RU2378013C1 - Method for preparation of virus vaccine against bird newcastle disease (for oral application) - Google Patents

Abstract

FIELD: veterinary science.

SUBSTANCE: invention relates to veterinary virology. Under the method proposed one grows a vaccine strain of "La-Sota" virus on chicken embryos. After that pearl-barley, heat treated, is mixed with allantoic fluid containing the vaccine strain, soaked with a stabiliser solution represented by a mixture of 40% peptone and 8% lactose, frozen at a temperature of minus 50±20°C during 8-10 hours and lyophilised during 30-32 hours.

EFFECT: proposed method enables production of an oral application vaccine for bird immunisation that is characterised by high immunological efficiency and storage stability.

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Description

The invention relates to the field of veterinary virology, and in particular to a virus vaccine against Newcastle disease (NB) in birds intended for oral use.

In the context of the economic development of poultry farming, mass immunization with the oral method significantly reduces labor costs and losses from post-vaccine stresses that adversely affect the health and productivity of the bird.

Particularly relevant and promising in the implementation of the oral method of bird immunization against NB are dry virus vaccines that can create a stable immunity to the disease when added in small quantities to the feed. Their advantage is compatibility with various poultry feeding technologies in any economic conditions.

A known method of manufacturing a vaccine against NB birds based on lactose granules from strain V-4 (Veter. Microbiol. - 1995, vol. 46, No. 1/3-p. 47-53).

In addition, a technology was developed for the production of granulated virus vaccine against bird NB for oral administration based on lactose granules (RF patent No. 2153356, 07.27.2000), which consists in spraying allantoic fluid in chicken embryos infected with pieces. "La Sota virus NB, on lactose agglomerates."

The disadvantage of this method is that to obtain a virus vaccine against NB birds for oral use, lactose agglomerates specially prepared for this purpose are used.

A known method of producing a virus vaccine based on heat-treated crushed barley and sorghum grains (M. Nasser, J.E. Johr, Y. V. Mebraty at all. Oral Newcastle disease Vaccination trials in Ethiopia - Avian Pathologe, 2000, 29, p. 27-34).

For this purpose, the grain was poured with boiled water and boiled for 2 minutes, cooled, the water was drained. After 5 minutes, the grain was cooled under running tap water, and then dried in the open air for 48 hours. They were impregnated with water during the day, after which it was drained. The impregnation process was repeated, kept overnight. The water was drained, the grain was dried in the open air for 48 hours. After that, an allantoic fluid (amniotic fluid of chicken embryos) containing the virus diluted 10 times with water was added to the grain in relation to 1 part of allantoic fluid and 4 parts of grain. The chickens were fed the prepared mixture immediately and 6 hours later at the rate of 10 g of vaccine per head. After immunization with a freshly prepared mixture, 100% of the chickens were resistant to control infection, and 95% were vaccinated with the mixture 6 hours after its preparation. Sorghum turned out to be unsuitable for these purposes - only 30% of the chickens were resistant to control infection, received a mixture of virus-containing allantoic fluid - grain immediately after its preparation.

The disadvantages of this method is to obtain a vaccine preparation based on crushed barley, which retains immunogenic properties for no more than 6 hours after its preparation - this time is not enough for a preliminary check of the infectious activity, safety, immunogenicity of the vaccine due to their duration in time, because of the shortest operation - determination of infectious activity - takes at least 92 hours. The above does not allow the use of the drug for its intended purpose in poultry farms.

The aim of the present invention is to develop a method of virus vaccine against Newcastle disease of birds for oral immunization of birds with high immunogenic efficiency, long shelf life.

This goal is achieved by the fact that heat treated pearl barley is mixed with allantoic fluid containing the La Sota vaccine strain of the Newcastle disease virus, soaked in a stabilizer solution, in the form of a mixture of 40% peptone and 8% lactose, frozen minus 50 ± 20 ° C for 8-10 hours and lyophilized for 30-32 hours.

The addition of pearl barley provides the adsorption of the vaccine virus on its surface and in the pores, obtaining the vaccine in the form of granules, and delivering the virus to the gastrointestinal tract.

Examples of specific performance.

Pearl barley weighing 1.0 kg is poured into a metal container with a volume of 3.5-4.0 l, pour 2.5 l of tap water and boil for 40 minutes over low heat with occasional stirring to avoid burning. It is washed for 3-5 minutes with tap water, poured with the same volume of distilled water and boiled for 40 minutes. Washed with distilled water. The same procedure is repeated a third time, pouring the cereal and rinsing with saline. The groats are placed in an oven, distributed in a thin layer and dried for 3-4 hours at a temperature of 130-140 ° C.

Stabilizers are prepared on distilled water — a 4% solution of lactalbumin hydrolyzate and a mixture of 40% peptone and 8% lactose. The stabilizer is sterilized by autoclaving at 1.2-1.5 ati for 1 hour.

Example 1

The virus vaccine is prepared as follows: 10.0 g of heat-treated pearl barley, soaked in a mixture of 5.0 ml of 40% peptone and 8% lactose, is poured with 10 ml of the allantoic fluid of the La Sota vaccine strain of bird NB virus grown on 9-10 day old chicken embryos for 92 hours at a temperature of 37 ± 0.5 ° C and a relative humidity of 60-70%. The mixture is applied to the pallet with a uniform layer of 3.0-5.0 mm high, frozen at minus 50 ± 2 ° C for 8-10 hours and lyophilized.

Example 2

The virus vaccine is prepared as follows: 10.0 g of pearl barley, heat-treated with 10.0 mm allantoic fluid of the La Sota vaccine strain NB virus grown on 9-10 day old chicken embryos for 92 hours at 37 ± 0.5 ° C and relative humidity 60-70%. The mixture is applied to the pallet with a uniform layer of 3.0-5.0 ml high, frozen at minus 50 ± 2 ° C for 8-10 hours and lyophilized. Lyophilization is carried out in a sublimation unit. After loading the drying chamber, a material temperature control sensor is connected, the chamber is sealed and vacuumized to a residual pressure of 80-100 mm Hg. Then turn on the circulation pump and during the first 10-14 hours of drying the heating of the shelves is carried out by natural heating to minus 20 ° C. The temperature of the material during the first 10-12 hours should not exceed minus 30-32 ° C. Further, the heating of the shelves in steps (2-3 hours) is increased to minus 15; 10; 5 ° C and plus 5, 10 and 30 ° C. The final temperature of the shelves (30 ° C) is maintained for 3 hours, while the temperature of the material should not exceed 20 ° C. Dosage of the vaccine is carried out at a temperature of the material plus 15-20 ° C for 5 hours. The drying time is 30-32 hours with a residual moisture content of the finished product of 2-3%.

The biological activity of the obtained virus vaccine sample No. 1 was 9.5 lg EID ₅₀ / g, sample No. 2 - 9.75 lg EID ₅₀ / g

Virus vaccine of both samples was sterile against bacterial and fungal microflora, harmless when administered intramuscularly and feeding a five-fold dose of the vaccine to chickens mixed with the daily feed rate, retained biological and immunogenic properties for 8 months of storage for sample No. 1 and 3 months for sample No. 2 at a temperature of 4-6 ° C and provided protection for 100% of vaccinated chickens when infected with virulent strain "T-53" of the NB virus intramuscularly at a dose of 1000 LD ₅₀ .

The table shows the immunogenic properties of the anti-NB virus vaccine for oral use.

The use of the virus vaccine against NB of birds from the La Sota strain for oral administration is ensured by preserving the biological activity, immunogenicity and effectiveness of the vaccination method compared to the dry vaccine from the La Sota strain (STO 00495549-0015-2006) used by aerosol , intranasal and oral (drink) methods. However, drinking this virus vaccine with drinking water creates certain practical difficulties, which reduces the quality and effectiveness of vaccination.

The data obtained indicate that the proposed method allows to obtain a vaccine for oral immunization of birds with high immunological effectiveness, stable when stored in conditions of 4-6 ° C for 8 months.

Table 1
The biological activity of the anti-NB virus vaccine from the La Sota strain for oral use The activity of the initial allantoic fluid lg EID ₅₀ / cm ³ The biological activity of the virus vaccine, lg EID ₅₀ / g Shelf life, month Samples one 2 10.5 0 10.0-10.25 10.5 1,5 10.0 10.25 2 9.5-9.75 9.75 chicken experience 3 9.5 9.75 6.0 9.5 chicken experience 8.25 7 9.5 8.0-8.25 8 9.5 8.0 13 8.5 7.0-7.5

Claims (1)

A method of manufacturing a vaccine against Newcastle disease of birds, comprising growing a vaccine strain of La Sota virus on chicken embryos, characterized in that heat treated barley barley is mixed with allantoic fluid containing the La Sota vaccine strain of Newcastle disease of birds, they are impregnated with a stabilizer solution in the form of a mixture of 40% peptone and 8% lactose, frozen at minus 50 ± 20 ° C for 8-10 hours and lyophilized for 30-32 hours