RU2562585C1 - Method for increasing biocidal and therapeutic efficacy of staphylococcal anatoxin vaccine - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: method is implemented by using a liquid synthetic nutrient medium for Staphylococcus growth, autoclaving the microorganisms and detoxifying in glutaric aldehyde solution and aethonium solution and activating the biopreparation 10 mg/l with colloidal silver ions.

EFFECT: higher biocidal and therapeutic efficacy of the Staphylococcal anatoxin vaccine.

4 tbl, 6 ex

Description

The invention relates to microbiology and biotechnology.

Known methods for increasing the production of staphylococcal inactivated vaccines for humane medicine and antistaphylococcal drug (ASP) manufactured by the Ignatov Research Center (Reference "Vidal Veterinarian". 2013, 56-58; A.I. Korotyaev, S. A. Babichev. Medical Microbiology, Immunology, Virology. St. Petersburg, 1998, pp. 330-335; A.I. Vorobyov. A Practitioner Handbook. M .: Medicine, 1990, p. 321-327).

A significant drawback of these drugs is the cultivation of production staphylococci strains on meat-hydralizate glycerin broth, the need to supply carbon dioxide, the use of a 0.5-0.7% formaldehyde solution for detoxification and polymerization of a complex of staphylococcal anatoxins, which has not only carcinogens, but also does not provide the completeness of detoxification of staphylococcal and other bacterial toxins. It should be noted that the name Ignatov's antistaphylococcal drug does not reflect the generally accepted terminology - toxoid or vaccine. In addition, the concentration of each of the 3 strains of staphylococci 10 MK / ml is very high, because on meat and peptone broth, the maximum accumulation is 3-5 billion / ml and no more, and the use of meat and peptone agar is technologically not profitable and not safe when working with live microorganisms.

The prototype is a method of producing a staphylococcal toxoid vaccine (RF patent No. 23377014, publ. 12/27/2009), which includes growing staphylococci on a liquid synthetic medium in 2-liter bottles for 10-12 days to 11-12 billion / ml suspension of staphylococci with subsequent autoclaving, detoxification and polymerization, first with a 0.2-0.3% solution of glutaraldehyde at 40 ° C for 2-3 days, and then with a 0.2-0.3% solution of ethonium for 2-3 days and sorption on 3-5 mg / ml of aluminum hydroxide.

The disadvantage is the lack of biocidal action against a number of microorganisms and therapeutic efficacy in purulent-necrotic skin lesions, dermatitis, eczema in cows with mastitis.

An object of the invention is to increase the biocidal and therapeutic effect of staphylococcal toxoid vaccine by activating a suspension (suspension) and a complex of staphylococcal toxins by colloidal silver ions.

The technical result is achieved by using staphylococci growing liquid synthetic nutrient medium, autoclaving microorganisms and detoxifying and polymerizing a complex of toxins initially with a 0.2-0.3% solution of glutaraldehyde at 38-40 ° C for 2-3 days, and then 0 , 3-0.5% ethonium solution, followed by activation of a biological product solution of 10 mg / l colloidal silver ions.

The results are presented by manufacturing technology and examples for the study of biocidal and therapeutic properties of a biological product. The manufacturing technology of the toxoid vaccine contains the following steps:

- the cultivation of Staphylococcus aureus on a liquid synthetic nutrient medium in 2-liter bio-bottles for 10-12 days at 37 ° C to a concentration of 11.0 ± 1.0 billion microbial bodies in 1 ml;

- autoclaving a suspension of staphylococci at 1.0 atm for 30 minutes;

- the first detoxification of staphylococcal toxins with a 0.2-0.3% solution of glutaraldehyde at 40 ° C for 2-3 days;

- the second detoxification of a complex of staphylococcal exo-, endo and superenterotoxins with 0.3-0.5% ethonium solution at 40 ° C for 2-3 days;

- activation of a suspension of staphylococci in combination with anatoxin 10 mg / l of silver ions and sorption of the biological product on aluminum hydroxide;

- packaging of the drug in vials and sterilization at 1.0 atm for 30 minutes.

The obtained staphylococcal toxoid vaccine was used to study the effectiveness of the prevention and treatment of burn and lacerations, mastitis in cows, dermatitis and trophic ulcers of carnivores.

The proposed technology for the production of staphylococcal toxoid vaccine differs from existing ones by growing staphylococci in a liquid synthetic nutrient medium instead of meat peptone glycerin broth and completely detoxifying and inactivating exo-, endo, and superenterotoxins initially with a 0.2-0.3% solution of glutaraldehyde at 40 ° C for 2-3 days, and then with 0.3-0.5% ethonium solution at 40 ° C for 2-3 days.

Example 1. Determination of the harmlessness of staphylococcal toxoid-acin (CAB)

The study of CAB toxicity was performed on 12 white mice weighing 20 + 2.0 g by double intraperitoneal (6 goals) and subcutaneous (6 goals) administration of 0.5 ml of a biological product with an interval of 3 days.

All animals without visible necrotic, inflammatory processes and general toxicosis remained alive when observed for 10 days.

Example 2. The test of harmlessness in dogs

For research, 8 outbred puppies at the age of 2-3 months weighing 3-4 kg were used. The experimental animals were injected 5 ml CAB with an interval of 5 days twice subcutaneously (4 goals.) And intramuscularly (4 goals.) Within 12 days, all animals remained alive, and at the injection site of the biological product there were no inflammatory and purulent-necrotic lesions.

Example 3. A comparative assessment of the biocidal action of the control and experimental toxoid vaccine without aluminum hydroxide are shown in table 1,

where ID is the content of infectious doses in 1 ml of culture and embryonic fluid, "+" - biocidal effect, "-" - does not work.

From the data presented in table 1, it follows that the control CAB with a 0.1% solution of glutaraldehyde and a 0.2-0.3% solution of ethonium biocidal effect in a dilution of 1: 1 and 1: 2, and CAB with 10 mg / L silver ions with 0.1% glutaraldehyde solution and 0.2-0.3% ethonium solution shows a biocidal effect in a dilution of 1: 1,1: 2, 1: 8 and 1:16 .

Example 4

Comparative evaluation of the effectiveness of treatment of burn wounds in dogs

A skin burn in a diameter of 5-6 cm in 12 outbred puppies aged 2-2.5 months was carried out by applying for 4-5 minutes several layers of gauze moistened with a 5% solution of carbolic acid, followed by washing with water. The treatment was carried out with a control CAB and an experimental CAB with 10 mg / L silver ions on the 2nd day after a skin burn with carbolic acid, first by washing and then by applying a tissue towel impregnated with CAB.

The results are presented in table 2.

From the results it follows that in the treatment of burn wounds in dogs with an experimental staphylococcal toxoid vaccine with colloidal silver ions, the wound healing process was completed 2-3 days earlier compared to the control staphylococcal toxoid-vaccine.

Example 5

A comparative assessment of the effectiveness of the control and experimental toxoid vaccine with silver ions in the treatment of cows with mastitis is shown in table 3.

From the data presented in table 3, it follows that the treatment time for 36 cows with mastitis, experimental toxoid vaccine with colloidal silver ions was halved.

Example 6

A comparative assessment of the treatment of dogs with dermatomycosis, control and experimental toxoid vaccine with colloidal silver ions is shown in table 4.

It is known that trichophytosis favus (scab) belongs to dermatomycosis under the general nosological form of "lichen". The causative agents of dermatomycosis are imperfect molds Asp. flavus and Asp. niger.

From the presented data it follows that the treatment of dogs with dermatomycosis with lesions up to 5-7 cm in 2-3 places of the skin with the help of an experimental toxoid vaccine with silver ions accelerated the healing of the affected skin by 4 days compared to the control toxoid a vaccine.

Claims (1)

A method of increasing the biocidal and therapeutic effectiveness of a staphylococcal toxoid vaccine, including growing staphylococci in a liquid synthetic medium, autoclaving, detoxifying a complex of toxins initially with a 0.2-0.3% solution of glutaraldehyde at 40 ° C for 2-3 days, and then a 0.3-0.5% solution of etonium followed by activation of a biological product solution of 10 mg / l with colloidal silver ions.