Strain "52/70-m" of bursal infectious disease virus for preparing inactivated vaccine and evaluation of their immunogenicity

FIELD: veterinary virology.

SUBSTANCE: the virus strain of bursal infectious disease (IBB) "52/70-M" is isolated from bursas of Fabricius of sick and killed chickens. The strain shows the expressed precipitating and infectious activity. Invention can be used for producing inactivated vaccine against IBB virus and for evaluation of properties of vaccines against IBB virus.

EFFECT: valuable properties of strain.

4 tbl, 3 ex

 

The invention relates to veterinary Virology and can be used in the production of inactivated vaccine against infectious bursal disease of poultry (IBD), as well as control the virulent strain for evaluation of the immunogenic properties of the vaccine strains of the indicated disease.

The main direction of prevention of IBD is the use of vaccines. This is done using live and inactivated vaccines, the effectiveness of which is determined by the quality, mainly their high immunogenicity and safety.

Vaccinated liquid with a high concentration of virus is the starting material in the production of vaccines, their quality assessment and receipt of diagnostic drugs, as well as inactivated vaccines. Further processing of the viral fluid depends on the desired final product and technological conditions. Vaccinated liquid used as part of diagnostic or vaccine preparation; antigen for immunization of animals when receiving hyperimmune diagnostic sera; the source material for obtaining precipitating antigen of the virus IBD and manufacture of inactivated vaccine.

When assessing the immunogenic vaccine strains are taken into account the results of seroconversion and data from which tasciotti vaccinated birds to infection with a virulent strain of IBD virus. The strain intended for the control of infection to assess the immunogenicity must be stable and free from contamination, quite virulent and possess high biological and antigenic activity.

According to the literature for the production of various immuno-biological properties of the virus IBD use different strains of IBD virus strain "ARTICLE" [1], strain "BG" virus IBD [2] and strain "Winterfield" [3].

However contagious strains of IBD virus, is given in [1] and [3], does not exceed 106,0TCD50/ml, and precipitating activity does not possess.

The closest to the proposed invention, the essential features is the strain "BG" virus IBD birds, intended for the production of immunobiological preparations [2]. Strain "BG" cultivated contamination 10-day-old SPF chicken embryos at 37,7°and humidity 54-66%. Within 72-96 hours of incubation the virus accumulates to 5.25 to 6.0 lg EID50/0,2ml, thus precipitating the title is not in one of the experiments did not exceed 1:16. It should be noted that the precipitating activity of strain "BG" often correlative with the value of the infectious titer. Title of activity of the antigen in the RDP was 1:4-1:16, and the maximum titer of 1:16 was installed, only 3 of the 10 experiments.

Instability obtained Raza is tatov and low titer precipitating activity of antigen hamper the production of immunobiological preparations of virus IBD based on this strain. Strain "BG" due to multiple passages in SPF-embryos lost their infectious properties and for this reason cannot be used as a control virulent virus in experiments to assess immunogenetic vaccine against IBD.

The purpose of the invention is achieved by obtaining a stable virulent strain of IBD virus with high and precipitating infectious activity, suitable for the manufacture of inactivated vaccine and as a reference virulent strain when assessing the immunogenicity of vaccines against IBD.

The strain 52/70-M" deposited in the State collection of viruses all-Russian state research Institute for control, standardisation and certification of veterinary preparations (VGNKI) 03.04.1992, under registration number - virus IBD - "52/70-M-DEP".

Obtaining strain. Material for isolation of the virus IBD is the homogenate of Gabrilovich bags sick or dead chickens. The affected bags homogenizers in 0.001 M phosphate buffer solution (pH of 7.2 to 7.4) in the ratio 1:1 (weight to volume), frozen and thawed three times, centrifuged at 4000 rpm for 30 min, the supernatant sterile sucked off, checked for sterility, biological and antigenic activity (table 1).

Virus isolation performed on SPF-chicken embryos 11-Nam who ate age. Viral material in a volume of 0.2 ml inoculant on chorioallantoic shell (HAO) and incubated at 37-38°for 96-120 hours. View embryos exercise daily and dead embryos within 3-5 days homogenized in phosphate-buffered saline (pH 7,2-7,4), frozen and thawed three times, centrifuged at 4000 rpm for 30 minutes the Supernatant sterile sucked off, checked for sterility, biological and antigenic activity.

Characteristics of the strain. The strain 52/70-M" corresponds to the common structure of the virus IBD and has morphological features characteristic of the pathogen IBD: the shape of the virion icosahedrons, the size of the virion - 58-60 nm. His infectious activity is suppressed by physical and chemical means (pH, heat, acetone, formaldehyde, UV irradiation). The virus does not manifest hemagglutinine activity.

A typical manifestation of strain is active reproduction on 11-day-old SPF-embryos, getting vaccinated liquid infectivity titer of 106,0-106,5EID50/ml and precipitating activity 1:32-1:64. Determination of the titer of infectivity spend titration method for SPF-chicken embryos with a 10-fold dilution of the virus suspension in physiological solution. The final accounting is conducted on 7-8 day by taking into account pathological changes in the organization who tries infected embryos. The strain 52/70-M" IBD virus is neutralized by specific antibodies. The neutralization reaction is conducted in SPF chicken-embryo.

Example 1. As a specific serum using hyperimmune serum to the virus IBD, To obtain hyperimmune serum to IBD virus infect chickens virulent strain 52/70-M" at the age of 35-40 days and after 15 days re-enter the virus in a volume of 0.5 cm in the breast muscle. Serum from chickens get 14 days after re-introduction of strain. Serum tested for activity and specificity in the reaction diffuse precipitation and neutralization (table 2).

Example 2. The use of strain 52/70-M to assess the immunogenicity of the vaccine against IBD.

One of the requirements in the production of vaccine is tested for immunogenicity, i.e. determination of the degree of stability of vaccinated birds to infection with a virulent strain of the virus geologicznego by infecting birds control virulent strain of the pathogen.

Table 3 presents the results of the control of infection of chickens vaccinated with different vaccines production, virulent strain 52/70-M". Virulent virus was administered intranasally to chickens of all groups in a dose of 0.2 ml after 21 days after immunization. After the observation of birds killed and conducted the autopsy for detection of patologoanatom the economic changes, characteristic of IBD virus. The results of the experiments indicate that virulent strain 52/70-M equally suitable for assessing the immunogenicity of both live and inactivated vaccines against IBD.

Example 3. Fabrication and testing of inactivated vaccines.

Inactivated vaccine against IBD virus prepared from virus strain "52/70-M"grown in developing SPF-chicken embryos. Vaccinated material inactivate a 0.1%solution In-propiolactone with subsequent check on the completeness of inactivation. Preparing an oil emulsion based on mineral oil Marcol-52 with the addition of arlacel.

The results of the evaluation of immunogenic activity of inactivated vaccine from strain 52/70-M are presented in table 4.

Sources of information

1. Gazizov PH Immunobiological properties of the vaccine strain "ST" and the effectiveness of specific prevention of bursal disease of poultry. - , Tartu, Estonia, SHA, 1990, p.21.

2. RF patent № 2127602, p.12, 24.11.1997,

3. RF patent № 2127604, p.12, 14.04.1998,

Table 1

Precipitating and biological activity of strain 52/70-M virus IBD
No. of experimentsPrecipitating activity in RDPBiological activity on SPF-embryos (EID 50/ml)
11:326,0
21:16of 5.75
31:16of 5.75
41:16of 5.75
51:646,5
61:646,5
71:32of 5.75
81:325,5
91:325,5
101:646,5

Table 2

Activity of immune sera in RDP and PH
No. of experimentsAntibody titer
RDPPH
11:1261:1024
21:641:512
31:1261:1024
41:2561:2048
51:1261:1024
61:1261:1024
71:641:512
81:641:512
91:641:512
101:64 1:512

Table 3

The results of the control of infection by strain 52/70-M" SPF-chickens vaccinated against IBD
A group of chickens vaccinated with vaccineTotal goalsThe results of the infection, sore/PaloThe percentage of immunogenicity
D-78201/195
KBq200/0100
FYR-706200/0100
Inactivated201/095
Control, unvaccinated2020/50

100
Table 4

Immunogenic activity of inactivated vaccine against IBD from strain 52/70-M"
№/№ experimentsThe dose of the virus on 1 goal. in EID 50/mlVaktsinirovanoThe results of the control of infection
InfectedIllThe percentage of intact
The first experience
1101010280
21010100100
31010100100
4Control1010100
The second experience
1101010190
21010100100
31010100100
4Control1010100
The third experiment
1101010280
21010100100
31010100
4Control1010100

The strain of the virus infectious bursal disease of poultry Bursitis infectiosa gallinal cem. Birnaviridal, pog. Avibirnavirus, VGNKI serotype 52/70-M for the manufacture of inactivated vaccine and evaluation of the immunogenicity of vaccines against infectious bursal disease.



 

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