RU2378011C1 - Method for prophylactics of cattle brucellosis - Google Patents

Abstract

FIELD: veterinary science.

SUBSTANCE: invention relates to veterinary science immediately dealing with prophylactics of brucellosis infection. Under the method proposed colostral age calves receive an intramuscular injection of brucelline in an amount of 2.5-3 ml and 30-40 minutes afterwards - an intramuscular injection of 4-6 ml of a immunomodulator containing, wt %: formaldehyde - 0.07-0.12, sodium chloride - 0.9-0.95 and distilled water - balance.

EFFECT: proposed method provides for efficient prophylactics of cattle brucellosis.

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Description

The invention relates to the field of veterinary medicine and, in particular to the prevention of brucellosis infection.

Brucellosis is a zoonotic infection with a high potential for transition to a chronic form.

The use of antibiotics for the treatment of acute and chronic forms of brucellosis is known. In particular, there is a known method for the prevention of brucellosis, including oral administration of an antimicrobial preparation to an animal together with an immunostimulant and / or low molecular weight nucleic acids (RF patent No. 2238759, IPC A61K 39/10, publ. October 27, 2004). Various antibiotics are used as an antimicrobial drug (rifampicin, tetracycline, chloramphenicol, streptomycin and others), and as an immunostimulant, an immunophore is used, which has the ability to stimulate various immune systems (B-system and phagocytes), increase and activate the growth and development of animals.

However, antibiotic courses, even in combination with an immunostimulant, do not always lead to a positive result, since the causative agent of brucellosis, being intracellular, is reliably protected by the cell membrane from the antibiotic circulating in the body. In addition, prolonged use of antibiotics in therapeutic doses causes a number of side effects. The use of the immunophore drug leads to an increase in the phagocytic immunity, which leads to incomplete inactivation of brucella, the preservation of the pathogen in a viable state, and its persistence in the body.

For the prevention of brucellosis, various vaccines are widely used - live and inactivated. In particular, there is a known method for the prevention of cattle brucellosis with an antigen based on the protein-polysaccharide complex of brucella (see RF patent No. 2037319, IPC A61K 39/10, publ. 06/19/1995), which is a suspension of cells of the vaccine strain of Br.abortus 19 brucella -BA in S-form, treated with acetone. The protein-polysaccharide complex is administered to calves aged 3-5 months once in a dose of 5-35 mg in 5 ml of a 0.9% sodium chloride solution.

However, the use of the vaccine causes undesirable side symptoms, such as reactogenicity, sensitization, agglutinogenicity, low protective activity.

Closest to the claimed is a method for the prevention of brucellosis, which consists in immunizing animals with a standard vaccine from strain Br. Abortus 82 and the introduction after 14-21 days of immunomodulators, which are used as solutions of T- and B-activin, which are administered once for 3 days (see RF patent No. 2065749, IPC A61K 39/10, publ. 27.08. 1996). After 12 months, serologically negatively reacting animals are revaccinated with the same vaccine in the same dose. The method allows to enhance the immunogenic properties of the vaccine from strain 82.

However, the method is time-consuming, requires careful execution, which in practice is difficult to implement. In addition, the immunity created in animals is not stable, it is only 12 months.

The invention is aimed at solving the problem of creating an economical, easy-to-carry out, effective method for the prevention of brucellosis, which provides a significantly faster immune level of animals from infection in a significantly short time.

To solve the problem in the method for the prevention of brucellosis, which consists in the intramuscular administration of a prophylactic drug and an immunomodulator, according to the invention, brucellin is used as a prophylactic drug, and as an immunomodulator, a drug containing, wt.%: Formaldehyde - 0.07-0.12 and sodium chloride - 0.9-0.95, distilled water - the rest, while first brucellin is administered once to young colostrum youngsters at a dose of 2.5-3 ml per head, and after 30-40 minutes, an immunomodulator is administered in doses 4-6 ml per head.

Known sources of patent and scientific and technical information do not describe a method for the prevention of brucellosis, which allows for effective protection in a significantly short time and provides a high level of immunity due to the use of brucellin as a prophylactic agent, as an immunomodulator of a formaldehyde-based drug.

In contrast to the known analogues in the present method, instead of the vaccine from strain Br.Abortus 82, brucellin is used.

By its nature, brucellin is a sterile biological product used for the allergic diagnosis of brucellosis; it is a filtrate of a 30-day-old culture of three types of brucella.

Typically, brucellin is administered to animals either under the skin of the eyelid (palpebral test) or intradermally in the center of one of the papillary folds (intradermal test).

In the inventive method, brucellin is administered intramuscularly.

An immunomodulator (formaldehyde-based drug) belongs to a new class of drugs that contribute to the activation of an antigen-specific immune response, the development of which eliminates the infectious agent. The drug causes the production of alpha and beta interferons, mobilizes and activates macrophages, limits the production of inflammatory cytokines, stimulates the production of antibodies to various antigens of an infectious nature, and increases the body's resistance.

The authors revealed for the first time that intramuscular administration of brucellin followed by (30-40 minutes) administration of an immunomodulator promotes the activation of complete phagocytosis in animals, which is not obvious.

All vaccines known to date are based on the activation of the lymphocytic immunity, and phagocytosis is considered as a nonspecific response of the macroorganism to the introduction of the pathogen.

In the claimed invention, the problem of forming a high level of immunity of the animal organism on the basis of activation of completed phagocytosis, increasing the functional activity of neutrophils, as well as changing the ratio of aerobic and anaerobic oxidation processes towards aerobic is solved.

The introduction of these drugs leads to the absence of allergic and serological reactions.

The foregoing allows us to conclude that there is a criterion of "inventive step" in the claimed invention.

The method is as follows.

Cows of colostrum age (from 2 to 9 days) are injected intramuscularly into the croup with 2.5-3 ml of brucellin in a concentration used for allergic diagnosis of brucellosis (Manual on the use of brucellin VIEV for allergic diagnosis of animal brucellosis, approved by the Department of Veterinary of the Ministry of Agriculture of Russia on 26.02. 96 g.).

After 30-40 minutes, calves are injected intramuscularly with 4-5 ml of an immunomodulator containing formaldehyde, sodium chloride and distilled water.

Both drugs are administered to animals once.

Brucellin working solution is a brownish-yellow clear liquid without opalescence, containing metabolic products and specific substances.

The immunomodulator is a colorless transparent liquid with a faint smell of formalin, it is prepared according to TU 9336-001-00670551-2005.

All subsequent brucellosis prophylaxis measures are carried out in accordance with Veterinary Rules VP 13.3.1302-96 approved by the Veterinary Department of the Ministry of Agriculture of the Russian Federation on June 18, 1996 (In the collection of sanitary and veterinary rules “Prevention and control of infectious diseases common to humans and animals ", Publishing house of the State Committee for Sanitary and Epidemiological Supervision of Russia and the Ministry of Agriculture and Food of Russia, Moscow, 1996. - P.20).

The effectiveness of the method was evaluated by the results of phagocytosis, the functional activity of neutrophils, biochemical parameters, the results of serological reactions (agglutination reaction (RA), complement fixation reaction (CSC) and immunodiffusion reaction (RID)), provided for by the above rules and by direct infection of animals with virulent culture B. abortus.

Below are examples that confirm the activation of the phagocytosis process, an increase in the functional activity of neutrophils, and an increase in the aerobic oxidation processes in the calf’s body.

Example 1, proving the activation of the phagocytosis process in animals.

Studies were conducted on 15 rats and 12 calves.

The animals were first injected with brucellin, and then after 30-40 minutes an immunomodulator. The drugs were administered intramuscularly, once.

In this case, brucellin was administered to rats at a dose of 0.1 ml, and an immunomodulator at a dose of 0.5 ml per head. Brucellin was administered at a dose of 3 ml to calves aged 2-9 days, and an immunomodulator at a dose of 5 ml per head.

Saline was injected with physiological saline: 0.6 ml in rats and 8 ml in calves.

The effectiveness of the process of bacterial phagocytosis in neutrophils was evaluated by indicators: phagocytic activity (% phage), phagocytic index (PI), phagocytic number (PS) and phagocytosis completion index (PPI).

The results of the study of phagocytosis are presented in table 1.

Table 1
Rat rats phagocytosis 28 days after administration of brucellin with an immunomodulator Characterization of animal groups Phagocytosis PS (abs.) % Phage FI (abs. Value) CONTROL 21.0 ± 2.0 79.5 ± 3.2 26.2 ± 2.0 EXPERIMENTAL (injected brucellin with immunomod.) 6.2 ± 1.9 80.6 ± 5.7 7.4 ± 2.0

It was found that 28 days after the administration of white rats with brucellin with an immunomodulator in immunized animals, the PS (from 21.0 ± 2.0 to 6.2 ± 1.9) and PI (from 26.2 ± 2 , 0 to 7.4 ± 2.0). The percentage of neutrophils involved in phagocytosis of brucella (FA) was similar to the control (80.6 ± 5.7% in the experiment and 79.5 ± 3.2 in the control).

At the same time, the completeness of brucella phagocytosis was recorded in the experimental groups.

The results of microscopic studies of rat blood showed that the number of brucella in the rat treated with these drugs compared with the control in the field of view is minimal.

Similar results on the activation of the phagocytosis process were obtained on calves. In animals, at the same time as in rats (after 28 days), after the administration of drugs (brucellin with an immunomodulator), a tendency toward a decrease in PS and PI in the process of brucella phagocytosis by neutrophils with a constant value of FA was observed.

Microscopic studies of the blood of calves treated with these drugs also showed the absence of the causative agent of brucellosis.

Example 2, confirming the increase in the functional activity of neutrophils in the blood of calves.

Blood tests of calves were carried out, which were administered intramuscularly once at the age of 2-9 days: brucellin at a dose of 3 ml and after 30 minutes an immunomodulator. Blood tests were performed on days 7, 15, and 35 after drug administration.

The results are presented in table 2.

table 2
Functional activity of calf blood neutrophils Indicators 7 days 15 days 35 days Counter. Experience Counter. Experience Counter. Experience Functional neutrophil activity: Stimulation Index 0.32 ± 0.04 0.34 ± 0.01 3.09 ± 0.44 5.36 ± 0.32 0.13 ± 0.11 0.57 ± 0.16

From table 2 it is seen that after the administration of drugs on days 15 and 35 there is a significant increase in the functional activity of neutrophils in the blood of the experimental group of calves as compared with the control, which indicates the possibility of complete destruction of the pathogen in the peripheral blood of animals.

Example 3, confirming the enhancement of aerobic oxidation processes in the body of calves.

28 days after the administration of the drugs, the biochemical blood parameters of the blood of the calves of the experimental and control groups described in Example 1 were studied. The results are presented in Table 3.

Table 3 Biochemical indicators of blood calves Characterization of animal groups Biochemical parameters ALT ACT QC LDH Protein Control 1.92 ± 0.5 16.92 ± 1.0 174.8 ± 10.1 1717.95 ± 114.4 71.85 ± 1.4 Experienced (processed: brucellin + immunomodulator) 7.43 ± 0.5 7.43 ± 0.5 263.80 ± 46.5 1890.52 ± 331.9 40.83 ± 3.0

Summarizing the results of biochemical studies (absolute indicators and their correlation with each other), we can assume that the introduction of brucellin with an immunomodulator enhances the aerobic oxidation processes in the body of immunized calves.

To confirm the specificity of the formed resistance of the organism to the causative agent of brucellosis, serological bacteriological and biological studies of experimental and control animals were carried out according to generally accepted methods.

In the blood serum of immunized calves, there was no increase in the level of specific antibodies in agglutination reactions, complement binding and immunodiffusion. These results indicated the absence of activation of the lymphocytic immunity.

Virus culture of B. abortus was infected with laboratory animals, control and treated with these drugs according to our scheme.

In the experiments, 10 rats were used. None of the animals treated with these drugs were infected.

Bacteriological studies confirmed the absence of the causative agent of brucellosis in the internal organs of the treated animals.

All control animals became ill; a virulent B. abortus culture was isolated from them.

Thus, the claimed method for the prevention of brucellosis in cattle has allowed to create a high level of animal immune defense to the causative agent of brucellosis by activating completed phagocytosis, increasing the functional activity of neutrophils and enhancing aerobic oxidation processes.

Claims (1)

A method for the prevention of cattle brucellosis, including intramuscular administration of a prophylactic drug and an immunomodulator, characterized in that brucellin is used as a prophylactic drug, and a drug containing, wt.%: Formaldehyde 0.07-0.12, sodium chloride as an immunomodulator 0.9-0.95, the rest is distilled water, with the first administration of brucellin once young colostrum youngsters at a dose of 2.5-3 ml per head, and after 30-40 minutes an immunomodulator is administered at a dose of 4-6 ml per head.