RU2764600C1 - Vaccine against escherichiosis of calves and piglets - Google Patents

Abstract

FIELD: pharmaceuticals.

SUBSTANCE: invention relates to the field of pharmaceuticals, namely to a vaccine against escherichiosis of calves and piglets. The vaccine includes in certain proportions: a cell-free culture medium containing inactivated thermolabile, thermostable and shiga-like Escherichia coli toxins obtained by mixing in equal proportions epizootic strains of Escherichia coli with genes of thermolabile, thermostable and shiga-like toxins inactivated with formalin to its final concentration of 0.4%, and also as adjuvants, a pyrogenal solution containing 100 mcg/ml of the active substance and 3% aqueous solution of polyacrylic acid.

EFFECT: invention provides an increase in the effectiveness of specific prevention of escherichiosis in calves and piglets, an increase in the immunogenicity, harmlessness and safety of the vaccine.

1 cl, 5 tbl, 7 ex

Description

The invention relates to biotechnology, in particular to the design of vaccines, and can be used in institutions involved in the creation of vaccine preparations for the prevention of escherichiosis in animals.

It has been established that effective means of specific prevention of escherichiosis in animals are vaccines containing inactivated exotoxins of pathogenic Escherichia coli (Biological preparations for the specific prevention and treatment of escherichiosis in animals / M.K. Pirozhkov // Diss. doc. veterinary sciences. - Moscow 2002. - 298 p.). Currently, there are studies on the use of biological products that include three types of pathogenic Escherichia toxins at once: thermolabile (LT) and thermostable (ST) and Shiga-like (STX) toxins, both in pure form and in combination with adjuvants (Karaev Ya.M. Immunogenic and protective properties of escherichiosis toxoid: thesis ... candidate of veterinary sciences / Ya.M. Karaev - Krasnodar: KubGAU, 2009. - 132 p; Candidate of Veterinary Sciences / A.S. Tishchenko - Krasnodar: KubGAU, 2011. - 124 p.). However, these studies have found that escherichial toxoid stimulates cellular and humoral immunity on the first day after the administration of drugs, and the use of adjuvants, although it enhances the immune response, but for a short time. Given these circumstances, it is necessary to look for new ways to increase the immunogenic activity of vaccine preparations based on inactivated E. coli toxins.

Known vaccine against escherichiosis animals (RF patent No. 2043771, class A61K39/108, 1995) made from somatic - O78, O141; protein adhesive - K88, K99, 987P, F41; capsular polysaccharide antigens - K80. K87, and also includes TL and GS-anatoxins. Currently, this vaccine has been upgraded by adding Escherichia strains and including, in addition to TL and TS, Shiga-like toxins (VT1 and VT2 anatoxins) of Escherichia coli inactivated with formalin in an amount of 0.4% with the addition of aluminum hydroxide gel as an adjuvant in an amount of 25% B 1 cm3 of the vaccine contains 10 billion Escherichia microbial bodies. Commercial name - Vaccine against escherichiosis of animals Verokolivak K88, K99, 987P, F41, TL-, TS-, VT1 and VT2 toxoids (manufacturer of the Federal State Unitary Enterprise Armavir Biofactory, Russia) (Vaccinal prevention of escherichiosis of farm animals // http://www .armbio.info/verokolivak.html; Vaccine against animal escherichiosis // http://www.armbio.info/catalog/1006-3).

The disadvantage of this improved vaccine is that it contains a very large amount of ballast antigens in the form of whole E. coli cells and an insoluble mineral adjuvant, and this negatively affects the formation of immune protection in animals, as well as the development of adverse reactions and complications in vaccinated animals. . In addition, the method of obtaining such a vaccine is very complicated, since it requires separate cultivation of 18 Escherichia strains.

It is known to use pyrogenal and polyacrylic acid separately as adjuvants to enhance the immunogenicity of escherichiosis toxoid containing inactivated toxins (TL, TS, STX) E. coli (Terekhov, V.I. The effect of various adjuvants on the immunogenic properties of escherichiosis toxoid / V.I. Terekhov, Y. M. Karaev, A. S. Tishchenko, A. V. Ivanov // Proceedings / KubGAU - 2009. - Issue No. 1 (part 1) Series: Veterinary sciences. - P. 100- 102.; Tishchenko A. S. et al. Influence of bacterial polysaccharide and polyelectrolyte on the immunogenic properties of escherichiosis toxoid // Young scientist. - 2016. - No. 20 (124). - P. 113-115.; Tishchenko A. S., Terekhov V.I., Serdyuchenko I.V. Immunogenicity of escherichiosis toxoid when used with pyrogenal and polyacrylic acid // Scientific support of the agro-industrial complex: collection of articles based on the X All-Russian conference of young scientists - Krasnodar: KubGAU, 2017 - S.287-288.).

The disadvantage of known studies is that escherichiosis toxoid using pyrogenal as an adjuvant and escherichiosis toxoid using polyacrylic acid as an adjuvant had low immunogenic properties, which negatively affected the formation and duration of post-vaccination protection in animals against E. coli toxins. In addition, white rats and rabbits acted as biological models in these studies, and it is not known how the components of the vaccine will behave in calves and piglets.

It is known the use of adjuvants, including polyacrylic acid and pyrogenal, in combination with escherichiosis toxoid in experiments on pregnant cows and pregnant sows (Tishchenko L.S. Influence of adjuvants on the immunogenic properties of escherichiosis toxoid: thesis ... candidate of veterinary sciences / A. S. Tishchenko - Krasnodar: KubGAU, 2011. - 124 pp. Tishchenko, A.S. Evaluation of the humoral immune response in pregnant sows immunized with escherichial toxoid in combination with adjuvants / A.S. Tishchenko, V.I. Terekhov / / Proceedings / KubGAU - 2011. - Issue No. 2 (29) - P. 144-147; Terekhov, V. I. Influence of adjuvants on the immunogenic properties of escherichiosis toxoid during vaccination of pregnant cows / V. I. Terekhov, A Tishchenko A.S., Novikova E.N., Vinokurova D.P., Kiyashchenko A.A., Kremyansky V. B. The spread of escherichiosis in piglets and the method of its specific prevention / Polythematic network electronic scientific journal of the Kuban State Agrarian University site. - 2018. - No. 137. - P.220-229), as well as the preventive efficacy of these drugs (Tishchenko, A.S. Preventive efficacy of escherichiosis toxoid in escherichiosis of piglets and calves / A.S. Tishchenko, V.I. Terekhov // Experience of international cooperation in the field ecology, forestry, veterinary medicine and hunting science: materials of the II-nd International scientific-practical conference / KubGAU - Krasnodar, 2010. - P.380-381).

The disadvantage of the known studies is that escherichiosis toxoid, using pyrogenal and polyacrylic acid separately as adjuvants, had a short immune response, which affected the effectiveness of specific prophylaxis.

The closest in technical essence is a technical solution that includes epizootic strains of Escherichia coli mixed in equal proportions with genes for thermolabile, thermostable and Shiga-like toxins and inactivated by formalin to its final concentration of 0.4%, (patent No. 2432174 dated 27.10.2011, IPC A61K 39/108, C12N 1/20, A61P 1/00).

The disadvantage of the known technical solution is the absence of adjuvants, and as a result, the low immunogenic activity of the drug.

The technical result of the invention is to increase the effectiveness of the specific prevention of escherichiosis in calves and piglets, increase the immunogenicity, harmlessness and safety of the vaccine.

The technical result is achieved by the fact that in the vaccine against escherichiosis of calves and piglets, including a cell-free culture medium containing inactivated thermolabile, thermostable and Shiga-like toxins of Escherichia coli, obtained by mixing in equal proportions of epizootic Escherichia coli strains with genes for thermolabile, thermostable and Shiga-like toxins, inactivated with formalin to its final concentration of 0.4%, followed by separation of the bacterial mass using sterilizing filtration, characterized in that it contains a pyrogenal solution containing 100 μg / ml of the active substance and a 3% aqueous solution of polyacrylic acid as adjuvants, which are introduced alternately followed by mixing in a cell-free culture medium containing inactivated thermolabile, thermostable and Shiga-like toxins of Escherichia coli, in the following ratio, wt. %:

pyrogenal solution containing 100 mcg/ml active substance 9-11 3% aqueous solution of polyacrylic acid 9-11 cell-free culture medium containing inactivated thermolabile, thermostable and Shiga-like toxins of Escherichia coli up to 100

The novelty of the technical solution lies in the fact that through the use of cell-free cultures of epizootic strains of Escherichia coli that produce heat-labile, heat-stable and Shiga-like toxins, inactivated by formalin, when used together with adjuvants: solutions of pyrogenal and polyacrylic acid, they provide an increase in the immunogenic properties of the vaccine and the effectiveness of prevention of escherichiosis in animals, as well as the safety of the biological product due to the absence of post-vaccination reactions after the application.

In the patent and scientific and technical literature, a set of features similar to the claimed one has not been found, which makes it possible to judge the inventive step of the proposed proposal.

The proposed vaccine meets the criterion of "industrial applicability" because it is easily reproducible and does not require complex technological equipment and conditions.

The resulting vaccine is packaged in 100 ml sterile vials, sealed and tested for sterility and harmlessness. In appearance, the finished vaccine is a clear liquid of light straw color without sediment and impurities. Shelf life in a dry and dark room at a temperature of 4-10 ° C - 1 year.

The resulting vaccine was tested for sterility and harmlessness.

At control inoculations of toxoid on MPA, MPB, Kitta-Tarozzi, Saburo, Endo medium, there was no growth of bacterial and fungal flora, which indicated the sterility of the preparation.

With intraperitoneal administration of the drug at a dose of 0.3 ml to white mice weighing 20-22 g, no inhibition and death were noted within 10 days, which is an indicator of the safety of the drug.

The effectiveness of the vaccine against Escherichia coli depends on the properties of the selected epizootic strains of Escherichia coli, the sequence of preparation and the percentage of components included in its composition, such as: 100 μg of the active substance - lipopolysaccharide (LPS) isolated from Salmonella typhi cells) (EAN: 4602379000047) - 8 ml (8 ampoules of 1 ml) - (8%); 9 ml (9 ampoules of 1 ml) - (9%); 10 ml (10 ampoules of 1 ml) - (10%); 11 ml (11 ampoules of 1 ml) - (11%) and 12 ml (12 ampoules of 1 ml) - (12%);

- 3% aqueous solution of polyacrylic acid - 8 ml (8%); 9 ml (9%); 10 ml (10%); 11 ml (11%) and 12 ml (12%).

Quantitative values for the introduction of adjuvants were selected experimentally, taking into account the immunogenic activity of the vaccine. With the introduction of solutions of pyrogenal and polyacrylic acid in a volume of 8%, the immunogenic activity increased, but still was not sufficient for long-term preservation of post-vaccination immunity. When adjuvants were administered in a volume of 12%, there was no increase in the production of specific antibodies more than when they were administered in a volume of 9-11%, and economic costs increased. In addition, following the increase in the input of polyacrylic acid, the risk of developing a local inflammatory reaction due to tissue irritation increased.

Five examples (1-5) are given of the dependence of the effectiveness of the vaccine against escherichiosis of calves and piglets on the ratio of the claimed components per 100 ml of the vaccine and two examples (6, 7) demonstrating the effectiveness of the use of the claimed invention.

For examples 1-5, epizootic Escherichia coli strains with genes for heat-labile, heat-stable and Shiga-like toxins were used, they were cultivated in nutrient broth at a temperature of 37°C for 6-7 days with double stirring daily, then they were inactivated with formalin to its final concentration 0.4% for 14 days with daily double stirring, after which the cultures were selected and mixed in equal proportions, the bacterial mass was separated using sterilizing filtration to obtain a cell-free culture medium, into which adjuvants were added alternately with subsequent mixing with the following volumetric quantities and percentage:

Example 1 - with the following ratio of components, wt. %:

pyrogenal solution containing 100 mcg active substance eight 3% aqueous solution of polyacrylic acid eight cell-free culture medium containing inactivated thermolabile, thermostable and Shiga-like toxins of Escherichia coli up to 100

With this ratio of components, the vaccine has a weak protective activity and protects against Escherichia infection caused by pathogenic Escherichia coli in calves in 75%, and in piglets in 72% of cases.

Example 2. With the following ratio of components, wt. %:

pyrogenal solution containing 100 mcg active substance 9 3% aqueous solution of polyacrylic acid 9 cell-free culture medium containing inactivated thermolabile, thermostable and Shiga-like toxins of Escherichia coli up to 100

With this ratio of components, the vaccine protects against Escherichia infection caused by pathogenic Escherichia coli in 86%, and piglets in 82% of cases.

Example 3. With the following ratio of components, wt. %:

pyrogenal solution containing 100 mcg active substance 10 3% aqueous solution of polyacrylic acid 10 cell-free culture medium containing inactivated thermolabile, thermostable and Shiga-like toxins of Escherichia coli up to 100

With this ratio of components, the vaccine protects against Escherichia infection caused by pathogenic Escherichia coli in 92%, and piglets in 87% of cases.

Example 4. With the following ratio of components, wt. %:

pyrogenal solution containing 100 mcg active substance eleven 3% aqueous solution of polyacrylic acid eleven cell-free culture medium containing inactivated thermolabile, thermostable and Shiga-like toxins of Escherichia coli up to 100

With this ratio of components, the vaccine protects against Escherichia infection caused by pathogenic Escherichia coli in 94%, and piglets in 90% of cases.

Example 5. With the following ratio of components, wt. %:

pyrogenal solution containing 100 mcg active substance 12 3% aqueous solution of polyacrylic acid 12 cell-free culture medium containing inactivated thermolabile, thermostable and Shiga-like toxins of Escherichia coli up to 100

With this ratio of components, the vaccine protects against Escherichia infection caused by pathogenic Escherichia coli in 84%, and piglets in 80% of cases. At the injection site, some animals, especially among piglets, developed dense, painful swelling and an increase in body temperature.

The vaccine against escherichiosis of animals is produced as follows.

The initial stages of obtaining Escherichia vaccine were carried out according to the RF patent for invention No. 2432174 dated 10/27/2011, IPC A61K 39/108, C12N 1/20, A61P 1/00.

Preliminarily, from epizootic strains of Escherichia coli, strains with genes for thermolabile, thermostable, and Shiga-like toxins were selected. For this purpose, AmpliSense E. coli-tox test systems (Central Research Institute of Epidemiology of the Ministry of Health of the Russian Federation, Moscow) were used. Confirmation of toxin formation on an artificial nutrient medium was carried out using a biotest on ciliates-stylonichia (RF patent for the invention 2262529 dated October 20, 2005, IPC C12N 1/10, C12Q 1/10). According to a well-known patent, to test the survival of ciliates, a culture of Escherichia coli grown on Hottinger broth with an incubation period of 1-3 days was used, then the medium with ciliates was mixed with the broth culture of Escherichia coli in a ratio of 3:1. In the presence of exotoxins in the culture medium, the number of dead ciliates should be at least 70-80%, while the number of dead ciliates in a control sample that does not contain a toxin should not exceed 5%.

The selected strains were individually inoculated into test tubes with 10 ml of nutrient broth for the accumulation of E. coli toxins, which included acid blood hydrolyzate (9-11%), baker's yeast autolysate (9-11%), peptone (0.9-1. 1%), sodium chloride (0.4-0.6%), disubstituted sodium phosphate (0.05-0.15%) and potassium chloride (0.01-0.03%) (RF patent for invention 2342425 dated 12/27/2008, IPC C12N 1/20, C12Q 1/04). Then they were placed in a thermostat and cultivated at 37°C for 6-8 hours until a slight turbidity appeared, indicating the growth of microorganisms. Next, the resulting cultures were transferred into flasks with 200-300 ml of a similar nutrient broth and incubated at 37°C for: producing thermolabile and thermostable - 6 days, and producing shiga-like toxin - 7 days. The flasks were shaken twice daily. Before the end of the incubation, 10 ml of culture liquid was taken from each flask, centrifuged at 10 thousand rpm for 30 min, and the presence of toxins was determined by biotesting on ciliates.

In the presence of toxins, formalin was added to the remaining cultures to a concentration of 0.3-0.4%. Inactivation of cultures was carried out for 14 days at a temperature of 37°C with daily double stirring. After completion of inactivation, equal volumes of cultures were combined, thus obtaining a complex preparation containing 3 types of antigens. Using sterilizing filtration, the microbial mass was separated from the culture medium, thus obtaining a cell-free culture medium of Escherichia coli (Escherichia antigen toxoid component). Further, adjuvants were introduced into the cell-free culture medium of Escherichia coli: the preparation was pyrogenal solution (lipopolysaccharide isolated from microbial cells of Salmonella typhi, manufacturer of N.F. OOO Khimdirect service, Moscow) with the following ratio of components, wt. %:

pyrogenal solution containing 100 mcg active substance 9-11 3% aqueous solution of polyacrylic acid 9-11 cell-free culture medium containing inactivated thermolabile, thermostable and Shiga-like toxins of Escherichia coli up to 100

Confirmation of the effectiveness of the use of the vaccine for the prevention of escherichiosis in calves and piglets is given in examples 6 and 7.

The use of the vaccine makes it possible to protect calves and piglets from Escherichiosis infection for at least 6 months (observation period).

To prove the effectiveness of the proposed vaccine, studies were conducted on white rats, calves and piglets. A satisfactory result was obtained by optimizing the input of solutions of pyrogenal and acrylic acid, which is demonstrated by the following experiments.

Experience 1 - presents the results of studies conducted on white rats.

4 groups of white rats were formed, 6 heads each, which were injected intramuscularly with 0.15 ml of the drug once, in the first group, the Escherichiosis vaccine was used without adjuvant (EV), in the second group, the Escherichiosis vaccine with a pyrogenal solution containing 100 μg of the active substance - lipopolysaccharide (EV+LPS), in the third - escherichial vaccine with polyacrylic acid (EA+PAA), in the quadruple - escherichial vaccine with solutions of pyrogenal and polyacrylic acid (EA+LPS+PAA). After 7, 14, 21, 60, 90 and 120 days in the reaction of indirect hemagglutination, the titer of antibodies to coli toxins was determined. The results of the experiment are shown in Table 1.

From the data in Table 1, it can be seen that in animals of all groups, after the introduction of the vaccine, antibodies are produced within 7 days. However, in groups 2 and 3, where it was administered with adjuvants, the level of antibodies was 1.7 times higher than in animals that were administered the vaccine without pyrogenal and polyacrylic acid, and in animals of group 4, the level of antibodies was 2 times higher. In the following days, in animals of group 1, the level of antibodies decreased, but in animals of groups 2, 3, and 4, it increased up to day 14, exceeding the values of group 1 by 1.9, 1.8, and 2.4 times, respectively. Subsequently, in all groups, a gradual decrease in antibodies by day 120 was established to a value of 1.2±0.7, 2.0±0.6, 1.8±0.7 and 3.6±0.4 log _2, respectively. In group 4 animals, the level of antitoxic antibodies remained the highest even 120 days after immunization and was sufficient for a protective effect against E. coli toxins.

Experience 2 - presents the results of studies conducted on calves and piglets.

For research, the vaccine was vaccinated in pregnant cows and heifers twice subcutaneously. The first time for 30, and the second time for 15 days before calving at a dose of 5 and 10 ml, respectively. Calves were also vaccinated twice, the first time at the age of 15-20 days, and the second time after 10-14 days at a dose of 1 and 2 ml, respectively.

Pregnant sows were vaccinated intramuscularly twice, the first time 30 days before, and the second time 20 days before farrowing at a dose of 5 ml; piglets were vaccinated for the first time at the age of 15-17 days, and the second - at the age of 28-30 days at a dose of 0.25 and 0.5 ml, respectively.

Example 6 - shows the results of studies on the use of a vaccine for the prevention of escherichiosis in calves.

In the economy of the Krasnodar Territory, which is unfavorable for calf Escherichiosis, a production test of the proposed vaccine was carried out. According to the veterinary laboratory, diarrhea in young cattle on this farm is caused by pathogenic strains of Escherichia coli. The effectiveness of the vaccine made according to the claimed method was compared with the effectiveness of the prototype, for this purpose 3 groups of pregnant cows were formed, 25 heads each.

The first group of cows was immunized with the claimed vaccine for the first time 30 days before calving at a dose of 5 ml, the second time 15 days before calving at a dose of 10 ml. The calves obtained from these cows were also immunized with the claimed vaccine for the first time at the age of 15-20 days at a dose of 1 ml, and the second time after 10-15 days at a dose of 2 ml. The vaccine was injected subcutaneously into the region of the upper third of the neck.

The second group of cows and calves derived from them were immunized with Escherichiosis vaccine with pyrogenal solution according to the scheme and in doses similar to those in the first group.

The third group of cows and calves derived from them were immunized with Escherichiosis vaccine with a solution of polyacrylic acid according to the scheme and in doses similar to those in the first group.

The fourth group of cows and calves derived from them were immunized with Escherichiosis vaccine without adjuvants according to the scheme and in doses similar to those in the first group.

The fifth group of cows was a control group; they and their calves were not immunized.

The effectiveness of the vaccines was assessed when the calves reached the age of 3 months and when they were transferred to another corps for group keeping. The evaluation criteria were morbidity and mortality rates.

The results of the experiment are shown in Table 2, from the materials of which it is clear that in the first group, escherichiosis infection was registered in 2 calves, while they managed to stop their diarrhea within 3 days, and on the 5th day the calves fully recovered. Thus, the prophylactic efficacy of the proposed vaccine was 92%.

In the 2nd group, escherichiosis infection was registered in 5 calves, and in one calf it proceeded very hard and ended in the death of the animal. The prophylactic efficacy of the analog was 80%.

In the third group, escherichiosis infection was registered in 6 calves, in two calves it proceeded very hard and ended in the death of animals. The prophylactic efficacy of the analog was 76%.

In the fourth group, escherichiosis infection was registered in 8 calves, of which in three calves it was very difficult and ended in the death of animals. Prophylactic efficacy of the prototype was 68%.

In the fifth group of 25 calves, 18 animals fell ill, of which 5 died. Thus, in group 3, the incidence was 72%, and mortality was 27.8%, which indicates a significant severity of the disease.

Therefore, the use of the vaccine against escherichiosis, obtained by the claimed method, made it possible to more effectively prevent the disease of calves with escherichiosis infection. The incidence of calves vaccinated with the claimed vaccine was 8%, and the safety was 100%, while when using analogue 1, the incidence of calves was 20%, and the safety was 96%, analogue 2 - the incidence of 24%, safety 92%, analogue 3 - the incidence of 32%, safety 88%. In the control group, the incidence of acute intestinal infection in calves was 72%, and the survival rate was 80%.

The immunogenic properties of the Escherichiosis vaccine were studied in calves obtained from vaccinated pregnant cows. Serum was taken 7, 14, 28, and 56 days after their last vaccination to determine the presence of antitoxic antibodies in the diffuse precipitation reaction.

The effect of vaccines on antibody titer in calves is presented in Table 3.

From the materials of table 3 it can be seen that the immunization of cows and calves by the claimed method provides a more intense and prolonged specific immunity against escherichiosis infection. This is evidenced by the amount of antibodies to E. coli toxins in calves, which are 2-16 times more in animals of the first group than in animals from the second and third groups, and 8-32 times more than in animals of the fourth and fifth groups.

Example 7. Substantiation of the effectiveness of the use of the vaccine for the prevention of escherichiosis in piglets.

In the farm long-term unfavorable for escherichiosis, piglets were formed into 5 groups of pregnant sows, 5 animals each.

In the first group, sows and piglets derived from them were immunized with the proposed vaccine (EV+LPS+PAK). Sows were immunized for the first time 30 days before farrowing at a dose of 5 ml, the second time - 20 days before at a dose of 5 ml intramuscularly in the base of the auricle. Piglets were immunized twice, the first time at the age of 15-17 days at a dose of 0.25 ml, and the second at the age of 28-30 days at a dose of 0.5 days intramuscularly in the area of the inner surface of the thigh.

The second group of sows and piglets derived from them were immunized with escherichiosis vaccine with pyrogenal solution (EV+LPS) according to the scheme and in doses similar to those in the first group.

The third group of sows and piglets derived from them were immunized with the Escherichiosis vaccine with polyacrylic acid (EV + PAA) according to the scheme and in doses similar to those in the first group.

The fourth group of sows and piglets derived from them were immunized with Escherichiosis vaccine without adjuvants (EV) according to the scheme and in doses similar to those in the first group.

The fifth group of sows was the control group; they and their piglets were not immunized.

The piglets were clinically observed for 90 days, the number of sick and dead animals was taken into account. The results of the study are presented in table 3. From the materials of the table it can be seen that in the 1st group, 53 piglets were obtained from 5 sows, of which 8 heads fell ill and 2 died. The incidence in the group was 15.1%, the mortality was 3.8% . Therefore, the preventive effectiveness of the proposed method was 84.9% with the safety of piglets in the group of 96.2%.

In the 2nd group, where the sows were immunized with a vaccine with pyrogenal solution, 52 piglets were born, of which 12 fell ill and 4 died. The morbidity in this group was 23.1%, mortality 7.7%, preventive efficacy 78.9%, with a survival rate of 92.3%.

In the 3rd group, 54 piglets were received, of which 14 fell ill, 6 died. Morbidity was 25.9%, mortality was 11.1%, preventive efficacy was 74.1%, and safety was 88.8%.

In the fourth group, where the sows were vaccinated with the vaccine without adjuvants, 53 piglets were born, of which 20 fell ill, 8 died. Morbidity was 37.7%, mortality was 15.1%, preventive efficacy was 62.3%, and safety was 84.9%.

In the 5th group, 53 piglets were received, of which 28 fell ill, 9 died. Morbidity was 52.8%, mortality 17%, safety 83%.

Therefore, the use of the vaccine against escherichiosis, obtained by the claimed method, made it possible to more effectively prevent the disease of piglets with escherichiosis infection. At the same time, the incidence decreased in comparison with analogue 1 by 8%, analogue 2 by 10.8%, analogue 3 by 22.6%, and in comparison with the group (control) by 37.7%, while the incidence decreased by 2- 4.5 times, which indicates the high efficiency of the proposed vaccine.

The immunogenic properties of the Escherichiosis vaccine were studied in piglets obtained from vaccinated pregnant sows. Serum was taken 7, 14, 28, and 56 days after their last vaccination to determine the presence of antitoxic antibodies in the diffuse precipitation reaction.

The effect of vaccines on antibody titer in piglets is presented in Table 5.

From the materials of table 5 it can be seen that the immunization of sows and piglets by the claimed method provides a more intense and prolonged specific immunity against escherichiosis infection. This is evidenced by the number of antibodies to E. coli toxins in piglets, which are 2-32 times higher in animals of the first group than in animals from other groups.

Claims (2)

Vaccine against escherichiosis of calves and piglets, including a cell-free culture medium containing inactivated heat-labile, heat-stable and Shiga-like toxins of Escherichia coli, obtained by mixing in equal proportions of epizootic Escherichia coli strains with genes for heat-labile, heat-stable and Shiga-like toxins, inactivated by formalin to its final concentration of 0 .4%, followed by separation of the bacterial mass using sterilizing filtration, characterized in that it contains a pyrogenal solution containing 100 μg / ml of the active substance and a 3% aqueous solution of polyacrylic acid as adjuvants, which are introduced alternately with subsequent mixing into a cell-free culture medium containing inactivated thermolabile, thermostable and shiga-like toxins of Escherichia coli, in the following ratio of components, wt.%: pyrogenal solution containing 100 mcg/ml active ingredient 9-11 3% aqueous solution of polyacrylic acid 9-11 cell-free culture medium containing inactivated thermolabile, thermostable and Shiga-like toxins of Escherichia coli up to 100