SI9400244A - Vaccine against trighophytosis of animals - Google Patents

Abstract

The invention relates to veterinary science, precisely to prophylaxis and the treatment of trichophytosis in animals. The invented vaccine against trichophytosis in animals differs from the known preparations in containing the additional antigen of the Trychophyton Mentagrophytes strain 27 (VGNK I) in a sufficient dose to give an immune reaction in an animal.

Description

VSEROSIISKY GOSUDARSTVENNY

NAUCHNO-RESEARCH INSTITUTE OF CONTROL

MOSCOW, RUSSIA

ANIMAL TRICHOPHYTOSIS vaccine

FIELD OF THE INVENTION

The present invention relates to biotechnology, and in particular to trichophytosis vaccines in animals.

BACKGROUND OF THE INVENTION

In the art to date, the known vaccine LTF-130 and the vaccine T-130 / K /, which mainly contain the Trichophyton faviforme antigen of the genus Trichophyton faviforme, are known as animal vaccine, mainly in animals. 130 (syn. Tr. Verrucosum) or an antigen of the genus Tr. favofirme 130 L, gelatinase sucrose stabilizer (see UK patent No. 1330240, C1.A5B, 1973; Inventor's Certificate USSR No. 955571, C1.A61K 39/00,

1980).

A disadvantage of the known vaccines is that they exhibit a pro-active effect in only one species Animals, namely livestock and that they only affect one type of agent, that is, Tr. f av and forms.

According to Bodin (1902) (see VM Leschenko. Laboratory Diagnosis Of Fungus Infections, M., Medicine, 1982, ρ.109), it should already be pointed out that the name of the genus Tr.faviforme 120 should be more precisely defined than Tr .verrucosum, because, according to George's Classification (1957) (same source, p. 34), the correct name for the cause of trichophytosis in animals is Tr.verrucosum.

The object of the present invention is the development of a vaccine with high immune capacity against trihofitosis in animals, which is caused by Tr.verrucosum, Tr.verrucosum var autotropicum, Tr.mentagrophytes, Tr.equimem, Tr.sarkisovii.

BRIEF DESCRIPTION OF THE INVENTION

The object of the invention was achieved by providing a vaccine additionally containing the Tr.mentagrophytes antigen VGNKI No.27 with the following component ratio, (weight percentages);

Antigen of the genus

Tr. verrucosum 130L

Ant and genus genus Tr.

mentagrophytes VGNKI No.27

Sucrose

With e 1 a t ina

Water

300-600 x 10 ^ microconidium / cm ^

300-600 χ 10 ⁶ microconidia / cm ^ 10.0 - 20.0

1.5 - 4.0 other

DETAILED DESCRIPTION OF THE INVENTION

The vaccine was prepared on the basis of the genus Tr.mentagrophytes VGNKI No.27.

This species was obtained from an epizootic white mouse species with directional, graded selection, based on spore formation in rapidly growing colonies with active accumulation of microconidia.

Type Tr. mentagrophytes is deposited in the Collection of Microorganisms at the All-Russia State Research Institute for Control, Standardization and Specification of Veterinary Preparations, Registration No.; 27 (All - State Research Institute for the Control, Standardization and Specification of Veterinary Products, Registry St: 27;

Op. prev.)

The genus of fungi Tr. mentagrophytes VGNKI 27 has the following characteristics and characteristics.

Morphological characteristics. The mycelium is colorless, flat, coated, of seven parts and can decompose into fragments, the end hyphae sometimes forming spirals (usually 3-5 turns), the diameter of the mycelium being 0.002-0.003 mm. Microconidia are numerous, round, pear-shaped, elongated, very similar to the rod-shaped form, exposed on the surface of the mycelium in clusters or, more often, individually, in size 0.0015-0.0052 mm.

Microconidia are individual. The number of groups is 1-5, more often 1-2, with a dimension from 0.01-0.03 to 0.003-0.008 mm.

Cultures of 20-30 days old can contain round shaped chlamydospores with double shaped membranes, always single, 0.003-0.005 mm in diameter. The arthrospores that develop in the individual sections of the mycelium are thin-walled and resemble oval or rod-shaped microconidia, measuring 0.003-0.009 mm.

Culture properties: Grows on agar in the form of a white, flat velvet colony with a small bulge in the middle. The part that grows is straight, the opposite is reddish-brown, the diameter of the colony is 50-90 mm (15-20 days old culture).

Rod No. 27 grows on Sabouraud glucose peptone agar, in the form of a flat, white, velvety colony in a cream colored hue in the center. The growing margin is straight, the opposite is pale yellow, sometimes darker in the middle, the size of the colony is 50-90 mm (15-20 days old culture).

It grows on the Henneberg medium in the form of a white, velvety colony, in the form of a radial fan. The growing margin is spidery and the opposite is reddish-purple, the colony having a diameter of 50-80 mm (culture 15-20 days old).

Disputes arise. The highest agar spore formation rate is 198.4-259.2χ10θ microconidia / cm ^.

Virulent properties. 2-3 weeks after intramuscular injection of Tr. mentagrophytes to Guinea pigs, rabbits, sheep, goats, calves, a specific surface focus develops at the injection site with a diameter of

2.0 to 2.5 cm. Heal itself within 15-20 days. The pathogen cannot be isolated mycologically. When a culture of this genus is used on dissected skin of non-immunized animals at a dose of 0.25x10 ⁶ microconidia / cm ³ , a typical trihofitosis focus occurs at the application site. The characteristics of the genus obtained from pathological materials correspond to those of the original species used.

Antigenic properties. After application of the genus of culture, a specific agglutination with a titer of 1: 40-1: 1280 develops in the body of calves, sheep and rabbits.

Immunological properties. After twice intramuscular administration of the vaccine prepared from these species, calves and rabbits, at a dose of 10-20x10 ^ live microconidia, the animals obtained hyperimmunity to subsequent infections with trihofitosis for a minimum of 5 years (monitoring time).

Rod Tr. mentagrophytes VGNKI No.27 differs from epizootic species in low virulence, absence of infection at contact between two animals, high activity in accumulation of microconidia, and immunogenicity in the case of intramuscular injection of animals.

Examples of vaccine preparation of the present invention

Example 1

For the preparation of the vaccine, 15-20 days old cultures of the genera Tr.verrucosum 130L and Tr were cultivated. mentagrophytes VGNKI 27 separately in Roux agar bottles (pH 6.26.8) at 26'-28 ^e C for 12-20 days.

The mycelium of each culture was removed, using special scrapers, from the surface of the nutrient medium under aseptic conditions and placed in sterile containers. The fungus substance of each genus was homogenized in blenders or colloidal mixers and then the Tr homogenate was obtained. verrucosum 130L was resuspended with sterile water in a microconidium concentrate 300x10 <0> / cm3 on the Tr.mentagrophytes homogenate VGNKI No. 27 -600xl0 ^ / cm ^. The homogenate was resuspended in the presence of penicillin or streptomycin at a dose of 100 units / cm homogenate. Homogenate of fungi of the genus Tr. verrucosum 130L was combined in equal volumes with a homogenate of a fungus of the genus Tr. mentagrophytes VGNKI 27. To the resulting suspension was added an equal amount of stabilizer (pH 6.2-6.8) containing 10% sucrose and 1.5% gelatin in water. The whole mass was thoroughly mixed, poured into sterile bottles, dried and lyophilized.

Example 2

The vaccine was prepared according to the procedure of Example 1. It was obtained from the homogenate of fungi of the genus Tr. verrucosum 130L and Tr. mentogrophytes VGNKI No.27 at a concentration of microconidia r o

450x10 / cm. The stabilizer was an aqueous solution of 15% sucrose and 22.5% gelatin.

Example 3

Animal trihofitosis vaccine was prepared by a similar procedure to Example 1 from a homogenate of fungi of the genus Tr. verrucosum 130L at a concentration of microconidia δθθχΐθθ / cm ^ and homogenates of the genera Tr. mentogrophytes VGNKI r o

No.27 at a concentration of microconidia of 300x10 / cm.

The stabilizer contained 20% sucrose and 4% gelatin in water

The reconstituted vaccine was in the form of a homogeneous porous, dry mass, in the form of green colored tablets. When stored in a closed, dry and dark place at 2 'to 10 ^e C, the vaccine retains its biological properties. The vaccine has a shelf life of 12 months. The vaccine is supplied with a bottle of sterile diluent.

Example 4

The series of vaccines obtained in Examples 1, 2 and 3 were diluted 1 dose into 1 cm ³ prior to administration. The inoculative dose of batches of vaccines obtained contained the following

C Q antigen concentrations, N x 10 / cm (Table 1).

Table 1

Antigen The vaccine from Of Example 1 The vaccine from Of Example 2 The vaccine from Of Example 3 Tr.verrucosum 130 L 5 7.5 10 Tr.mentogrophytes No.27 VGNKI 10 7.5 5

The vaccine of Examples 1, 2 and 3 was used to immunize calves by administering two intramuscular doses of 1 cm over a 10-14 day interval.

For comparison, vaccines of the genus Tr verrucosum 130L and Tr were prepared in a similar manner. mentagrophytes VGNKI 27 with a microconidium antigen concentration and tested at a r q inoculative dose of 7.5x10 / cm. 35 days after the second immunization, the calves were infected with a virulent culture of the genus Tr. verrucosum No.153 (TV-153), Tr.mentagrophytes No. 1 251 (TS251), Tr. sarkisovii No. 724 (TS-724), Tr. verrucosum var. autotropheim 118 (TVA-118). At the same time, six non-immunized calves were infected with the same cultures for comparison.

The results of experimental calf infections are summarized in Table 2, below.

Table 2

Gather on St. animals B iopr and rights for ir immune and z and ran j e number nun and z and ran and h calves With t.obo1 e 1i h animals by experimental inf ecc i j i TV- TM- TS- TV- TE- 153 251 724 118 20 1 9 The vaccine from Of Example 1 9 0 0 0 0 0 2 10 The vaccine from Of Example 2 10 0 0 0 0 0 3 7 The vaccine from Of Example 3 7 0 0 0 0 0 4 10 Vaccine of the genus Tr. ve r rucosum 1 30L 10 0 4 2 0 1 5 9 Vaccine of the genus Tr.menta- grophy t es VGNK1 No.27 9 5 0 1 2 2 6 6 Not immune to z and wound calves 2 2 2 2 2 I z Tabe 1 e it can be seen that first, second i n 1 the third series

vaccines develop hyperimmunity (100%) against trihofitosis in experimentally infected animals. Immunization of calves with monocep.ivi fails to provide sufficiently stable immunity in calves infected with heterologous cultures that induce trihofitosis. Non-immunized calves that do not have trihofitosis in their history are susceptible to experimental infection with virulent cultures that cause t r i hof i toza.

Example 5

We used three experimental batches of vaccines, as in Example 4, for immunization of 14 goats, followed by infection with the virulent cultures of the genera Tr.verrucosum No.153 and Tr.mentagrophytes No.251. We did not vaccinate two comparison animals.

None of the vaccinated animals underwent trihofitosis, whereas the comparative two animals showed all symptoms of infection in the form of fertilization of virulent cultures from the focus of infection.

Example 6

Ten rabbits of the chinchila type, weighing 2.5 to 3.0 kg, were immunized with the vaccine according to Example 2. Prior to administration, the vaccine was diluted in a 1 dose / 1 cm diluent.

The concentration of antigens at one inoculative dose was

7.5x10® microconidia in Tr.verrucosum 130L and

7.5x10® microconidia in Tr.mentagrophytes VGNKI No.27.

The vaccine was administered to the rabbit twice intramuscularly at doses of 0.5 cm. In comparison, ten rabbits were vaccinated, also intramuscularly in the hindlimb region, with a vaccine of the genus Tr.mentagrophytes No.135, containing 20x10® microconidia / cm ³ in a single inoculative dose. After 30 days, rabbits were infected epicutaneously with virulent cultures of the genera Tr.mentagrophytes No.251 and Tr.mentagrophytes No.18. The results of the infection were monitored for 25 days. We found that the administration of the vaccine contributed to the development of reliable immunity of all immunized rabbits, while using a known Mentawak vaccine we did not detect a reliable effect: two rabbits out of ten were infected with trihofitosis and virulent cultures the genera have flowed out of the foci of infection in the form of fertilization.

Example 7 calves were immunized with the vaccine version of Examples 1 and

3, with the concentration of antigens in inoculative doses as in

Example 4. The vaccine was administered to each group of calves (groups of 10 animals) intramuscularly, twice, at doses of n 1 cm, every 10-14 days. Four and ten months after the second immunization, both groups of calves were infected with virulent cultures of the genus Tr. verrucosum No.153 and Tr.mentagrophytes No.251. All calves remained healthy. Therefore, the vaccine of the present invention maintains stable immunity even 4 and 9 months after infection.

The vaccine is highly effective against trichophytosis in animals. The vaccine is administered twice a day for professional purposes

About an interval of 10-14 days in the following doses: 1 cm calves from 14 days to 4 months of age and 2 cm calves older than 5

About months; 0.5 cm of sheep (goats, goats) at 3 to 6 months of age and 1 cm 'to 6 months old. Rabbits weighing 2.5 - 3.0 kg are administered 0.3-0.5 cm 'for professional purposes.

For curative purposes, we administer a double dose of the vaccine to sick animals twice in the interval of 10-14 days.

The use of the vaccine of the present invention will increase the efficiency of the control of trihofitosis in animals in veterinary practice.

Claims (1)

PATENT APPLICATION An animal vaccine vaccine containing Trichophyton verrucosum 130L antigen, sucrose, gelatin and water, characterized in that it also contains an Trigen antigen. mentagrophytes VGNKI 27, with the following ratio of components (weight percentages); Ant and genus genus Tr. verrucosum 130L 300-600 χ 10 ⁶ mi crocon and d and j / cm Antigen of the genus Tr. mentagrophytes VGNKI No.27 300-600 χ 10 ⁶ mi crocon and d and j / cm Sucrose 10.0 - 20.0 Gelatin 1.5- 4.0 Water the rest FOR