RU2347584C1 - Vaccine against newcastle disease - Google Patents

Abstract

FIELD: medicine; biotechnologies.

SUBSTANCE: vaccine contains virus material from "La-Sota" strain, saccharose, enzymatic peptone, casein, aerosil AM-1-300 and resin KB-4P-2. At that virus material from "La-Sota" strain used in vaccine is suspension from whole hen's eggs with developed 12-13-day embryos infected with culture of "La-Sota" strain at 9-10 day of their incubation. Vaccine also additionally contains α-tocopherol with the following ratio of components, wt %: suspension from whole hen's eggs with developed 12-13-day embryos infected with culture of "La-Sota" strain at 9-10 day of their incubation - 10.73-12.47, saccharose - 4.16-4.60, enzymatic peptone - 2.50-3.06, casein - 0.83-1.53, α-tocopherol - 0.04-0.08, aerosil AM-1-300 - 1.00-2.00, resin KB-4P-2 - the rest.

EFFECT: high immunogenicity, harmlessness for bird and ecological safety for servicing personnel.

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Description

The present invention relates to the field of biotechnology of viral veterinary drugs, in particular to the vaccine against Newcastle poultry disease OROVAC-NB-2, intended for oral immunization during veterinary measures to combat the specified disease.

Known vaccine against Newcastle disease containing viral material from the La Sota strain, sucrose, enzyme peptone, casein, Aerosil AM-1-300 and KB-4P-2 resin with a certain ratio of components [1].

The disadvantage of this vaccine is its low efficacy in oral immunization of birds, and its use for aerosol use is environmentally unsafe for staff due to the high concentration of aerosil (9% by weight).

Also known is the vaccine against Newcastle disease Orovak-NB containing viral material from the La Sota strain — extraembryonic fluid of chicken embryos, sucrose, enzyme peptone, casein, aerosil AM-1-300 and KB-4P-2 resin with a certain the ratio of components [2].

This vaccine is not sufficiently active during oral immunization, since a significant consumption of a biological product is required to obtain the proper effect, which affects the cost of the measures taken in case of Newcastle disease.

The aim of the invention is to increase the immunogenic activity of the vaccine and its effectiveness.

This goal is achieved by the fact that the vaccine as a viral material from the strain "La Sota" contains a suspension of whole eggs with developed 12-13-day-old embryos infected with the culture of the strain "La Sota" on the 9-10th day of their incubation and additionally α-tocopherol in the following ratio of components, wt.%:

Whole chicken slurry eggs with developed 12-13 day old   embryos infected culture strain "La Sota" at 9-10 day of their incubation 10.73-12.47 Sucrose 4.16-4.60 Peptone enzymatic 2,50-3,06 Casein 0.83-1.53 α-tocopherol 0.04-0.08 Aerosil AM-1-300 1.00-2.00 Resin KB-4P-2 Rest

The invention is illustrated by the following examples.

Example 1

Fertilized chicken eggs are used to obtain the Orovak-NB-2 vaccine against Newcastle disease. On the 9-10th day of incubation, they are infected with an infectious dose of the La Sota vaccine strain culture and continue incubation for 3 days. On the 12-13th day of embryo development, whole chicken eggs in the required quantity are placed in the flask of the RT-1 apparatus for grinding them together with the shell.

Grinding is carried out at 4000-8000 rpm for 1.5-2 minutes.

To 14-15 l of the resulting suspension of viral material add 5-6 kg of sucrose, 3-4 kg of enzymatic peptone, 1-2 kg of casein and 0.05-0.10 kg of α-tocopherol, followed by mixing in the same mode.

A suspension of the viral material with stabilizing substances is placed in a ball drum, in which it is mixed and dehydrated, in a 1: 4 ratio, with KB-4P-2 ion-exchange resin coated with hydrophobic aerosil AM-1-300, taken by weight, onto the surface of its particles in the amount of 1.25-2.50%.

The mixing of these components and obtaining a dry biological product is carried out in the mode of operation of the speed of rotation of the ball drum on the break-in machine, equal to 60-90 rpm, for 5-7 minutes

Prepare 3 composition of the proposed vaccine. The composition of the variants of the vaccine "Orovak-NB-2" are shown in table 1.

The vaccine is packaged in 100-200 g in sterile bottles or double plastic bags. The vials are sealed with sterile rubber stoppers and wrapped in aluminum caps, and the bags are hermetically sealed. The shelf life of the vaccine is 12 months, provided that it is stored in a dark and dry room, at a temperature of 6 ± 2 ° C.

The finished vaccine "Orovak-NB-2" is a loose powder of gray color. The biological activity of the drug is 8-8.6 lg EID ₅₀ / g ^-1 , the mass fraction of moisture in the range of 10-14%. In 1.0 g of the biological product contains 14-18 doses.

Example 2

The biological activity of 3 formulations of the Orovak-NB-2 vaccine, prepared according to Example 1 and stored for 3, 6 and 12 months at a temperature of 6 ± 2 ° C and minus 10 ± 2 ° C, is studied.

Biological activity is determined by titration on 9-10-day-old chicken embryos and expressed in lg EID ₅₀ / g ^-1 . The specificity of the REC response is determined by setting the hemagglutination reaction.

The results of the study of the biological activity of the vaccine "Orovak-NB-2" depending on storage conditions are presented in table 2.

From the data of table 2 it follows that the biological activity of the vaccine is maintained at the initial level for 12 months at a storage temperature of minus 10 ± 2 ° C, and a temperature of 6 ± 2 ° C contributes to a slight decrease by 0.15-0.25 lg EID ₅₀ / g ^-1 .

Example 3

Evaluation of immunogenicity 2 of the composition of the vaccine "Orovak-NB-2", prepared according to example 1, with an initial biological activity of 8.0 lg EID ₅₀ / g ^{-1 is} carried out on 40 broilers 12-14 days of age. Ten-fold dilutions of the vaccine are prepared successively.

The vaccine dose is fed for 2 days, once a day (in the morning at the rate of 5 g of feed per 1 goal.). Immunization efficiency is established after 7, 14 and 21 days according to antibody titers detected in rtga and according to the results of the control infection of vaccinated chickens with a culture of virulent strain "T-53" of the NB virus at a dose of 3.0 lg LD ₅₀ / cm ³ .

The results of immunization of broiler chickens with Orovak-NB-2 vaccine at a dose of 6-7 lg EID ₅₀ / g ^-1 / goal provide reliable protection of chickens from infection control.

Then, according to the Reed-Mench method, the dose most suitable for use is calculated that provides protection of at least 90% of the livestock from the control infection dose, which in this case is 6.8 · 10 ^6.0 EID ₅₀ / g ^-1 (6.83 lg) or 71 mg / goal.

Example 4

Explore the harmlessness of 1 composition of the vaccine "Orovak-NB-2" prepared according to example 1.

The vaccine is fed to chickens 20-270 days of age (30 goals in each age group), in a titrated 10-fold dose (7.83 lg EID ₅₀ / g ^-1 or 710 mg per chicken). The chickens are preliminarily kept on a hungry diet for 8 hours. The vaccine is fed 2 days in a row in the morning feeding. Observation of the bird is carried out for 10 days, carry out selective weighing of chickens with 5 goals in each group. All experimental chickens remained alive.

Establish the absence of signs of toxicosis and residual virulence of the vaccine. The clinical condition of experimental chickens did not differ from control intact ones. An increase in live weight in experimental chickens by 1-2% is noted in comparison with the control.

Example 5

A study of the immunogenic properties of the 3 composition of the vaccine "Orovak-NB-2" prepared according to example 1, as well as two samples of this vaccine, stored for 3 and 12 months at a temperature of 6 ± 2 ° C.

Test 3 samples of the vaccine is carried out similarly as described in example 3 on groups of broiler chickens 12-14 days of age (100 goals in each group). All chickens are inoculated with test samples of the vaccine in a single dose of 6.8 · 10 ^6.0 EID ₅₀ / g ^-1 .

The results are presented in table 3.

The results of the table indicate that all tested samples of the proposed vaccine are highly immunogenic: the chickens ’immune response to the Orovak-NB-2 vaccine reaches its maximum 2-3 weeks after vaccination (the antibody level on day 21 ranges from 5.1-5.8 log ₂ ), and the immunizing dose of the vaccine used provides protection in all groups of 100% of the chickens from the control of infection with the culture of the virulent strain “T-53”.

Orovak-NB-2 vaccine from La Sota strain, manufactured by contact dehydration technology for oral administration, is harmless to poultry, does not cause post-vaccination reactions, delayed onset of laying and dynamics of increase in chicken productivity; highly immunogenic (protection from the NB virus by 80-100%) and environmentally friendly for staff.

The use of a suspension of whole chicken eggs with developed 12-13-day-old embryos infected with the La Sota strain culture for 9-10 incubations as a virus-containing material allows to increase vaccine production, provide the bird with additional protein-mineral fertilizing, and increase productivity poultry and thereby reduce the economic costs of waste-free vaccine production and veterinary measures to prevent Newcastle disease poultry by reducing the vaccination dose.

Table 1 Components The content of components, wt.% Composition 1 Composition 2 Composition 3 Suspension of whole chicken eggs with developed 12-13-day-old embryos infected with the culture of the La Sota strain on the 9-10th day of their incubation 10.73 11,565 12.47 Sucrose 4.60 4,387 4.16 Peptone enzymatic 3.06 2,792 2,50 Casein 1,53 1,196 0.83 α-tocopherol 0.08 0.06 0.04 Aerosil AM-1-300 1.00 1,5 2.0 Resin KB-4P-2 Rest Rest Rest

table 2 The vaccine "Orovak-NB-2" The biological activity of the compounds, Ig EID ₅₀ / g ^-1 After cooking Storage at temperature (° С) for, month 3 6 12 6 ± 2 Minus 10 ± 2 6 ± 2 Minus 10 ± 2 6 ± 2 Minus 10 ± 2 Composition 1 8.6 8.45 8.6 8.35 8.6 8.35 8.6 Composition 2 8.0 7.9 8.0 7.85 8.0 7.8 8.0 Composition 3 8.5 8.45 8.6 8.32 8.5 8.35 8.5

Table 3 Vaccine dilution Dose of the vaccine, lg Geometric mean antibody titer (log2) Control infection results 7 days 14 days 21 days Palo (goal) Survived (goal) Protective effect (%) 10 ^-1 7 0.4 7 8 0 5 one hundred 10 ^-2 6 0.4 four 4.8 one four 80 10 ^-3 5 0 1.8 2 3 2 40 10 ^-4 four 0 1,2 1.8 3 2 40 10 ^-5 3 0 0.2 0.2 5 0 0 10 ^-6 2 0 0 0 5 0 0 10 ^-7 one 0 0 0 5 0 0

Table 4 The shelf life of the vaccine, months Geometric mean antibody titer (log2) Control infection results 7 days 14 days 21 days Palo (goal) Survived (goal) Protective effect (%) Freshly cooked 2.5 4.8 5.8 0 one hundred one hundred 3 2.5 4.4 5,6 0 one hundred one hundred 12 2.0 4.1 5.1 0 one hundred one hundred

Information sources

1.SU 1785098 A, 08/10/1995.

2. RU 2159129 C, 03/31/1999.

Claims (1)

Newcastle disease vaccine containing viral material from the La Sota strain, sucrose, enzyme peptone, casein, Aerosil AM-1-300 and KB-4P-2 resin, characterized in that it is a virus material from the La strain Sota "contains a suspension of whole chicken eggs with developed 12-13-day-old embryos infected with the culture of the La Sota strain on the 9-10th day of their incubation, and additionally α-tocopherol in the following ratio, wt.%:
Suspension of whole chicken eggs with developed 12-13-day-old embryos infected culture strain "La Sota" on day 9-10 their incubation 10.73-12.47 Sucrose 4.16-4.60 Peptone enzymatic 2,50-3,06 Casein 0.83-1.53 α-tocopherol 0.04-0.08 Aerosil AM-1-300 1.00-2.00 Resin KB-4P-2 Rest