87 strains. Thus, in animals infected with the PC-85 strain, abortion and the birth of a non-viable offspring were mainly observed, and pneumonia prevailed in the clinic of the disease when infected with the LS-87 strain. Thus, based on a study of the biological properties of the strains "PC-85" and "LS-87, interstrain differences in virulence were established with respect to experimental and natural animals.

When studying the antigenic properties of strains "PC-85" and "LS-87, it was found that the antigens and sera obtained for strains" PC-85 "and" LS-87 "react with sera and antigens from strains" LS-87 " and PC-85 are two orders of magnitude lower than with homologous ones. Thus, it was found that the strains "PC-85" and "LS-87" significantly differ in antigenic terms.

A comparative study of the immunogenic properties of PC-85 and LS-87 strains showed that immunization of white mice with PC-85 strains prevents white mice from developing chlamydial infection after infection with PC-85 strain by 80% and 36% from the strain "LS-87". Immunization of white mice with the strain "LS-87" protected them from developing chlamydial infection after infection with the strain "LS-87" by 87% and 30% from the causative agent of chlamydial abortion of pigs (strain "PC-85").

Table 5
Comparative characteristics of pig chlamydia strains: "PC-85" - the causative agent of chlamydial abortion and "LS-87" the causative agent of chlamydial pneumonia No. p / p Data Strains PC-85 LS-87 1. 2. 3. 4. Pathogenicity For white mice, the percentage of death in the method of infection 1. Intranasally 57 ± 2 73 ± 2 2. Subcutaneously 54 ± 3 31 ± 4 3. Intraperitoneally 73 ± 2 34 ± 3 Guinea pigs as a percentage 1. Subcutaneously get sick get sick 2. Intraperitoneally get sick get sick 3. Intranasally get sick 25 ± 2 death from pneumonia Pigs 1. 2. 3. 4. 1. intravenously, intramuscularly abortions, births of dead and non-viable piglets pneumonia, the birth of dead piglets Antigenic properties The serum obtained for the strains: Antigen from strain "PC-85" Antigen from strain "LS-87" 1. PC-85 160 ^x 40 2. Rostinovo-70 80 40 3. Laurie 10 twenty 4. LS-87 40 160 x is the reciprocal of antibody titers Immunogenic properties The survival rate of white mice immunized with strains: 1. PC-85 80 + 4 36 ± 5 2 LS-87 30 ± 4 87 ± 2 The control 23 ± 2 30 ± 4

Thus, in experiments on the study of the immunogenic properties of the PC-85 and LS-87 strains, it was found that the preparations made on their basis ensure the formation in vaccinated animals of intense immunity from infection with homologous strains, and weak - from a heterologous strain.

The data obtained in the study of the immunogenic properties of “pork” strains: “PC-85” and “LS-87” indicate their different immunogenic activity and, as a result, weak cross-immunity.

Thus, the work we carried out on a comparative study of the properties of pork strains of chlamydia PC-85 and LS-87 showed that the isolated strains differ significantly, although they belong to the same type of chlamydia - Chl. psittaci. The obtained data served as a scientific prerequisite for the design of the anti-chlamydial pork vaccine. Therefore, to increase the efficiency of the created anti-chlamydial swine vaccine, two strains were taken: the pathogen of chlamydial abortion of pigs "PC-85" and the pathogen of chlamydial pneumonia of pigs "LS-87", thereby we achieved a significant increase in the immunogenicity of the vaccine created by expanding the spectrum of antigenicity and immunogenicity of the drug , which is a hallmark of the created anti-chlamydia vaccine from the existing prototype.

Example 1. The preparation of native suspensions of chlamydia pigs.

Chicken embryos of 6-7 days of age are used, half of which are infected in the vitelline cavity of suspensions of freshly-transmitted strains of chlamydial pigs PC-86, of the pathogen of chlamydial abortion of pigs and the other half of LS-88, the causative agent of chlamydial pneumonia of pigs, diluted to 10 ^-4 at a dose of 0 3 ml. Infected embryos are incubated at 37-38 ° C for two days, then ovoscopy is performed, the dead are taken and destroyed (nonspecific death), and live embryos are re-infected into the allantoic cavity with a suspension diluted to 10 ^-3 at a dose of 0.3 ml, respectively, with strains "PC-86" and "LS-87." Next, infected embryos are incubated at 37-38 ° C for 7-8 days. The specific death of 90-95% of infected embryos occurs 6–9 days after the initial infection. Fallen embryos are cooled and opened. From each embryo, 4-5 ml of chlamydia-containing allantoic fluid is aspirated and 1-2 g of chorin-allantoic and vitelline membranes are taken together with the accompanying yolk.

Chlamydia-containing material is stored frozen at minus 20-30 ° C until use. Allantoic fluid before freezing is clarified by centrifugation at 3 thousand rpm for 30 minutes.

To obtain a vaccine semi-finished product from the chlamydia-containing material of the two strains, a 10% initial suspension is prepared in phosphate-buffered saline (FSB) pH 7.2-7.4 as follows: the chlamydia-containing membranes are weighed, thermolysed by freezing and thawing three times and suspended until 20% concentration in a colloid mill, adding the calculated amount of the FBI. The suspension is centrifuged at 3000 rpm for 30 minutes and the middle layer of the centrifugate is suctioned off - a suspension of chlamydia. The tissue sediment and the upper yolk layer are re-homogenized in the same amount of PBS, centrifuged under the same conditions, and the chlamydia suspension is again suctioned off.

The frozen chlamydia-containing allantoic liquid is thawed as quickly as possible in a water bath at a temperature of 37 ° C, vigorously shaking the bottle to prevent the formation of large flakes. Thawed slightly opalescent allantoic liquid is diluted in a ratio of 1: 1 with the FBI.

Example 2. Combining strains of chlamydia and inactivation.

Chlamydia-containing suspensions of infected membranes and allantoic fluid of strains "PC" and "LS" are combined and formaldehyde is added in a final concentration of 0.2%. The suspension after adding the reagents is mixed and incubated for 48 hours at a temperature of 37-38 ° C in order to inactivate chlamydia. Chlamydia from an inactivated suspension is precipitated by centrifuging it at 18-20 thousand rpm for 60 minutes. The supernatant along with the upper yolk layer is removed, the precipitate of elementary chlamydial bodies is weighed and resuspended in a ratio of 1:25 with PBS containing 0.1% glutaraldehyde.

The standardization of the suspension according to the content of elementary bodies of chlamydia is carried out by determining its optical density. For this, a 1:10 dilution is prepared from the prepared suspension of chlamydia (4.5 ml of the PBS are added to 0.5 ml of suspension) and the absorption coefficient is measured at a 56 M FEK at a wavelength of 540 nm in cuvettes with a working length of 10 nm. Optimum for obtaining an immunogenic vaccine is an absorption coefficient of 1.4-1.6 units, which corresponds to a content of 10-15 mg of antigen in one ml of suspension.

A standardized suspension of chlamydia is passed through a colloidal mill, potassium alum is added in a final concentration of 0.5% and homogenized again for 5 minutes, then the suspension is poured into glass cylinders and placed in a refrigerator at 4-6 ° C for 1-2 days. Under these conditions, chlamydia is adsorbed on potassium alum and precipitated in the form of flakes. The supernatant is carefully aspirated and removed, and the precipitate is resuspended to the original volume with a phosphate-buffered saline containing 0.1% glutaraldehyde. The resulting semi-finished vaccine with a preservative is stored in a refrigerator at 4-6 ° C.

Example 3. Preparation of a vaccine.

For the manufacture of 1 liter of vaccine, 450 ml of the inactivated Chlamydia psittaci antigen of the LS-87 and PC-86 strains containing 10-15 mg / ml of chlamydia adsorbed on potassium alum with 0.1% glutaraldehyde are taken. 550 ml of a sterile oil-lanolin mixture are added in a ratio of 84-16. Pass through a colloidal mill and packaged in sterile vials of 100 ml.

When changing the minimum or maximum in the ratios of the components during the manufacture of the vaccine, the emulsion is stratified into water and oil fractions, which reduces the immunogenicity of the emulsion vaccine.

Example 4. The control of the vaccine for sterility, safety and antigenic and immunogenic activity.

The vaccine is tested for sterility, durability of the emulsion and harmlessness by conventional methods.

10 series of vaccines were tested in laboratory animals and pregnant gilts. White mice weighing 16-18 g were vaccinated intraperitoneally at a dose of 0.1 ml, guinea pigs subcutaneously 1.0 ml, followed by monitoring the condition of experimental animals for 14 and 23 days, respectively.

The death of laboratory animals after vaccination during the observation period has not been established. The general condition of the vaccinated animals in all series of experiments was satisfactory. The test series of the vaccine against chlamydia pigs were sterile, harmless (tables 6, 7).

Table 6
The results of the study of the sterility of the experimental series of vaccines against chlamydia pigs No. p / p Vaccine series numbers Bacterial culture media BCH MPA IISP Saburo 1. No. 1 sterile sterile sterile sterile 2. No. 3 sterile sterile sterile sterile 3. Number 5 sterile sterile sterile sterile 4. Number 8 sterile sterile sterile sterile 5. Number 13 sterile sterile sterile sterile 6. Number 14 sterile sterile sterile sterile 7. Number 19 sterile sterile sterile sterile 8. Number 20 sterile sterile sterile sterile 9. Number 23 sterile sterile sterile sterile 10. Number 24 sterile sterile sterile sterile Table 7
The results of a study of the safety of experimental series of vaccines against chlamydia pigs in white mice and guinea pig No. p / p Vaccine series numbers Vaccinated Out of control Animals fell b / m m / s b / m m / s b / m m / s 1. No. 1 12 4 12 4 0 0 2. No. 3 12 - 12 - 0 - 3. Number 5 fifteen 4 fifteen 4 0 0 4. Number 8 fifteen - fifteen - 0 - 5. Number 13 10 4 10 4 0 0 6. Number 14 12 2 12 2 0 0 7. Number 19 12 - 12 - 0 - 8. Number 20 12 4 12 4 0 0 9. Number 23 12 - 12 - 0 - 10. Number 24 10 4 10 4 0 0 TOTAL: 122 22 122 22 0 0 Note: b / m - white mice;
m / s - guinea pigs.

Immunogenicity control is carried out on guinea pigs. Animals are vaccinated at a dose of 0.5 ml subcutaneously in the scapular region on the right and left (0.25 ml each). Vaccination promotes the production of specific chlamydial complement-binding antibodies on days 30-35 in titers of 1: 5-1: 80 (arithmetic average titer not lower than 1:20) (table 8).

Control of the immunogenic activity of the vaccine against pig chlamydia is carried out on white mice. Mice are vaccinated subcutaneously in a dose of 0.1-0.15 ml. Control infection of experimental and intact animals is carried out with the strains of chlamydia “PC-86” and “LS-88” intraperitoneally at a dose of 0.5 ml 30 days after immunization. The survival rate of white mice after control of infection with virulent cultures of chlamydia strain PC-86 and “LS-88” ranged from 73-85% (79.6 ± 4.3%) and 73-80% (76.6 ± 2.4 %) respectively. The average survival rate of vaccinated mice after challenge was 78.2 ± 2.1% (Table 9). In the groups of control mice after infection, 100% clinical disease and death of 60 to 80% of the animals were noted. The survival rate of intact mice after infection with the PC-86 and LS-88 chlamydia strains ranges from 20-26% (24 ± 2.4%) and 28-40% (35.6 ± 4.7%), respectively.

In experiments on sows under experimental conditions, it was found that a single intramuscular injection of a vaccine in a dose of 2.0 ml to sows protects them from the development of chlamydial abortion and pneumonia of pigs after infection with virulent chlamydia strains “PC-86” and “LS-88” ( table 10).

The vaccine is used as a preventive measure in farms and breeding enterprises unsuccessful for pig chlamydia. Clinically healthy animals are vaccinated once. Patients with chlamydia of animals before vaccination are subjected to treatment with tetracycline antibiotics. Sows and repair pigs are vaccinated 15-20 days before insemination or mating. The vaccine is administered to pigs intramuscularly in the neck at a dose of: adult animals - 2 ml; 3-6 month old pigs - 1 ml. In vaccinated animals at the injection site, a local reaction is observed in the form of a slight swelling, which disappears within 2-4 weeks. Immunity in vaccinated animals occurs one month after vaccination and lasts for six months. The shelf life of the vaccine is 1 year.

The production test of the vaccine manufactured in the Virology Laboratory at VNIVI (Kazan) was carried out under the conditions of the Sosnovoborsky pig farm of the Republic of Tatarstan, at the G.D. Guy Sengeleevsky district of the Ulyanovsk region, on the farm "Zvenigovsky" Mari-El and farms of Chuvashia, Urmarsky district: agricultural holding "MIR", KLP them. "Michurina", PSH "Chakinskoe". The data on vaccination against chlamydia of pigs in dysfunctional farms of Tatarstan, Mari-El, Chuvashia and the Ulyanovsk region are presented in table 11, from which it follows that as a result of the vaccine, the number of abortions and stillbirths in 6 dysfunctional farms ranged from 10 to 10, 6-66% (average 48%); mortality and forced slaughter of piglets ranged from 8-43% (on average 26%).

As a result of the use of the vaccine, the number of abortions and stillbirths in these households ranged from 1.0-16.0 (an average of 8%), i.e. decreased by 6 times. The incidence, mortality and forced slaughter of piglets from chlamydial infection also decreased significantly (2.3 times).

For the period 1995-2000, more than 200,000 doses of the vaccine against chlamydia of pigs were produced, which are used in 22 farms that are unsuccessful for chlamydia of pigs in the Russian Federation (table 12), including two large pig-breeding complexes: Sosnovoborsky of the Republic of Tatarstan and Zvenigovsky Republic of Mari El.

Experiments on a wide production test of the inactivated emulsion vaccine against pig chlamydia that we developed in various regions have shown that the vaccine is highly effective and, when systematically used, stabilizes the epizootic situation of chlamydia and improves the health of the pig population from chlamydia infection.

Table 10
Data before vaccination of pigs against chlamydia in dysfunctional farms of the Russian Federation No. p / p Regions and republics of the Russian Federation Number of households The number of vaccinated pigs Of the Republic 1. Tatarstan 8 22450 2. Mari El Republic 3 9600 3. Chuvashia 1 400 4. Bashkortostan 1 3000 5. Mordovia 2 1650 Areas 1. Perm 3 2400 2. Kursk 1 6000 Primorsky Krai 3 2125 TOTAL: 22 47625

Claims (1)

A vaccine for specific prophylaxis of pig chlamydia, containing the inactivated Chlamydia psittaci antigen, adjuvant and preservative, characterized in that it contains inactivated and purified chlamydial antigens from strains of PC-85 - the causative agent of chlamydial abortion in pigs and LS-87 - the causative agent of chlamydial pneumonia vitelline and allantoic membranes of chicken embryos, taken in equal amounts with a concentration of chlamydia of 10-15 mg / ml, adsorbed on potassium alum and emulsified in oil-lanolin adjuvant containing 0.1% glutaraldehyde as a preservative.