Treatment of respiratory mycoplasmosis in poultry. Infectious bursitis of chickens (Gumboro disease)
General information about the disease
Infectious bursal disease (IBD, Gumboro disease, chicken infectious bursitis) is a highly contagious viral little-studied disease of chickens 2-15 weeks of age, characterized by damage to the bursa of Fabricius, nephroso-nephritis, intramuscular hemorrhages and diarrhea. The disease, first recorded in Gumboro County (Delaware, USA) in 1957, received wide use in many countries of America, Asia, Africa, including the CIS countries. Since 1991, the disease has been registered in the Republic of Belarus.
Infectious bursal disease is common mainly in industrial poultry farms. The reason for this is considered to be the constant import of poultry. Birds 3-6 weeks of age are most often affected. The earliest outbreaks reported were in 11-day-old chicks and the latest at 84 days of age. Morbidity reaches 100%, mortality averages 20-40%. The source of infection is sick and recovered birds; the main route of spread of the virus is aerogenic. Transmission factors: poultry slaughter products, contaminated feed, water, clothing and footwear of service personnel. There are acute, subacute and chronic course of the disease. When chickens 3-6 weeks of age are infected, the disease, as a rule, is acute and subacute. When the disease occurs in chickens under 3 weeks of age that do not contain maternal antibodies, a latent (subclinical) form of the disease develops.
The economic damage caused by the disease is caused by the death of chickens, a decrease in body weight gain, and an increase in the percentage of rejection of birds and carcasses. The virus has a pronounced immunosuppressive effect, selectively affecting one of the central organs of the immune system of birds - the Bursa of Fabricius. As a result, the effectiveness of vaccinations against infectious laryngotracheitis, Newcastle disease, infectious bronchitis, Marek's disease decreases, and secondary infections become more active.
Most chickens infected with the infectious bursal disease virus show signs of liver damage, and during bacteriological examination of the organ, the causative agent of salmonellosis is isolated. Economic losses in farms unfavorable for IBD increase significantly due to complications: dermatitis with extensive necrotic lesions of feather follicles and skin in the back and wings, hepatitis and necrotic enteritis are often observed, and the incidence of chicken colibacillosis, Marek's disease, and eimeriosis increases. The danger of immunosuppression of this virus lies not only in the reduction of the immune response of the bird's body, but also in the fact that the antibodies produced to various antigens during and after the disease are functionally defective.
Infectious bursal disease virusThe virus was first discovered and described as an independent pathogen in 1962. According to modern classification, it belongs to the family Birnaviridae.
Sustainability.
The virus is resistant to ether, chloroform and UV irradiation. At a temperature of +56°C it lasts for 5 hours, at +60°C for 30 minutes, and at 30°C in the presence of 0.5% phenol for 1 hour. When exposed to a 0.5% formalin solution, it is inactivated in 6 hours, iodine preparations - in 2 minutes, 0.5% chloramine - in 10 minutes.
Antigenic variability and relatedness.
Two serotypes of the virus have been identified: Cu-1 (contains proteins VP-1, VP-2, VP-3, VP-4 and VPX), isolated from chickens, and 23/82 (contains proteins VP-1, VPX, VP-3 and VP- 4), isolated from turkey poults. The antigenic relatedness of these serotypes does not exceed 10-30%.
Spectrum of pathogenicity.
Under natural conditions, the IBD virus affects chickens of all breeds, but White Leghorn chickens are most seriously ill. Birds are most sensitive at 3-6 weeks of age. When laying hens, as well as 1-10-day-old chickens, are infected, no symptoms of the disease are observed.
Localization of the virus.
On the 3rd day after experimental infection of 3-5 week old chickens, the virus accumulates in the bursa of Fabricius, spleen and, in lower concentrations, in the brain and blood. In one-day-old chicks, 3 days after inoculation of the virus, its high concentration in the bursa of Fabricius, liver and kidneys. During experimental infection of chickens, the virus can be isolated within 10 days, but pathomorphological changes in the bursa of Fabricius persist for 10 weeks after infection.
Antigenic activity.
Already on the 21st day after infection of chickens 3-5 weeks old with the virus, virus-neutralizing antibodies (in titers up to 1:718) and precipitating antibodies (in titers up to 1:640) are detected in the blood serum. Antibodies in laying hens are transmitted transovarially to their offspring. In the blood serum of chickens obtained from immune chickens, specific antiviral antibodies are detected within 3-4 weeks after hatching.
Experimental infection.
Experimental infection is carried out on 20-30-day-old chickens by inoculating the viral material onto the conjunctiva, as well as orally. Infection is accompanied by the development in birds of clinical signs and pathological changes characteristic of this disease.
During intraperitoneal and intracerebral infection of 1-3-day-old white mice with the Becht strain, itching, ataxia, coma were noted, and in the brain - non-purulent lymphocytic encephalitis. Intracerebral infection of rats and hamsters is possible.
Infection with the turkey virus is accompanied by the formation of virus-neutralizing and precipitating antibodies in the absence of clinical signs of the disease. Pigeons, geese, quail and ducks are not susceptible to this infection.
Cultivation.
The virus is cultivated in chicken embryos (CE), free from maternal antibodies, when they are infected into the allantoic cavity or onto the chorion-allantoic membrane. The death of embryos occurs on the 3-8th day after infection.
The virus reproduces well in chicken embryonic kidney cell culture, causing CPD on days 3-5 after infection. In cultured FE fibroblasts, the virus forms plaques. The possibility of cultivating the virus in a continuous cell culture MA-104, Vero has been demonstrated.
Hemagglutinating properties.
Under normal conditions, without prior special treatment, the virus does not have hemagglutinating properties.
Pathogenesis
The pathogenesis of infectious bursal disease has not been fully studied. It has been established that the target cells for virus reproduction are lymphocytes of the bursa of Fabricius of chickens. B-lymphocytes carrying class M immunoglobulins on their surface turned out to be highly sensitive to the virus. The virus has a pronounced cytopathic effect, causing necrosis of lymphoid nodules and inflammatory processes in the interstitial tissue of the bursa of Fabricius. The death of a large number of lymphoid elements causes the development of secondary immunodeficiency in recovered birds.
It has also been shown that the latent course of IBD is accompanied by the phenomena of atrophy and delymphatization of the bursa of Fabricius against the background of the absence or very weak manifestation of an inflammatory micro- and macrophage reaction in the interstitium of the organ. This form of the disease is also characterized by the development of an immunosuppressive state in chickens associated with the phenomena of B-lymphocyte necrosis.
Morphologically, lymphocyte necrosis is revealed by the phenomena of karyopyknosis, karyorrhexis, vacuolization of the cytoplasm with the formation of apoptotic bodies. Apoptosis, unlike necrosis, does not cause a pronounced inflammatory response. The phenomena of B-lymphocyte necrosis in chickens with the subclinical form of Gumboro disease are found not only in the bursa of Fabricius, but also in the spleen, cecal tonsils and peripheral blood.
The pathogenesis of the disease also depends on the influence of immune complexes circulating infected lymphocytes. Hemorrhages in skeletal muscles, liver and other organs are caused by wall damage blood vessels. The presence of urates in the kidneys and an increase in uric acid levels in the blood indicate kidney damage. Increased activity of lactate dehydrogenase and glutamate oxalate transaminase in the blood serum confirms liver damage.
Clinical signs
Incubation period infectious bursal disease short. When chickens are experimentally infected, clinical signs appear within 2-3 days. The disease can occur acutely, subacutely and latently, depending on the immune status of the livestock. In susceptible groups, as a rule, the incidence rate can reach 100%. In acute cases, the disease usually lasts 4-8 days.
The characteristic symptom of IBD is diarrhea, accompanied by the discharge of watery, whitish-yellow droppings. In sick chickens, depression is observed, and in a later stage - trembling of the head and neck, coma. Morbidity and mortality increase rapidly and reach a maximum on the 3-4th day of illness. The distinctive signs of the disease are suddenness and high incidence, rapid recovery of the livestock. Mortality can reach 20-40%. In subsequent chick hatchings, occasional outbreaks are less severe and often go unnoticed.
In recent years, the number of outbreaks of latent Gumboro disease has increased significantly. In this case, infection of chickens leads to the development of an immunodeficiency state in birds due to necrosis of B-lymphocytes. As a result, the effectiveness of vaccinations against a number of viral diseases decreases. There have been outbreaks of chronic respiratory diseases leading to the development of swollen head syndrome. Saprophytic microflora are often isolated from the affected tissues: cocci, pseudomonads, etc.
Pathomorphological changes
The corpses of dead chickens are usually well fed. At autopsy, signs of dehydration and anemia are noted. The goiter is empty. Changes in the bursa of Fabricius are pronounced and very natural, lesions of which are also found in cases of asymptomatic infection. The organ is enlarged 1.5-2.5 times. The serous membrane is gray-yellow. The mucous membrane is swollen, reddened, with hemorrhages. In the lumen of the bursa between the folds of the mucous membrane, serous-fibrinous exudate is found, in severe cases– hemorrhagic exudate and cheese-like fibrin clots. Atrophy of the thymus, aplasia of the red bone marrow, esophageal and cecal tonsils, and serous-hemorrhagic inflammation of the spleen are noted.
Dotted and spotty hemorrhages are found in the pectoral muscles, on the medial side of the thighs and wings. The liver may be slightly enlarged, with marks from the ribs visible on the surface. The kidneys are enlarged, light gray to dark brown in color, with a clear pattern of tubules and ureters filled with urate. In addition, catarrhal enteritis and hemorrhages in the mucous membrane of the glandular stomach and in the cecal tonsils are noted.
Histological examination of the Bursa of Fabricius in the initial period notes necrosis of lymphocytes, and then of the reticular stroma with the formation of necrotic detritus in the majority of lymphoid nodules. Atrophied nodules, glandular structures and cysts may be detected. In the thymus at the beginning of the disease, delymphotization of the cortical layer, depletion of lymphocytes in the medullary zone with a simultaneous increase in the number and size of Hassal's bodies and hyperplasia of reticular cells are observed. In the red bone marrow, a decrease in the total number of cellular elements and an activation of the macrophage reaction are detected. In the spleen of sick chickens, necrosis of individual lymphocytes in the periarterial couplings (T-lymphocytes) and lymphoid nodules (B-lymphocytes), hyperemia of red pulp vessels, micro- and macrophage reactions are detected. Similar changes are found in the esophageal and cecal tonsils.
1. Serous-hemorrhagic or fibrinous-necrotic inflammation of the bursa of Fabricius.
2. Serous- hemorrhagic inflammation spleen.
3. Atrophy of the thymus, bone marrow, esophageal and cecal tonsils.
4. Dotted and spotty hemorrhages in the muscles of the thighs and wings (on the medial side), in the serous integument.
5. Granular dystrophy of the liver and kidneys, overflow of the ureters with urates.
6. Serous-fibrinous pericarditis, aerosacculitis, pleuroperitonitis, perihepatitis (complication).
7. Histo: total necrosis of lymphocytes in the bursa of Fabricius, thymus and spleen, serous-inflammatory edema of interstitial tissue, infiltration with histiocytes and pseudoeosinophils in acute and subacute cases; atrophy and depletion of lymphocytes in the bursa of Fabricius, thymus, spleen, esophageal and cecal tonsils, absence of micro- and macrophage reaction in latent course diseases.
Diagnostics
Diagnosis of infectious bursal disease is carried out taking into account epidemiological data, clinical symptoms, pathomorphological changes, as well as a complex of laboratory tests. It should be borne in mind that this disease is difficult to detect, is masked by other infections, and only in a typical course is it diagnosed based on clinical signs and autopsy data. Therefore on early stage The disease and its latent course require laboratory tests.
Laboratory diagnostics of IBD includes the isolation of the virus in developing SPF chicken embryos (CE) or in the culture of chicken embryo fibroblasts (FEC), its identification in the neutralization reaction (RN) and the immunodiffusion reaction (IDR), performing a bioassay on sensitive chickens, identifying the viral antigen by reaction immunofluorescence (RIF), indirect hemagglutination inhibition reaction (IHRA), latex agglutination reaction (RAL), electron microscopy, counter immunoelectrophoresis reaction, as well as detection of specific antibodies in PH, RID, VIEF, indirect hemagglutination reaction (IRHA), enzyme-linked immunosorbent assay (ELISA) ) and conducting histological studies.
Selection of pathological material.
Bursa Fabricius, spleen, liver, and kidneys are taken from 5-10 corpses of dead or sick chickens killed for diagnostic purposes. Organs are placed in clean, dry, sterile penicillin vials. The material is placed in a thermos with ice and stored until sent to the laboratory.
For serological testing, paired samples (25-30) of blood serum from sick birds are sent, taken at an interval of 21 days. The resulting serums are placed in clean, dry, sterile penicillin vials under rubber stoppers and placed in a thermos with ice.
Material sent to the laboratory is supplied with a covering letter. In the laboratory, the material is stored frozen or filled with a 50% aqueous solution of glycerol.
In the laboratory, pieces of pathological material are homogenized in 0.01 M phosphate-buffered saline (pH = 7.2) or meat-peptone broth in a ratio of 1:10, frozen and thawed three times, followed by centrifugation for 30 minutes at 5000 rpm. 100 U/ml penicillin and 0.1 mg/ml streptomycin are added to the supernatant liquid, incubated for 12 hours at 4°C, and checked for sterility.
Virus isolation in chicken embryos.
To isolate and titrate the pathogen, use SPF embryos of 9 days of age in an amount of at least 10. The prepared homogenate in an amount of 0.2 ml is applied to the chorion-allantoic membrane or into the allantoic cavity of the embryo. 5-10 uninfected embryos are left in the control.
The death of chicken embryos during the first day is considered nonspecific. If there is a virus in the test material, death of TBE occurs within 3-5 days. Dead embryos are dissected to detect specific pathological changes. The chorion-allantoic membrane (CHM) is edematous. There is a delay in growth and development, the presence serous-hemorrhagic swelling of the skin in the head, neck, limbs and abdominal wall. In the lungs, congestive hyperemia and pulmonary edema, necrotic nephrosis, granular myocardial degeneration, enlarged liver and spleen are detected. The bursa of Fabricius is usually not affected.
From dead embryos with growth retardation and pronounced pathomorphological changes, allantoic fluid and CAO are collected into sterile tubes and checked for sterility by inoculating 0.2 ml of the fluid with MPA and MPB. Whether the isolated agent belongs to the Gumboro disease virus is determined by setting the RN and RID.
Infection of cell cultures.
To isolate the virus, 24-48-hour cultures of EC fibroblasts are used. The timing of the onset of CPE depends on the dose of the virus and the number of passages. During its initial isolation, specific changes are observed after 2-3 passages of virus-containing material. CPD manifests itself 48-72 hours after infection of the cell culture and is characterized by vacuolization and rounding of cells, the phenomena of karyopyknosis and karyolysis. The specificity of cytopathic changes is confirmed by staging RN.
Bioassay on chickens.
For infection, 21-day-old SPF chickens or 35-40-day-old birds from a commercial flock with well-developed Fabrician bursae are used. To do this, from the total number of chickens intended for bioassay, 5-10 heads are randomly caught, killed, individual absolute body weights and bursae are determined, and the bursal index (BI) is derived using the formula:
where Ms is the mass of the bursa of Fabricius (g),
BW – body weight of birds
Chickens in the flock of which the bursa index is 4 or higher are used to perform a bioassay. Infection of chickens with an index below 4 does not cause an acute course of the disease. Determination of the bursal index has important diagnostic value, since in infected chickens there is a decrease in this indicator by 3-9 times.
Before infection, blood serum samples are taken from 10-20 birds selected for bioassay for serological testing (RID, RN, RNGA, ELISA) for the presence of antibodies to the Gumboro disease virus. The test material is administered intranasally in a dose of 0.5 ml.
The bioassay is considered positive if the infected chickens develop characteristic clinical signs of the disease after 2-5 days (diarrhea, dehydration, general depression, dirty gray feathers). At autopsy of sick and recovered birds, characteristic pathological changes are noted.
15 days after infection, a repeat serological test of the blood serum is performed in the surviving chickens to identify a diagnostic (4-fold) increase in titer specific antibodies and then they are killed in order to establish pathological changes.
Neutralization reaction.
It is used to identify the Gumboro disease virus and to detect specific antiviral antibodies. The reaction is carried out on chicken embryos. It uses normal and specific hyperimmune serum, and the virus isolated on BE is used as an antigen. Before the reaction, the serum is inactivated in a water bath at +56°C (30 min), after which penicillin (1000 U/ml) and streptomycin (1 mg/ml) are added.
Hyperimmune and normal serum are poured into sterile test tubes using dry sterile pipettes of 0.5 ml. Then each of them contains 0.5 ml of the test antigen in dilutions from 10 -1 to 10 -9. After shaking, the tubes are kept at +37°C for 30 minutes. The resulting mixtures in a dose of 0.2 ml are injected into the allantoic cavity of 9-day-old embryos. Ovoscopy is performed twice a day. Dead embryos are dissected. On the 10th day, all embryos are dissected to identify specific changes in organs and tissues. The pH results are expressed by the neutralization index, which is determined according to the generally accepted method. The reaction is considered positive if the difference in titers with normal and hyperimmune sera is 2 lg or more.
Immunofluorescence reaction.
It is a rapid diagnostic method, as it allows a diagnosis to be made within 2-3 hours from the moment of delivery of the pathological material.
Imprint smears (at least 3) are prepared from the bursa of Fabricius of dead birds or killed for diagnostic purposes on thin, dry, fat-free glass slides, fixed for 10-20 minutes in acetone, washed twice in two shifts with 0.01 M phosphate-buffered saline ( pH=7.2-7.4), then dried and painted according to the generally accepted method. Control fingerprint smears are prepared from the bursa of healthy chickens.
The reaction is considered positive if at least 3 lymphocytes with a specific bright green glow of the antigen in the cytoplasm (small granules or a diffuse halo around the nucleus) are detected on all preparations.
In addition to fingerprint smears for RIF, cryosections of the bursa of Fabricius can be examined. To do this, the organ is frozen in petroleum ether, cooled to minus 76°C in a mixture of acetone and dry ice. They are then frozen to the microtome block. Sections are prepared with a thickness of 4-5 microns.
Inhibition reaction of indirect hemagglutination.
This technique can be used to indicate the viral antigen in pathological material, as well as to identify the virus isolated in EC and CC.
From pathological material from sick and dead birds, chorion-allantoic membranes of dead birds after TBE infection, a homogenate is prepared in a sterile 0.85% sodium chloride solution (pH = 7.2-7.4) in a 1:1 ratio. After freezing and thawing three times, the homogenate is centrifuged at 5000 rpm. within 25 minutes. The supernatant liquid is drained for testing in the RTNGA. Allantoic fluid samples are used in their native form after centrifugation for 15-20 minutes at 3000 rpm. Suspensions from control birds and EC are prepared in the same way. In addition, specific immune serum to the Gumboro disease virus and a positive erythrocyte antigen from the BelNIIEV diagnostic kit are used for serological diagnosis of this disease in the RNGA.
RTNHA is performed using the micromethod using a Takachi microtiter. In this case, a twofold dilution of the test material from 1:2 to 1:128 is prepared in the first row of wells of the panel in a volume of 0.025 ml. To do this, add 0.025 ml of 0.85% sodium chloride solution containing 0.5% glycerol to each well. Add 0.025 ml of the test material to the first well and, while mixing, transfer 0.025 ml to another well, etc. (up to a dilution of 1:128. In the second row, a virus-free suspension is prepared in the same way. To each dilution of both rows, add 0.025 ml of immune serum, diluted 1-2 times less than its limiting titer in the RNGA. The panel with the mixtures is shaken and placed in a thermostat for 60 minutes at 37 ° C. The reaction is taken into account after the sedimentation of erythrocytes.
The reaction is considered positive if, in the first row with the test material, agglutination is inhibited in the first two or three wells, provided that the second row is completely agglutinated.
The latex agglutination test is used to indicate viral antigen. Pathological material is homogenized in 0.01 M phosphate-buffered saline (pH = 7.2) in a 1:1 ratio, frozen and thawed three times. The virus-containing suspension is centrifuged at 3000 rpm. 30 minutes. The supernatant liquid is drained to perform RAL.
Hyperimmune serum to the IBD virus is obtained by immunizing chickens with an inactivated oil-emulsion vaccine. The gamma globulin fraction is isolated from the serum by precipitation with ammonium sulfate.
Latex antibody diagnosticum is prepared by mixing a latex suspension (2% particle concentration) with an equal volume of gamma globulin fraction in an appropriate buffer solution. The mixture is incubated for 16-18 hours at 4°C, then centrifuged for 30 minutes at 3000 rpm. and wash the precipitate three times with a buffer solution. The sensitized latex is resuspended in the same solution to a final particle concentration (0.5-2.0%) and 0.05% sodium azide is added. The finished diagnosticum is stored in the refrigerator at 4°C.
The latex agglutination reaction is performed on a glass slide. Apply 25 µl of antigen diluted from 1:2 to 1:512 with a dispenser, add 25 µl of latex antibody diagnosticum and mix, turning smoothly. The reaction results are taken into account after 2-5 minutes using a three-point system: sharply positive reaction(+++) – clear agglutination, large flakes in a clear liquid; positive (++) – agglutination is visible, but the background is not completely cleared; negative (-) – cloudy homogeneous liquid. The control is a mixture of latex with a 0.85% sodium chloride solution (diagnosticum control), with positive and negative antigens (positive and negative control).
Immunodiffusion reaction (IDR).
This technique is widely used both for the purpose of indicating and identifying the Gumboro disease virus and for determining specific antibodies. When diagnosing RID, kits for diagnosing Gumboro disease VNIVIP or ARRIAH are used.
In the laboratory, the pathological material is weighed, an equivalent amount of 0.85% sodium chloride solution or distilled water is added, homogenized, frozen and centrifuged at 3000 rpm. within 10 minutes. The supernatant is drained and used for RID testing.
To set up the reaction, use 1.25% agar with 8% sodium chloride and 0.5% phenol. Petri dishes are filled 24-72 hours before use, pre-dissolving the agar by steaming and pouring 20 ml into the dishes. The agar layer should be at least 3 mm.
To study samples, a linear order of wells is used. Make 3 rows of vertical holes with a diameter of 5 mm, at a distance of 5 mm. Agar plugs are removed with a needle or tweezers.
Each sample of the tested diagnostic serum is diluted with physiological solution (0.85% sodium chloride solution), first 1:2, and then sequentially in twofold steps up to 1:256.
Before use, dry reference antigens are dissolved with distilled water or 0.85% sodium chloride solution to a volume of 0.5 ml. Dry diagnostic sera are first diluted with distilled water in the volume indicated on the label, and then a series of successive two-fold dilutions are prepared.
When examining pathological materials, positive serum is added to the central row of wells in a working dilution of 0.05 ml, and normal, positive (1 well each) and test antigens in a volume of 0.05 ml are added to the peripheral rows. When studying bird blood sera, a positive antigen in a working dilution in a volume of 0.05 ml is added to the central row of wells, and normal, positive (1 well each) and test sera in dilutions of 0.05 ml are added to the peripheral rows.
After filling the wells, the Petri dishes are placed in a thermostat at a temperature of 37°C. The reaction is recorded 24 and 48 hours after the reaction is performed. The cups are viewed against a dark background in a directed beam of light. The reaction is taken into account only if there are precipitation lines between the positive antigen and the positive serum in the control and the absence of precipitation lines between the positive antigen and normal chicken serum, as well as positive serum and normal antigen.
The formation of 1-2 precipitation lines between the wells with the test material and positive serum is taken as a positive result when examining pathological materials for the detection of viral antigen, and when antiviral antibodies are detected, the presence of precipitation lines between the wells with the test serum and the positive antigen.
Counter immunoelectrophoresis is used both for the purpose of indicating and identifying the Gumboro disease virus and for determining specific antibodies. The essence of the technique is the simultaneous movement of protein molecules with different electrophoretic mobility in agar with the formation of a precipitate from homologous antigens and antibodies. To set up VIEF, use devices PEF-3, EF-2 or similar brands and a set of diagnostic kits for setting up RID. The reaction is carried out on glass plates coated with a 1% agar solution in 0.85 M veronal-medinal buffer (pH = 8.6). Wells with a diameter of 4 mm are cut out in the agar, at a distance of 4 mm from each other.
A positive antigen and test sera are added to the wells at the anode, and a positive antigen and the pathological material to be tested are added to the wells at the cathode. Electrophoresis is carried out for 1.5 hours at a current of 4 mA/cm. Negative antigen and serum serve as controls
The plates are viewed in oblique light against a dark background. The reaction is considered positive if one or two lines of precipitation form between the wells containing the test antigen or serum and the positive serum or antigen.
The indirect hemagglutination reaction is based on the ability of antibodies to agglutinate red blood cells sensitized with a specific antigen. RNGA can be used both for serological diagnosis and for studying the serological epizootology of Gumboro disease.
When staging RNGA, erythrocyte antigen and control (positive and negative) sera from the BelNIIEV diagnostic kit are used to diagnose infectious bursitis (Gumboro disease).
The reaction is carried out using the micromethod in a Takachi microtitrator. In the wells of horizontal rows of plexiglass plates, successive twofold dilutions (from 1:2 to 1:1024) are prepared from the test plates in a 0.85% sodium chloride solution containing 1% glycerol, in a volume of 0.025 ml (1 drop). Add 0.025 ml of test serum to the first wells, mix, and transfer 0.025 ml of the mixture to the next wells, etc. From the last wells after mixing, 0.025 ml of contents are removed into disinfectant solution. Add 0.025 ml of a 1% suspension of virus-sensitized erythrocytes to each well with an appropriate serum dilution and shake.
At the same time prepare: control of erythrocyte antigen for spontaneous agglutination (0.025 ml of negative serum and 0.025 ml of erythrocyte antigen are added to 2-3 wells); positive control (0.025 ml of positive serum and 0.025 ml of erythrocyte antigen are added to 2-3 wells); negative control (0.025 ml of negative serum and 0.025 ml of erythrocyte antigen are added to 2-3 wells).
Plexiglas plates with components are placed in a thermostat for 1-1.5 hours at t=37°C until the red blood cells settle.
The reaction is recorded only in the case when the control of the erythrocyte antigen for spontaneous agglutination and with obviously negative serum is negative, and the control with positive serum is positive.
A positive reaction is characterized by the appearance of a sediment of red blood cells in the form of an umbrella at the bottom of the well. A negative reaction is manifested by the precipitation of red blood cells in the form of a point or ring with smooth edges.
When agglutination of erythrocytes with test sera in a dilution of 1:8 and above, RNGA is considered positive, and 1:4 and below is considered negative.
Enzyme immunoassay is widely used as the most specific test for detecting specific antiviral antibodies in the blood serum of immune birds. To perform this reaction, a set of ARRIAH diagnostic kits is used to determine antibodies to the infectious bursal disease virus (Gumboro disease) using the ELISA method. The essence of this technique is to detect the antigen-antibody complex on the surface of the wells of a polystyrene plate. The resulting specific complex interacts with the anti-species immunoperoxidase conjugate against chicken Ig G and causes decomposition of the substrate, staining the contents of the wells of the plate.
Before preparing working solutions, the kit and components are kept for 30 minutes. at room temperature (18-20°C).
Solution No. 1. Dissolve 0.97 g of tris (hydroxymethyl) aminomethane (bottle 5.1), 6.61 g of tris (hydroxymethyl) aminomethane hydrochloride (bottle 5.2) and 11.7 g of sodium chloride (bottle 5.3) in 1000 cm 3 of distilled water. After measuring the pH of the resulting solution (which should be between 7.4-7.6), 1.0 ml of Tween-20 liquid detergent (bottle 7) is added to it. This solution is used for diluting control sera, test samples, anti-species conjugate and inter-stage washing.
Solution No. 2. To prepare the substrate buffer, 5.37 g of sodium dihydrogen phosphate (bottle 6.1) is dissolved in 50 ml of distilled water. The contents of bottle 6.2 (1.51 g of citric acid) are also dissolved in 50 ml of distilled water. Then mix 224.3 ml of sodium dihydrogen phosphate solution with 25.7 ml of citric acid solution, add 50 ml of distilled water. The resulting solution should have pH=4.9-5.0. If necessary, add acidic or alkaline components.
Solution No. 3. 1.0 cm 3 of whole lyophilized positive serum against Gumboro disease (bottle 1) is dissolved in 0.5 ml of solution No. 1. The resulting solution can be stored for 3 days at 4 °C.
Solution No. 4. 1.0 cm 3 of whole lyophilized negative serum against Gumboro disease (bottle 1) is dissolved in 0.5 ml of solution No. 1. The resulting solution can be stored for 3 days at 4 °C.
Solution No. 5 The contents of solution 4 with the anti-species conjugate are dissolved in 0.5 ml of solution No. 1. To obtain a working dilution of 1:200, 0.05 cm 3 per 10.0 ml of solution No. 1 (per 1 tablet) is taken from this bottle. Prepare before use. Cannot be stored.
Solution No. 6. One tablet of hydroperite is dissolved in 20 ml of distilled water. Store in a place protected from light at 4°C for no more than 20 days.
Solution No. 7. Substrate-indicator mixture. A tablet of orthophenylenediamine (substrate) is dissolved in 20 ml of solution No. 2, shaken until completely dissolved and 0.4 ml of solution No. 6 is added for every 20 ml of this solution. Cannot be stored.
A plate is taken from the kit, in the wells of which the purified antigen of the Gumboro disease virus is adsorbed. Samples of the blood serum samples under study are diluted 1:100 with solution No. 1. For this purpose, add 1 ml of solution No. 1 to 0.01 ml of serum.
0.1 ml of solution No. 1 is added to the wells of rows B1-12...H1-12, and 0.2 ml of diluted serum samples is added to wells A2-11 and titration is carried out in vertical rows, 1:100 to 1:12800 . Dilutions of 1:100 control (negative and positive) sera are added to wells A1 and A12 and titration is also carried out in vertical rows. From the last wells H1 and H12, 0.1 ml is removed.
The tablet is carefully shaken, covered with a lid and transferred to a thermostat for 2 hours at a temperature of 37°C. Then the wells of the plate are freed from the contents by shaking and washed three times with solution No. 1. Add 0.1 ml of solution No. 5 to all wells of the plate, place in a thermostat for 1 hour, and wash three times with solution No. 1. Then 0.1 ml of solution No. 1 is added to all wells. Leave at room temperature for 10 minutes. The reaction is stopped by adding 0.05 ml of a 0.5% sulfuric acid solution to each well.
Detection of specific antibodies in blood sera can be carried out without titration of the test sera. In this case, 1 ml of blood serum at a dilution of 1:400 is added to all wells of the A2-12...H2-12 plate. Positive control sera are added to wells A1 and B1 of the vertical row, and negative control sera at a dilution of 1:400 are added to the next two C1 and D1, and wells E1 and F1 serve as a conjugate control - 1 ml of solution No. 1 is added to them.
The results of the analysis are recorded after stopping the reaction in one of the following ways: visually - by the intensity of coloring of the contents or instrumentally - using a spectrophotometer with a vertical beam at a wavelength of 492 nm. During visual recording, the color of the contents of the wells of the test sample plate is compared with the color of the wells of the control samples. The titer of the test serum is taken to be its last dilution, at which a color visible to the eye is observed, more intense than that of the negative sample. Samples with a dilution of 1:400 and higher are considered positive. When assessing the results without serum titration, the reaction is assessed according to the principle - “yes” - a positive reaction (specific antibodies are present in the sample) or “no” - a negative reaction (there are no specific antibodies in the sample).
Instrumental photometric recording makes it possible to quantify titers of specific antibodies by determining extinction. The final dilution of the test serum is considered to be its last dilution, in which the extinction exceeds the control by 2.0-2.1 times.
Histological examination.
From the corpses of dead or forcedly killed birds, pieces of the bursa of Fabricius, thymus, spleen, liver, kidneys, heart, and skeletal muscles are taken. Organs with labels are placed in a glass container and filled with a 10% formaldehyde solution for fixation. The volume of fixing liquid must exceed the volume of the pieces being fixed by at least 10 times. Fixation is carried out at room temperature (18-20°C) for 24-48 hours. The criteria for completing fixation are: uniform compaction of organs and the same color on the surface and on the section. Fixed pieces with labels are placed in a container with a fixative liquid or in plastic bags with cotton wool soaked in a fixative and sent to the laboratory with a covering letter.
In the laboratory, the material is compacted by freezing with liquid nitrogen or on semiconductor stages, or by embedding in paraffin. Histological sections are obtained on freezing or sleigh microtomes and stained with hematoxylin-eosin. Stained sections are examined under a light microscope.
In the bursa of Fabricius, at the onset of the disease, massive necrosis of lymphocytes and then reticular cells is noted with the formation of necrotic detritus in place of the majority of lymphoid nodules. Lymphoid nodules are replaced by glandular structures. At the same time, a local immune reaction is detected, in which, along with necrosis of lymphoid nodules, extensive lymphocytic proliferations are formed with the formation of small lymphoid nodules in them. Edema, infiltration of pseudoeosinophils and histiocytes, and hyperplasia of reticular cells are observed in the interstitium.
It should be taken into account that during the latent course of the disease, atrophy and delymphatization of lymphoid nodules develop due to necrosis of lymphocytes.
In the thymus during the acute course of Gumboro disease, necrosis of lymphocytes is found in the cortical, less often in the medulla, and an increase in the number and size of Hassall's bodies. Inflammatory hyperemia of blood vessels, micro- and macrophage reactions, and hyperplasia of reticular cells are noted. Subacute and latent course is accompanied by early accidental involution of the organ.
In the spleen at the onset of the disease, hyperemia of red pulp vessels, macrophage infiltration, and necrosis of lymphocytes in periarterial couplings (T lymphocytes) and lymphoid nodules (B lymphocytes) are detected. At a later date, pronounced plasmatization of the organ is revealed.
In the kidneys during the acute course of Gumboro disease, vascular hyperemia, vacuolar degeneration and necrosis of epithelial cells, and destruction of convoluted tubules are recorded. In some areas, extensive lymphocytic-histiocytic proliferations are detected. The chronic course is characterized by the phenomena of nephrosclerosis, the accumulation of urate crystals in the lumens of the tubules.
In the liver, hyperemia of the central veins of the lobules, focal hemorrhages, granular and fatty degeneration of hepatocytes, as well as accumulations of lymphocytes, macrophages (less often microphages) in the interstitium of the organ are noted.
In skeletal and cardiac muscles, hyperemia of blood vessels and sometimes granular degeneration are detected. In a chronic course, weakly expressed lymphocytic-histiocytic proliferations are found between muscle fibers.
Differential diagnosis
At differential diagnosis gumboro disease highest value has an exclusion of tumor diseases of birds, adenovirus infection, reduced egg production syndrome (SSY-76), infectious bronchitis, influenza and Newcastle disease, as well as streptococcosis, pasteurellosis, colibacillosis, eimeriosis, prostagonymosis, hypovitaminosis A, nutritional dystrophy and radiotoxicosis.
Marek's disease affects chickens from 4 to 30 weeks of age, occurring in the form of enzootics, less often epizootics. Chickens get sick more often, and cockerels less often. During life, sick birds exhibit impaired coordination of movement, paresis of legs and wings. Gray eyes, hyperplasia of feather follicles, weakness, and progressive exhaustion are noted. In dead chickens, diffuse or focal sebaceous neoplasms are found in the bursa of Fabricius, spleen, liver, intestinal wall, and thickening of the sciatic nerves is noted. During histological examination, atrophied lymphoid nodules are found in the Bursa of Fabricius, replaced by cysts or glands, and a growth of interfollicular connective tissue. The development of tumors in internal organs, characterized by the proliferation of lymphoblasts, histiocytes and plasma cells, is often noted.
Pathological diagnosis:
1. Growth of sebaceous tumor tissue in the spleen, liver, kidneys, ovary, testes, heart, lungs, wall of the glandular stomach and intestines, thymus, bursa of Fabricius or their atrophy.
2. Hyperplasia of feather follicles.
3. Gray eyes, pupil deformation.
4. Neuritis with sharp thickening in sciatic nerves and in the nerves of the brachial and lumbar plexuses.
5. Histo: proliferation of lymphoblasts, histiocytes, plasma cells and reticulocytes in tumor nodes; atrophy of the lymphoid nodules of the bursa of Fabricius, their replacement with cysts, glands, growth of internodular connective tissue.
Additional laboratory tests are carried out: a bioassay is performed on chickens, blood serum is examined in RID, and histopathological examination of affected organs and tissues is carried out.
Chicken leukosis usually occurs in the form of enzootics. Birds older than 8-12 months of age become ill. Sick chickens experience lethargy, diarrhea, and exhaustion. When autopsying dead birds, diffuse or focal fat-like growths of tumor tissue are found in the bursa of Fabricius, spleen, liver, kidneys, heart and other organs.
Histological examination reveals proliferation of tumor cells in the bursa, forming a tumor focus, the infiltrative growth of which leads to destruction of the organ. In the liver, kidneys, heart, the growth of immature lymphoid cells.
Pathological diagnosis:
1. Growth (diffuse or in the form of nodes) of sebaceous tumor tissue in the bursa of Fabricius.
2. Tumor-like sebaceous nodes in the spleen, liver, kidneys, wall of the glandular stomach and small intestine, heart, lungs
3. Exhaustion and general anemia.
4. Histo: proliferation of immature lymphoid cells in tumor nodes.
Laboratory diagnosis is based on histological examination of the pathological material, staging of RSK and RNGA.
Rous sarcoma is accompanied by progressive cachexia, decreased egg production, diarrhea, drooping abdomen, anemic skin and visible mucous membranes. At autopsy, numerous tumor nodes are revealed in the skin, skeletal muscles, as well as in the spleen, intestinal mesentery, liver, kidneys, and ovary. In the presence of a large number of tumor metastases in the internal organs, atrophy of the lymphoid nodules of the bursa of Fabricius and delymphatization are observed. There is a 2-3 times thickening of the internodular connective tissue septa, their hyperemia and swelling.
Pathological diagnosis:
1. Tumor nodes in the skin, skeletal muscles, spleen, liver, kidneys, intestinal mesentery, ovary.
2. Exhaustion, general anemia.
3. Histo: proliferation of poorly differentiated polymorphic cells in tumor nodes, atrophy and necrosis of elements of the parenchyma of internal organs; atrophy and delymphotization of lymphoid nodules, hyperemia and serous edema of the interstitium in the bursa of Fabricius.
To confirm the diagnosis, a virological study is performed.
Adenoviral infection mainly affects young animals 2-3 months of age. The pathogen is highly contagious. The disease is accompanied by drowsiness and anemia of the mucous membranes. In dead or forcedly killed chickens, hemorrhages in the muscles, enlargement of the liver with bile overflowing the gallbladder are found. The kidneys are swollen, with hemorrhages.
Histological examination reveals numerous micronecrosis, hemorrhages (under the capsule), interstitial lymphocytic-histiocytic proliferations in the liver, and eosinophilic and basophilic inclusions in the nuclei of hepatocytes. In the Bursa of Fabricius, atrophy of lymphoid nodules, depletion of lymphocytes in the medulla, interstitial edema, and thinning of the folds of the mucous membrane are detected.
Pathological diagnosis:
1. Alterative hepatitis, bile overflow of the gallbladder.
2. Acute catarrhal-hemorrhagic enteritis.
3. Enlarged spleen.
4. Hemorrhages in skeletal muscles, liver, kidneys, and in the wall of the gallbladder.
5. General anemia, exhaustion.
6. Histo: liver - intranuclear basophilic and eosinophilic inclusions, vacuolar degeneration and necrosis of hepatocytes, hemorrhages, interstitial lymphocytic-histiocytic proliferations; pancreas – pancreatitis with the presence of intranuclear polychromatophilic inclusions in the cells of the glandular tissue; bursa of Fabricius - atrophy of lymphoid nodules, depletion of lymphocytes in the medulla, interstitial edema, thinning of the folds of the mucous membrane; spleen – serous splenitis with intranuclear inclusion bodies in reticulocytes.
Additional laboratory tests are carried out: virus isolation on chicken embryos.
Reduced egg production syndrome (RES-76) affects chickens of all breeds during the laying period. The virus is capable of persisting in the body of birds and becoming more active under stress caused by the onset of oviposition. The disease is accompanied by ovarianitis and catarrhal salpingitis. Histological examination of the uterine section of the oviduct reveals basophilic intranuclear inclusions in the epithelial cells. Changes in the bursa of Fabricius are not pathognomonic.
Pathological diagnosis:
1. Ovariitis, hemorrhages in the ovary.
2. Catarrhal, catarrhal-hemorrhagic salpingitis.
3. Reduction in shell thickness (by 30-60%) and its depigmentation (in eggs of colored breeds) to a faint yellow or white color.
4. Gangrenous dermatitis (complication).
The decrease in egg production in chickens is taken into account, and early and retrospective diagnostics are used using RZGA.
Infectious bronchitis is characterized by damage to the respiratory tract (respiratory form) in chickens and the oviduct in adult birds with a decrease in egg production (reproductive form). At autopsy, dead and forcedly killed chickens show acute catarrh upper respiratory tract, serous-fibrinous aerosacculitis, in chickens - atrophy of the ovary and oviduct, deformation of the egg follicles. Histological examination also reveals granular degeneration of the epithelium of the convoluted tubules of the kidneys.
The nephroso-nephritis form of infectious bronchitis is recorded in chickens 3-9 weeks of age. In a sick bird, watery diarrhea, depression, and prostration are observed. Presenters pathological processes detected at autopsy and histological examination: granular and fatty degeneration kidneys and accumulation of urates in renal tubules and ureters, fatty degeneration (small-droplet) of the liver, pseudoeosinophilic reaction in the interstitium of these organs.
Pathological diagnosis:
respiratory form
1. Serous-catarrhal rhinitis, conjunctivitis.
2. Serous-catarrhal fibrinous tracheitis and bronchitis.
3. Focal catarrhal or catarrhal-fibrinous pneumonia.
4. Serous-fibrinous aerosacculitis.
5. Exhaustion.
nephroso-nephritis form
1. Nephroso-nephritis, accumulation of urates in the ureters.
2. Overflow of the rectum and cloaca with whitish feces mixed with urates.
3. Visceral uric acid diathesis.
reproductive form
1. Atrophy of the ovary and oviduct.
2. Cyst of the ovary with the presence of a caseous-fibrinous mass in its lumen.
3. Yolk peritonitis.
Additional research is carried out: isolating the virus on EC and CC, placing a bioassay on 9-day-old embryos (in which, already on the 5-6th day after infection of the embryos with the virus, a “dwarfism effect” is noted - a growth retardation of 3-4 times compared with control), as well as serological reactions - RN, RID, RNGA.
Bird flu affects chickens of all ages and is acute. Morbidity and mortality rates can reach 100%. The disease is characterized by nervous phenomena, swelling of the head, cessation of egg production, diarrhea, cyanosis of the comb and earrings followed by their necrosis. At autopsy, numerous pinpoint and spotty hemorrhages are found on the mucous membranes and serous tissues, a hemorrhagic ring in the mucous membrane of the glandular stomach at its border with the muscular stomach, and acute catarrhal enteritis. Histological examination reveals micronecrosis in the brain, and depletion of lymphocytes in the bursa of Fabricius, thymus, spleen, esophageal and cecal tonsils.
Pathoanatomical diagnosis:
1. Hemorrhagic diathesis.
3. Cyanosis of the comb and catkins.
4. Serous-fibrinous pericarditis and pleuroperitonitis.
5. Catarrhal, catarrhal-hemorrhagic enteritis.
6. Serous edema of the subcutaneous tissue.
7. Unchanged spleen.
Histo: micronecrosis in the brain; delymphatization of the bursa of Fabricius, thymus, spleen, esophageal and cecal tonsils.
Laboratory diagnosis is based on the isolation of the virus in chicken embryos, the production of RGA and RZGA with specific hyperimmune sera, and histological examination of the pathological material.
Newcastle disease affects birds of all ages and occurs acutely, subacutely and chronically. The acute course is characterized by fever, depression, drowsiness, diarrhea (watery feces, greenish-yellow in color). When breathing, you can hear wheezing and bubbling in the throat. Cyanosis of the comb and catkins is observed.
In subacute and chronic cases, nervous symptoms, shortness of breath, cough, and wheezing are observed.
The main pathological changes are characterized by the phenomena of hemorrhagic diathesis, the appearance of a hemorrhagic ring in the mucous membrane of the glandular stomach at its border with the muscular stomach. Fibrinous-necrotic, erosive-ulcerative enteritis with the formation of scab-buds, granular degeneration of parenchymal organs, venous hyperemia and pulmonary edema are also detected. Histopathological examination reveals non-purulent lymphocytic encephalitis in the brain; in the organs of the immune system - massive necrosis of lymphocytes, processes of destruction of lymphoid nodules.
Pathological diagnosis:
1. Hemorrhagic diathesis.
2. Hemorrhagic ring in the mucous membrane of the glandular stomach at its border with the muscular stomach.
3. Cyanosis of the comb and catkins.
4. Fibrinous-necrotic, erosive-ulcerative enteritis with the formation of scab-buds.
5. Serous edema of the subcutaneous tissue.
6. Slight enlargement of the spleen.
7. Histo: non-purulent lymphocytic encephalitis; death of lymphocytes and destruction of lymphoid nodules in the bursa of Fabricius and spleen.
To make a final diagnosis, additional laboratory tests are carried out: isolation of the virus on chicken embryos and its identification using RTGA, RN, RSK, RIF, ELISA. Data from histological examination of the brain are taken into account.
Streptococcosis occurs at lightning speed or acutely. In a sick bird, drowsiness, ruffled feathers, depression, diarrhea, anemic comb and earrings, and sometimes paralysis of the limbs are observed. Pathological changes are characterized by serous-hemorrhagic edema of the subcutaneous tissue, serous-fibrinous inflammation of the serous integument, and splenic hyperplasia. The liver is in a state of granular degeneration with miliary foci of necrosis.
Pathological diagnosis:
1. Cyanosis of visible mucous membranes and skin.
2. Serous-hemorrhagic edema of the subcutaneous and intermuscular tissue.
3. Venous hyperemia, granular and fatty degeneration of the liver, foci of necrosis in it.
4. Enlarged spleen.
5. Acute venous hyperemia and pulmonary edema.
6. Acute venous hyperemia, granular degeneration and serous glomerulonephritis of the kidneys.
7. Acute catarrhal enteritis.
8. Serous-fibrinous peritonitis, pericarditis, perisplenitis, ovariosalpingitis (with subacute and chronic course).
9. Ovarian hyperemia, deformation of egg follicles and hemorrhages in them.
Laboratory tests are carried out: microscopy of organ smears, isolation of a pure culture of the pathogen, determination of its virulent properties.
With chicken pasteurellosis, sick birds experience lethargy, thirst, fever, depression, diarrhea, cyanosis of the comb and earrings, and in a chronic course, diphtheritic inflammation of the wattles. When opening dead and forcedly killed chickens, signs of septicemia are found, lobar pneumonia. During histological examination, pronounced atrophy and delymphotization of lymphoid nodules with simultaneous thickening of interfollicular connective tissue septa are recorded in the bursa of Fabricius.
Pathological diagnosis:
acute course
1. Cyanosis of the comb and earrings.
2. Croupous pleuropneumonia.
3. Serous-fibrinous pericarditis.
4. Hemorrhagic diathesis.
5. Granular dystrophy and miliary necrosis in the liver and myocardium.
6. Acute catarrhal, catarrhal-hemorrhagic duodenitis.
7. Enlarged spleen (not always).
chronic course
1. Fibrinous-necrotic inflammation of the barbs, sometimes their falling off.
2. Croupous, necrotizing pneumonia.
4. Fibrinous-purulent arthritis.
5. Exhaustion.
Histo: atrophy of the lymphoid nodules of the bursa of Fabricius, thickening of the interstitium. To clarify the diagnosis, a bacteriological study is carried out and a bioassay is performed on chickens.
Colibacillosis is characterized by fever, depression, diarrhea, and increasing exhaustion. At autopsy they find: catarrhal inflammation of the small intestine, serous-fibrinous inflammation of the serous covers, air sacs. Microscopy of sections of the bursa of Fabricius reveals focal hemorrhages, hyperemia and edema of the interstitium, and atrophy of lymphoid nodules. A depletion of lymphoid elements in the cortical and medullary zones of the nodules and swelling of the cytoplasm of reticular cells are noted.
Pathological diagnosis:
1. Serous-fibrinous perihepatitis, perisplenitis, pericarditis, peritonitis, aerosacculitis.
2. Dot and spotty hemorrhages in parenchymal organs, mucous and serous membranes.
3. Acute catarrhal enteritis.
4. Enlarged spleen.
5. Granular dystrophy and miliary foci of necrosis in the liver.
6. Serous-fibrinous polyarthritis (with chronic course).
7. Histo: hemorrhages in the bursa of Fabricius, atrophy of lymphoid nodules, hyperemia and edema of the interstitium.
Additional research is carried out: isolating a culture of the pathogen, determining its virulence.
Pathological diagnosis:
1. Catarrhal-hemorrhagic erosive-ulcerative typhlitis and proctitis.
3. Granular dystrophy of the liver, kidneys and myocardium.
4. Exhaustion.
5. Histo: atrophy of the lymphoid nodules of the bursa of Fabricius, swelling of the interfollicular connective tissue, accumulation of histiocytes, pseudoeosinophils and eosinophils in it.
To make a final diagnosis, microscopy of feces or scrapings of the intestinal mucosa is performed to identify eimeria oocysts.
Prostagonymosis occurs in the form of enzootic outbreaks, most often on farms located near water bodies. In sick birds, depression, drowsiness, diarrhea (white or greenish feces), decreased egg production, and an enlarged abdomen are observed. When opening dead or forcedly killed birds, ovarianitis, purulent-fibrinous salpingitis and peritonitis, hyperemia and edema of the bursa of Fabricius are discovered. Microscopy of bursa sections reveals serous-inflammatory edema of the interstitium, infiltration of pseudoeosinophils and eosinophils.
Pathological diagnosis:
1. Ovariitis, deformation and rupture of follicles.
2. Purulent-fibrinous peritonitis.
3. Purulent-fibrinous salpingitis, egg stones in the lumen of the oviduct.
4. Acute catarrhal enteritis.
5. Granular dystrophy of the liver, kidneys, myocardium.
6. Hyperemia and swelling of the bursa of Fabricius.
7. Exhaustion, exicosis.
8. Histo: Serous-inflammatory edema and microphage infiltration of internodular connective tissue in the bursa of Fabricius.
Additional laboratory tests are carried out - microscopy of the droppings of sick birds to detect helminth eggs.
Hypovitaminosis A is recorded in chickens, as well as productive birds. Symptoms: immobility, stunted growth, conjunctivitis, laryngitis, general anemia. Upon opening, miliary gray-yellow nodules (hyperplasia and keratinization of the epithelium of the mucous glands) are found on the mucous membrane of the pharynx and esophagus, and granular degeneration and urate deposition are found in the kidneys.
Pathological diagnosis:
1. Hyperkeratosis of the skin, dullness and brittleness of feathers.
2. Fibrinous conjunctivitis, xerophthalmia, keratomalacia, panophthalmitis.
3. Catarrhal rhinitis, laryngitis, tracheitis.
4. Millet-like nodules in the mucous membrane of the pharynx and esophagus.
5. Visceral uric acid diathesis and gout.
6. Exhaustion and general anemia.
To establish a final diagnosis, a biochemical study of the liver, blood, egg yolks, and feed is carried out for the content of vitamin A and carotene.
Nutritional dystrophy, most often observed in chickens 15-20 days of age, is characterized by depression, diarrhea, and exhaustion of the bird. At autopsy, the absence of fat in the fat depot, atrophy of skeletal muscles and internal organs are recorded. Histological examination of sections of the thymus and bursa of Fabricius reveals early involution of the organ, accompanied by atrophy and devastation of lymphoid nodules.
Pathological diagnosis:
1. Lack of fat in the fat depot.
2. Atrophy of skeletal muscles and internal organs.
3. Serous edema in the subcutaneous and intermuscular tissue.
4. General anemia, exicosis.
5. Histo: accidental involution of the bursa of Fabricius and thymus.
A chemical and toxicological study of feed is carried out. An increase in acid and peracid numbers is detected.
Radiotoxicosis is recorded in birds of all ages. Sick chickens and chickens experience anorexia, thirst, and swelling in the head area. An autopsy reveals hemorrhagic diathesis, serous infiltration of subcutaneous tissue, and histopathology reveals atrophy and necrosis of lymphoid tissue in the central and peripheral organs of the immune system.
Pathological diagnosis:
1. Hemorrhagic diathesis.
2. Serous swelling of the subcutaneous tissue in the head area.
3. Subacute or chronic catarrhal, erosive and ulcerative enteritis.
4. Atrophy of the bursa of Fabricius, thymus, spleen, Harder gland.
5. General anemia, exhaustion, exicosis.
6. Histo: focal necrosis or atrophy of lymphoid tissue in the thymus, bursa of Fabricius, Harder's gland, spleen, esophageal and cecal tonsils.
Radiological examination of feed is carried out.
Prevention and control measures
In order to protect poultry farms from the introduction of the pathogen, veterinary specialists are obliged to: use eggs for incubation from poultry farms that are free from infectious bursal disease; equip poultry houses with birds of the same age; observe inter-cycle technological breaks with thorough cleaning, disinfection, disinfestation and deratization; carry out disinfection of eggs and containers being carried out; provide the necessary veterinary requirements for keeping and feeding birds.
When a diagnosis of IBD is established, restrictions are imposed on the farm, under the terms of which it is prohibited: the export of eggs for incubation, young and adult birds, feed, tools and equipment to farms free from IBD and for sale to the public; stocking flocks with recovered poultry.
It is allowed to sell eggs for food purposes after disinfecting them with formaldehyde vapor.
After clinical trial All sick and suspected chickens are slaughtered and disposed of. The remaining healthy chickens are vaccinated, followed by the slaughter of all birds that have reached slaughter conditions from the premises where the disease was recorded. Stop incubating eggs and accepting young animals for rearing. The incubator is disinfected. Laying eggs for incubation is resumed no earlier than 10 days after the hatching of the last batch of incubated eggs.
Separate maintenance personnel are assigned to each poultry house. Overalls are disinfected daily in a steam-formalin chamber.
Poultry slaughter is carried out with complete gutting of the carcasses. If there are no pathological changes in the carcasses, they are used after boiling at t=100°C (for 90 minutes). Carcasses with intramuscular hemorrhages, edema, and deposits of uric acid salts are sent along with internal organs for technical disposal.
For wet disinfection, use a 4% solution of sodium hydroxide, a clarified solution of bleach (containing at least 3% active chlorine), and a 2% aqueous solution of formaldehyde with an exposure of at least 6 hours.
Litter and deep litter are disinfected biothermally.
Restrictions are lifted after final veterinary and sanitary measures and in the absence of clinical and pathological changes in at least 3 batches of young animals reared up to 60 days of age after a preventive break.
Specific prevention
According to the observation of a number of researchers, general veterinary and sanitary measures do not provide complete recovery farms from infectious bursal disease. Therefore, in the complex of measures to prevent and eliminate this infection, the main place is given to specific prevention using live and inactivated vaccines.
Protection of chickens from infection with the IBD virus is achieved by creating a high level of passive antibodies in young animals by immunizing replacement chickens with inactivated vaccines and using live virus vaccines as transovarial immunity decreases.
Live virus vaccines are used to immunize chickens. Birds are vaccinated orally at 7-14 days of age (depending on the level of passive immunity and the epizootic situation), and revaccinated 14 days later. 6 hours before immunization, the supply of water and feed in poultry houses is stopped. The vaccine is dissolved in warm water and feed the chickens in such a way that there is one dose of vaccine (10-15 ml) per head. The supply of water and food is resumed no earlier than 2 hours after drinking the vaccine. Establish clinical observation of the condition of the bird.
The disadvantage of most live virus vaccines against infectious bursal disease is that the vaccine strains of the virus, along with immunogenicity, have pronounced immunosuppressive properties. Therefore, live vaccines, depending on the degree of reactogenicity, are divided into moderate-, weak- and apathogenic with high immunogenic activity. The effectiveness of reactogenic strains has been established when vaccinating chickens with high levels of maternal antibodies; they are unacceptable for susceptible chickens.
One of the indicators of the reactogenicity of vaccine strains for Gumboro disease is a decrease in the level of antihemagglutinins in birds vaccinated against Newcastle disease. A correlative relationship has been established between the immunogenic activity of vaccine strains of the virus, its damaging effect on the morphological structure of the bursa and a decrease in the bursal index. The most immunogenic strains cause more pronounced lesions of the bursa of Fabricius. A decrease in the bursal index was established when reactogenic vaccines were administered to weakly immune birds of early age. Therefore, before vaccinating chickens against IBD, we recommend determining their immune status and background levels of specific antibodies. To do this, send blood serum and immune organs from 10-20 chickens to the laboratory of diseases of birds and bees of the BelNIIEV or to the Department of Small Animal Diseases of the VGAVM for research.
It has been shown that even weakly pathogenic vaccines against IBD are capable of causing pathomorphological changes in the organs of the immune system, thereby causing the development of an immunodeficiency state in birds. For example, a single immunization of chickens with the dry live virus vaccine “Bursin-2” leads to a sharp decrease in the size of lymphoid nodules due to the processes of delymphotization of their cortical layer. In the medulla, phagocytosis of destroyed cells is recorded with the formation of many round cavities like a honeycomb. In chickens vaccinated twice with this vaccine, the lesions of the bursa of Fabricius intensify. There is an almost complete disappearance of lymphocytes, atrophy and destruction of lymphoid nodules, fibroplasia and transformation of the organ into glandular structure. Cortical atrophy is observed in the thymus.
Double vaccination of chickens with attenuated vaccines “Gambovac” and “Humboral-ST” (made in France) causes almost complete atrophy of the bursa of Fabricius with the loss of its functional part.
The results of our studies showed that when administered orally, a dry live virus vaccine from pcs. “D 78” (produced in Holland) in immune chickens, processes of atrophy and delymphatization develop in the thymus, bursa of Fabricius and cecal tonsils, leading to immunosuppression.
We have also established that oral immunization of birds against Gumboro disease with a dry live virus vaccine from pcs. “Winterfield 2512” (produced by the All-Russian Research Institute of Animal Health, Vladimir) is accompanied by active immunomorphological changes in the organs of the immune system and does not cause an immunodeficiency state. In the bone marrow of chickens immunized with this vaccine, we recorded an increase in the number of myeloblastic cells, an increase in the leukoerythroblastic index, as well as the bone marrow index of eosinophil and pseudoeosinophil maturation. In the thymus of immune chickens, an increase in the specific volume of lymphoid tissue was observed, and in the bursa of Fabricius, hyperplasia of lymphoid nodules was observed. In the spleen and cecal tonsils of chickens immunized with this vaccine, we detected an increase in the density of nodular lymphoid tissue, an increase in the size of lymphoid nodules, as well as active accumulation of plasma cells. In this regard, the use of dry live virus vaccine from pcs. “Winterfield 2512” (manufactured by ARRIAH) is most preferable for immunizing chickens against Gumboro disease, given the high immunogenicity and low reactogenicity of this biological product.
BelNIIEV has developed a live embryonic virus vaccine against Gumboro disease from the KMIEV-15 strain. It is currently undergoing extensive production testing. Studying the effectiveness of this biological product is of particular interest, given the antigenic relationship of the vaccine and epizootic strains of the virus circulating in the Republic of Belarus.
At the age of 110-120 days, inactivated vaccines are used for replacement young animals. They are administered once, intramuscularly, into the pectoral muscle area in a dose of 0.5 ml. The vaccine injection site is treated with 70% ethanol. It has been established that the introduction of inactivated vaccines increases the antibody titer in birds previously immunized with live vaccines. The immunizing effect is more pronounced the later it was used inactivated vaccine. Immunization of replacement chickens with inactivated vaccines creates more intense and long-lasting immunity than when using live vaccines.
Despite the immunization of replacement chickens, there is often significant variability in passive antibodies in a flock of chickens, which creates conditions for bird diseases with weak level transovarial immunity. This necessitates the need to stimulate the post-vaccination immune response through the use of immunotropic agents.
We have conducted studies to study the influence of a number of immunostimulants (sodium thiosulfate, thymalin, ASD-2, levamisole) on the immunogenic properties of the liquid sorbed inactivated vaccine of the ARRIAH against infectious bursal disease (manufactured in Vladimir, Russia). It has been established that immunization of replacement chickens together with sodium thiosulfate and thymalin helps to create a more intense antiviral immunity than when using a single vaccine. The drugs ASD-2 and levamisole did not significantly affect the immunogenicity of this vaccine.
Method of immunization of replacement young chickens against IBD with liquid sorbed inactivated vaccine ARRIAH together with sodium thiosulfate:
The bird is vaccinated at 110-120 days of age. First prepare a fresh 35% sodium thiosulfate solution. The resulting solution is sterilized by boiling for 30 minutes, and after cooling, 50 ml of a 35% aqueous solution of sodium thiosulfate is mixed with 200 ml (bottle - 400 doses) of the vaccine. The resulting mixture (containing 7% sodium thiosulfate) is administered once, intramuscularly, into the pectoral muscle area, in a dose of 0.6 ml (thus, the volume of the administered vaccine increases by 0.1 ml). The injection site is treated with 70% ethanol.
We have established that immunization of replacement young animals against Gumboro disease with a liquid sorbed inactivated vaccine (produced by the All-Russian Research Institute of Animal Health, Vladimir) together with the immunostimulant sodium thiosulfate (in a 7% aqueous concentration) activates the processes of immunomorphological restructuring in the central (red bone marrow, thymus, bursa Fabricius) and peripheral (spleen, blood) organs of the immune system of birds, which helps to increase the level of specific antibodies by 20-50%., creating more intense post-vaccination immunity in breeding stock, and thereby enhancing immune protection in chickens due to a high level of transovarial immunity. .
Method of immunization of replacement young chickens against IBD with liquid sorbed inactivated vaccine ARRIAH together with thymalin:
The bird is immunized at 110-120 days of age. On the day of vaccination, 10-20 heads of replacement chickens are selectively weighed to determine the average weight of the birds. Timalin is administered together with the vaccine at a dose of 1 mg/kg body of birds. The drug is available in powder form in 10 mg bottles.
Example. The average weight of birds is 2 kg. Therefore, 1 dose of the vaccine should contain 2 mg of thymalin. 800 mg (80 bottles) of thymalin are dissolved in a vaccine bottle (200 ml - 400 doses). The resulting mixture is administered once, intramuscularly, into the pectoral muscle area, in a dose of 0.5 ml. The injection site is treated with 70% ethanol.
Our studies have shown that immunization of birds together with thymalin promotes an increase in specific antibodies in the blood by 1.5-4.4 times compared to the use of a vaccine without an immunostimulant.
At the same time, sodium thiosulfate is a cheaper and more accessible drug. Therefore, its use during the period of immunization of replacement chickens is preferable.
Assessment of the intensity of post-vaccination immunity.
To assess the intensity of immunity in Gumboro disease, serological methods are used: RID, VIEF, RN, RNGA and ELISA. In all chickens vaccinated with live virus vaccines, antibodies are detected in the blood serum starting from 20-30 days of age. Specific antiviral antibodies in replacement chickens vaccinated at the age of 110-120 days are detected in chickens up to 46 weeks of age. Maternal antibodies are detected in chickens up to 17-20 days of age.
19.04.2018
Vaccines for the prevention of chicken infectious bronchitis (IB) and Gumboro disease are live and inactivated preparations of imported and domestic production. These drugs are produced in the form of monovalent as well as polyvalent vaccines that protect poultry from 2–4 different infectious diseases. A huge role here is played by live vaccines that provide a rapid immune response, including drugs of modern technologies.
Vaccination of birds against IBV is carried out intraocularly, intranasally, orally and by spray method (coarse and fine spray), against Gumboro disease - orally and in ovo.
Infectious bronchitis of chickens
Infectious bronchitis of chickens (IB) is a highly contagious viral disease, manifested by damage mainly to the respiratory and reproductive organs and kidneys. The disease affects birds of all ages and is especially dangerous for chickens.
The causative agent of IB
Caused by an RNA virus with high genetic variability. Mutations of the pathogen are facilitated by a number of factors, including the reproduction of the virus in poultry of different ages, mixed infections, and the co-circulation of vaccine and field viruses in the same flock.
There are many strains of chicken infectious bronchitis virus circulating around the world, which makes the diagnosis and prevention of this disease difficult. Currently, the greatest danger to the Russian poultry industry is posed by viral strains of the Massachusetts serotype, viruses of serotype 793B, as well as highly contagious QX strains and some other pathogens. Several strains circulate simultaneously in poultry farms, but 1–2 main serotypes usually predominate.
Certain serotypes of infectious bronchitis virus can multiply in various tissues of the bird's body.
IBV serotype Massachusetts (Mass) primarily affects the respiratory system, causing severe respiratory disease. Immunization against Massachusetts serotype viruses can be carried out from the first days of life.
Respiratory strains of IBV viruses lead to death (mortality rate 15–35%) and create a favorable background for the development of bacterial infections.
The Massachusetts serotype has become widespread throughout the world and was identified in the 40s of the twentieth century in Europe and the USA.
Later it turned out that a number of strains of IBV viruses also infect the excretory organs, and respiratory symptoms can be expressed to varying degrees.
The nephroso-nephritis form of IB is characterized by weak and short-term respiratory signs followed by depression, the mortality rate of young animals ranges from 25–30% to 70%. Nephropathogenic properties are most pronounced in the QX strain, which came to Europe from Asia and has been circulating in Russia since the early 2000s.
The highly contagious and pathogenic strain QX actively reproduces in the tissues of the respiratory tract, kidneys, ovaries and lymphoid tissues of the cecum and colon.
In the 90s of the last century, scientists identified the nephropathogenic serotype 793B, which causes high mortality in broilers. Immunization of chickens against this pathogen is usually carried out during the second vaccination. But there are vaccines that can be used from one day of age.
Serotypes close to Massachusetts, as well as 793B, are the most common strains of IBV viruses in poultry farms in Russia and Europe; the largest number of immunobiological drugs have been developed to protect against these strains. When used together, sufficient cross-protection against the QX strain is formed.
Some strains of the IBV virus (including M41) can reduce the egg production of poultry for a long period (causing a drop in productivity parameters from 30 to 89%), deteriorate the quality of egg shells and change their color.
As a result, laying hens are found on farms with normally expressed secondary sexual characteristics, but unable to lay eggs due to adhesions of the oviducts as a result of salpingo-oophoritis, provoked by the multiplication of the IBV virus in chickens at an early age.
Among the new virulent strains that have appeared over the past few years, it is necessary to highlight VAR 2 (Variant 2), circulating in Asia and the Middle East, and recently in Central Europe. This virus mainly affects the kidneys, respiratory and reproductive organs.
Considering the rapidly increasing infectious pressure of IBV types QX and 793B and new variant strains of type 2 in Europe, the Middle East and Russia, Phibro Animal Health Corporation (USA) has developed a live attenuated vaccine TABIC IB VAR 206. It was created on the basis of a field strain Option 2 (IS/1494/06).
The TABIK IB VAR 206 vaccine is produced using the TAbic technology, patented and owned by Phibro (production of live vaccines in the form of sterile water-soluble tablets). This promising development has been appreciated by other major vaccine manufacturers.
Boehringer Ingelheim, based on scientific developments in the production of effervescent forms of Sanofi under license from Phibro Animal Health, began producing vaccines in the form effervescent tablets(NEO line). This dosage form significantly reduces storage space and facilitates the work of veterinarians.
During the study of strains of infectious bronchitis pathogens in chickens, interesting facts were revealed. For example, a combination of some strains of IBV pathogens can cause cross-protection from other strains. For example, immunization with strains Ma5 of serotype Massachusetts and serotype 793B protect birds from the highly pathogenic strain QX (vaccination occurs with one strain, revaccination with another strain). The phenomenon of such a synergistic effect of vaccines is called protectotype. It was discovered by Jane Cook, and today it is the main concept in IB immunity.
IBC protection programs
The infectious bronchitis virus is constantly changing. PCR studies on sequencing the IBV genome, carried out in the early 2000s in Russia, showed that about a third of the viruses were previously unstudied and constituted a group of local strains.
Programs to protect chickens from infectious bronchitis include live and inactivated vaccines. The main purpose of vaccination is to develop immunity in birds against a wide range of viral agents. Immunization against IBV is carried out once or twice (depending on the recommendations of the drug manufacturer and the epizootic situation on the farm). Chicks are vaccinated from the first day of life, regardless of the level of maternal bodies. A third (additional) revaccination against IBV should be carried out in both parent and commercial flocks before the start of laying (on days 98–120).
Live vaccines are the main tool for protecting chickens from parent flocks, broilers and laying hens from IBV. They create early specific protection that develops in the chicken within 2 weeks. The main disadvantage of live vaccines is the potential ability of the vaccine strain to revert to the wild type and restore virulence due to mutations. Viruses of respiratory diseases are able to compete for the same receptor sites in the mucous membrane of the upper respiratory tract. Therefore, when using two live vaccines with a different set of strains in an IBV immunization regimen, it is necessary to maintain a time interval of at least 14 days.
Inactivated vaccines used for young laying hens and parent flocks (re-vaccination), they cause the production of maternal antibodies. In order for vaccination with inactivated vaccines to be effective, it is necessary to first use a live vaccine for primary antigen exposure, at least four to five weeks before using the inactivated vaccine. The content of several heterologous strains in an inactivated vaccine causes the formation of high levels of antibodies to a larger number of strains of the IBV virus. Primary vaccination with a live vaccine and revaccination with an inactivated one provide protection on average in 95% of cases, while the combination of two inactivated immunobiological drugs provides approximately 90% protection.
Identification of the type of virus circulating in the herd using PCR and other methods in specialized laboratories will help you choose the right drug for immunization.
From the moment they are born, chickens are at risk not only of IBV, but also of Newcastle disease. A fairly large number of drugs have been created for comprehensive protection against these diseases.
The company Ceva Sante Animale proposed a vaccine for vaccinating day-old chicks against Newcastle disease and infectious bronchitis (strain H-120) VITABORN L.
A high-quality vaccine against infectious bronchitis of chickens BRONIPRA-1 (strain H-120) for use on the first day of life is offered by the Spanish company Laboratorios HIPRA, S.A. For areas unaffected by Newcastle disease, the company also has bivalent vaccines HIPRAVIAR-B1/H120 and HIPRAVIAR-CLONE/H120, which are successfully used from the first day of life using the large-droplet spray method.
Monovaccines for the prevention of infectious bronchitis in chickens
|
Vaccine |
Description |
Strain and serotype of the pathogen |
Manufacturer |
|
AVIVAC-IBC |
live dry |
A/91 serotype 793/B |
NPP "AVIVAC", Russia |
|
AVIVAC-IBC |
live dry |
H-120 serotype Massachusetts |
NPP "AVIVAC", Russia |
|
Bioral H120 NEO |
live tablet |
H120 serotype Massachusetts |
Boehringer Ingelheim, France |
|
BRONIPRA-1 |
live dry |
H-120 serotype Massachusetts |
|
|
Vaccine against infectious bronchitis of chickens from the variant strain RV-07 live dry |
live dry |
strain RV-07 |
|
|
live dry |
H-120 serotype Massachusetts |
FSBI "ARRIAH", Russia |
|
|
Vaccine against infectious bronchitis of chickens from strain N-120 live dry |
live dry |
H-120 serotype Massachusetts |
OJSC "Pokrovsky Plant of Biological Preparations" |
|
Vaccine against infectious bronchitis of chickens, multi-strain inactivated emulsified |
inactivated emulsified |
Taganrog serotype 793/B + Kaluga strain + H-52 serotype Massachusetts |
FSBI "ARRIAH", Russia |
|
Live dry virus vaccine against chicken infectious bronchitis (IBV) from strains N-120, RV-07 |
live dry |
H-120 serotype Massachusetts, variant strains RV-07 |
Kronvet, Russia |
|
Volvac IB Mass MLV |
live dry |
modified Massachusetts serotype virus |
Boehringer Ingelheim, Germany |
|
Galliwack IB 88 |
live lyophilized |
CR88121 serotype 793B |
Boehringer Ingelheim, France |
|
Galliwack IB 88 NEO |
live tablet |
CR88121 serotype 793B |
Boehringer Ingelheim, France |
|
Gallimun 793B |
dry inactivated |
variant strains of serotype 793B |
Boehringer Ingelheim, France |
|
live dry |
H-120 serotype Massachusetts |
FKP “Shchelkovo Biocombine”, Russia |
|
|
Nobilis IB 4/91 |
live dry |
4/91 serotype 793B |
Intervet/MSD, Netherlands |
|
Nobilis IB Ma5 |
live dry |
Ma5 serotype Massachusetts |
Intervet International/MSD, Netherlands |
|
Pulvak IB H120 |
live dry |
H-120 serotype Massachusetts |
Zoetis Inc., USA |
|
Pulvak IB QX |
live dry |
serotype QX (L1148) |
Zoetis Inc., USA |
|
Pulvak IB Primer |
live dry |
H-120 serotype Massachusetts + variant strains D274 |
Zoetis Inc., USA |
|
Sevak iBird |
live dry |
1/96 serotype 793B |
Ceva Sante Animale, France |
|
Sevak MASS L |
live dry |
B-48 serotype Massachusetts |
Ceva Sante Animale, France |
|
Sevak BRON 120 L |
live dry |
H-120 serotype Massachusetts |
Ceva Sante Animale, France |
|
TABIK H-120 |
live dry tablet |
H-120 serotype Massachusetts |
Phibro Animal Health, Israel |
|
TABIK IB Var |
live dry tablet |
233A serotype 793B |
Phibro Animal Health, Israel |
|
TABIK IBVAR2-06 |
live tablet |
Phibro Animal Health, Israel |
|
|
HatchPack IB H120 |
live frozen |
H-120 serotype Massachusetts |
Boehringer Ingelheim, France |
Immunization of chickens against IBV can also be carried out polyvalent drugs:
– produced by Ceva Sante Animale: Sevak Megamun ND-IB-EDS-SHS K, Sevak NB L, Sevak VITABRON L;
– produced by Intervet/MSD: Nobilis Ma5 + Clone 30, Nobilis IBmulti + ND + EDS, Nobilis IBm + ND + EDS, Nobilis RT + IBmulti + G + ND;
– produced by Laboratorios HIPRA, S.A: HIPRAVIAR-TRT4, HIPRAVIAR-CLONE/H120, HIPRAVIAR-B1/H120, AVISAN MULTI;
– produced by Boehringer Ingelheim: Volvac ND + IB + EDS KV, Gallimun 303, Gallimun 407;
– produced by Abic Biological Laboratories Ltd (a division of Phibro Animal Health): VH + H120, Quadractin VP 2, SSY + NB + IBK;
– produced by Zoetis: Provak 4, Pulvak Aero;
– produced by AVIVAC: AVIVAC-IBK + NB, AVIVAC NB + IBK + IBB + SSYA + REO
and some other vaccines.
Gumboro disease
Gumboro disease, or infectious bursal disease (GD, IBD) is highly contagious viral disease chickens 2–20 weeks of age, accompanied by damage to the bursa of Fabricius, to a lesser extent to other lymphoid organs and kidneys, the presence of hemorrhages in the muscles of the thigh, chest, wing and in the mucous membrane of the glandular stomach. Along with Marek's disease, IBD is the main immunosuppressive disease in poultry.
IBB - a blow to poultry farming
The Gumboro virus was first discovered in the 50s of the twentieth century in the USA. Today it circulates in all countries of the world with developed poultry farming and causes great economic damage. From time to time, outbreaks of highly virulent strains of IBD are recorded in Europe, leading to mortality of 10 to 30% of young poultry.
The pathogen is resistant to external environment. In droppings, water, and feed, it does not lose its infectious properties for 56 days; on poultry farm equipment - up to 122 days or more.
Infectious bursal disease can occur in both acute and subclinical forms, accompanied by retardation in the growth and development of chickens, suppression of their immunity, susceptibility to viral, bacterial and other diseases.
The subclinical form of the disease, no less than its acute course, causes significant damage to farms. According to Intervet/MSD, the profit from raising broiler flocks free of Gumboro disease is on average a third higher than that obtained from raising birds with subclinical disease.
It is possible to detect the IBD virus by ELISA, PCR, diffuse precipitation reaction on agar gel and some other methods.
Modern methods of protection
Live vaccines IBB is used for vaccination of healthy broiler chickens and replacement young animals of meat and egg breeds. They provide rapid formation of immunity. The frequency of vaccination is double or single, depending on the recommendation of the manufacturer of a particular drug. Live Gumboro disease vaccines are given 6 to 8 weeks before the inactivated vaccine. The disadvantages of live IBD vaccines include immunosuppression, which provokes an insufficient response to vaccination and increases the likelihood of developing other infectious and invasive diseases.
The range of industrial strains is quite extensive. For example, vaccines containing a moderately attenuated strain of the Gumboro disease virus are produced, such as AviPro Presize (Elanco) - LC-75, Nobilis Gumboro 228E (Intervet/MSD) - strain 228E, HIPRAGAMBORO-GM97 (Laboratorios HIPRA, S.A.). To protect poultry, the intermediate vaccine strain Winterfield 2512 is used, which is part of the imported and domestic immunobiological preparations Sevak TRANSMUN (Ceva Sante Animale), HIPRAGAMBORO-CH/80 (Laboratorios HIPRA, S.A.), AVIVAC-IBB (NPP AVIVAC). There are vaccines containing weakly attenuated (hot) strains, for example TABIC MV (Phibro Animal Health) - strain MV, etc.
Prevention of Gumboro disease is complicated by the presence of maternal heterogeneous antibodies in chickens. With a high level of maternal antibodies, the vaccine virus is quickly recognized and neutralized by the cells of the chicken's immune system.
Thanks to special innovative technology, Ceva Sante Animale specialists created immune complex vaccine Sevak TRANSMUN, which allows solving the problem of preventing Gumboro disease in chickens with a heterogeneous level of maternal antibodies. The vaccine is administered once to chicken embryos at the age of 18.5 days using the in ovo or day-old broiler chickens subcutaneously. Once the vaccine virus has replicated, the immune response culminates in the production of protective antibodies against Gumboro disease.
At the origins of the creation of immunocomplex vaccines was the company Embrex, owned by Zoetis, a manufacturer of equipment for in ovo vaccinations.
There are other similar drugs. The Zoetis company has developed the drug Bursaplex based on strain 2512 of the Gumboro virus and antibodies from the hyperimmune blood serum of SPF chickens.
Cloned live vaccine HIPRAGAMBORO-CH/80 has a minimal immunosuppressive effect on the bird's body and has high antigenic activity and immunogenicity. Intended for use in safe, unfavorable and endangered breeding and commercial poultry farms. Contains a culture of fibroblasts of SPF embryos of chickens infected with the cloned virus CH/80 of the Gumboro disease strain Winterfield 2512. Chickens are vaccinated twice starting from the age of 7 days.
Along with immune complex preparations, recombinant vaccines do not require monitoring the level of maternal antibodies.
Recombinant live vaccine Vaxitek HVT+ IBD is manufactured by leading health protection expert Boehringer Ingelheim. The vaccine contains the V2 gene, cloned from the Faragher 52/70 strain, using the turkey herpes virus as a vector. The drug is prescribed to chickens of meat and egg breeds once at one day of age or in ovo and provides protection against both classical and variant and highly virulent strains.
Thanks to ongoing developments in the field of recombinant and immunocomplex vaccines, it is possible to take a step towards eradicating a number of animal viruses.
But it is too early to write off classical drugs. A properly selected traditional live vaccine based on an intermediate strain provides the necessary level of protection. This is evidenced by a number of studies, including those from Phibro Animal Health specialists.
Domestic manufacturers offer vaccines with a wide range of current vaccine strains. The drugs are produced on modern equipment and meet international standards. A great contribution to the protection of poultry health and ensuring food security in Russia is made by the vaccines of NPP AVIVAC, FKP Shchelkovo Bioplant and FGBU ARRIAH.
In 2018, a new drug was registered by the Federal Center for Animal Health (ARRIAH). The basis of the live dry vaccine Gamboromix is a combination of Gumboro disease strains Winterfield 2512 and GD, positioned as “intermediate” and “hot” variants of the virus.
Monovalent vaccines against Gumboro disease
|
Vaccine |
Strain |
Manufacturer |
|
|
AviPro Presize |
live dry |
Elanco, Germany |
|
|
AVIVAC-IBB |
live dry |
NPP "AVIVAC", Russia |
|
|
AVIVAC-IBB |
live dry |
NPP "AVIVAC", Russia |
|
|
AVIVAC-IBB |
live dry |
Winterfield 2512 |
NPP "AVIVAC", Russia |
|
AVIVAC-IBB |
liquid inactivated |
NPP "AVIVAC", Russia |
|
|
Bursaplex |
live dry |
2512 + antibodies of hyperimmune blood serum of SPF chickens |
Zoetis Inc., USA |
|
Bursin Plus |
live dry |
Lukert, protein stabilizer H |
Zoetis Inc., USA |
|
Vaccine against infectious bursal disease from the VNIVIP strain, live, dry |
live dry |
FKP “Shchelkovo Biocombine”, Russia |
|
|
Virus vaccine against infectious bursal disease from the BG strain |
live dry |
FSBI "ARRIAH", Russia |
|
|
Virus vaccine against infectious bursal disease from the Winterfield 2512 strain |
live dry |
Winterfield 2512 |
FSBI "ARRIAH", Russia |
|
Gamboromix |
virus vaccine against infectious bursal disease live dry |
Winterfield and BG |
FSBI "ARRIAH", Russia |
|
Nobilis Gumboro D78 |
live dry |
Intervet/MSD, Netherlands |
|
|
Nobilis Gumboro 228E |
live dry |
Intervet/MSD, Netherlands |
|
|
Pulwak Bursa F |
live dry |
Zoetis Inc., USA |
|
|
dry live |
Phibro Animal Health, Israel |
||
|
SEVAK IBD L |
live dry |
Winterfield 2515, G-61 |
Ceva Sante Animale, France |
|
SEVAK GUMBO L |
live dry |
Ceva Sante Animale, France |
|
|
HIPRAGAMBORO-SN/80 |
live dry |
Winterfield 2512, clone CH/80 |
Laboratorios HIPRA, S.A., Spain |
|
HIPRAGAMBORO-GM97 |
live dry |
Laboratorios HIPRA, S.A., Spain |
|
|
Transmoon IBD |
dry live |
Winterfield 2515 + immunoglobulin complex from hyperimmune blood serum of SPF chickens |
Ceva Sante Animale, France |
Inactivated vaccines against IBD are used as part of polyvalent drugs for parent stock. They ensure the creation of the proper level of maternal antibodies in chickens.
Polyvalent vaccines against Gumboro disease are presented:
– Nobilis RT + IBmulti + G + ND (Intervet/MSD);
– HIPRAVIAR-TRT4 (Laboratorios HIPRA, S.A.);
– Sevak ND-IB-IBD-EDS K (Ceva Sante Animale);
– Provac 4 (Zoetis Inc.);
– Vaxitek HVT + IBD, Bursa Guard REO (Boehringer Ingelheim);
– Quadractin VP 2 (Abic Biological Laboratories Ltd, a division of Phibro Animal Health)
and some other vaccines, including those manufactured in Russia.
Number of impressions: 2243
Author: V. Lavrenova, marketer at the publishing house "Agricultural Technologies"
Broilers are one of the most popular crosses. Alas, the rapid growth and precocity of these chickens is associated with some health problems. In particular, it has become not uncommon for a whole population of broilers to be decimated by Gumboro disease. Let’s find out what it is and how to protect your household right now!
What is Gumboro disease?
Gumboro disease, also called infectious bursal disease, was first recorded in the United States in 1962 in the city of Gumboro, which gave the name to the disease. Later, outbreaks of a similar disease were recorded in Mexico, England, and Belgium. Currently, outbreaks have been recorded on all continents. The disease is caused by a virus of the Birnaviridae family.
The main “target” of Gumboro disease is leukocytes, which are actively destroyed in the bursa of Fabricius and other organs of the immune system (thyroid gland, spleen, almond gland); the kidneys are also severely affected.
Bursal disease can affect broilers at any age, but chickens between 2 and 9 weeks of age are particularly at risk.
Its danger is that it is very quickly transmitted from one individual to another, and infection can occur both by contact and through food, water, and equipment. Because of this, on large industrial enterprises there is a risk that the staff themselves may become carriers of the virus. Gumboro disease has a very severe consequences and is associated with significant financial losses. Not only does infection among livestock occur quite quickly, the destruction of leukocytes is associated with the onset of immune depression in birds. Sick broilers become very vulnerable and often begin to suffer from colibacillosis, coccidiosis, enteritis, which most often leads to the death of the bird.
The virus that causes this disease is quite stable and persists in the external environment for a long time. For example, in the droppings of infected birds, in water or feed, it persists for up to 56 days. On inventory and equipment of poultry farms even longer - more than 120 days.
Symptoms
There are two types of disease:
- Clinical type.
- Hidden (subclinical) type.
The clinical type of Gumboro disease is characterized by acute course and obvious external manifestations. It is most often recorded in broilers at the age of 3-6 weeks. The incubation period does not last long - a maximum of 1-2 days; infection of the livestock occurs very quickly.
The main symptoms of Gumboro disease are:
- severe diarrhea, mostly white;
- ruffled plumage, weakness and depression of the bird;
- chills, significant loss of appetite, signs of incoordination may be observed;
- dehydration and susceptibility to pathogenic organisms.
Most often, outbreaks of the disease can be short-term in nature - about 2-3 days. After recovery, the birds continue to experience a decrease in immunity for some time. Mortality from bursal disease, as a rule, reaches 5-6%, however, it can be much higher - 40% or more.
The hidden type, although it does not have such obvious external manifestations, is considered more insidious and dangerous. It can manifest itself in a general depressed state in birds. The latent course of Gumboro disease is associated with deterioration in feed conversion, stunted growth of broilers and their immune vulnerability.
Treatment
There is no specific treatment for Gumboro disease. The best way The way to prevent an unwanted epidemic is vaccination. For these purposes, so-called live and inactivated vaccines are used.
It is very important to detect foci of the disease in time and isolate the sick bird. Particularly affected individuals are recommended to be killed.
Pathological changes that will help confirm the diagnosis are as follows:
- enlarged and edematous bursa of Fabricius, it can be yellowish to brown due to areas of hemorrhage;
- hemorrhages in the kidneys and muscles;
- dehydration and anemia.
Since the virus is very stable, it is recommended to “relocate” sick birds from the chicken coop to another place. And in the room, carry out repeated treatment with formaldehyde and phenol.
Video “Diseases of chickens”
The video below will tell you what ailments befall the feathered inhabitants of farmsteads!
Infectious bursal disease of chickens
Infectiosis Bursitis gallinarum (Gumboro disease) An acute viral disease of chickens and turkeys, mainly 2-15 weeks of age, characterized by inflammation of the bursa of Fabricius, joints, intestines and internal hemorrhages.
HISTORICAL REFERENCE- the disease was first recorded in 1956 in Gumboro County (USA). In 1962, Kostrov described Gumboro disease as a disease. Winterfeld and Hitchner (1962) isolated a virus from sick chickens that caused nephroso-nephritis in sick broilers. Therefore, this disease is sometimes called nephroso-nephritis. Later, Karnayup (1965) proved that the symptoms of nephroso-nephritis are concomitant, the main and permanent changes are found in the bursa of Fabricius, which is why the disease began to be called infectious bursitis.
The disease is widespread in many countries of America, Europe, and Asia, where industrial poultry farming is developed. Data from serological studies show that the infection rate of herds ranges from 2 to 100%. And the reason for this is considered to be the constant import of poultry.
PATIENT- RNA virus from the genus Aviovirus of the Reoviredae (reovirus) family. The virion size is 70-75 nm. When 9-day-old embryos are infected in the yolk sac, the virus causes their death after 6 days. In addition to growth retardation, it causes
the appearance of edema, necrotizing lesions in the liver, which are typical for all viruses of this group. 3 days after the introduction of virus-containing material into the fibricium bursa, changes characteristic of a natural infection occur. In chicken embryo fibroblast culture, the virus causes a cytopathic effect. Virus-neutralizing and precipitating antibodies are formed in recovered birds.
RESISTANCE - the virus is resistant to ether, chloramine and pH 2.0, sensitive to trypsin. Indoors, the virus persists in droppings for 52 days. At 56°C it does not die within an hour. A solution of chloramine (0.5%) inactivates the virus in 10 minutes, formaldehyde (0.5%) in 6 hours.
EPISOOTOLOGICAL DATA- chickens of all ages are susceptible to the pathogen, but especially broilers aged 2-15 weeks. The most sensitive are 3-6 week old White Leghorn chickens. In adult chickens the disease is asymptomatic.
The source of the infectious agent is sick chickens that shed the virus in their droppings.
Infectious bursitis is an extremely contagious disease that is easily transmitted when birds are housed in close quarters. Chickens become infected through contaminated feed and water. A vertical route of transmission of the virus through infected eggs cannot be ruled out. Infected care items, equipment, clothing, and personnel play a certain role in the transmission of the pathogen.
The possibility of spreading the virus through the air has been proven. The reservoir of the pathogen can be flour beetles.
In fresh epizootic foci, the disease is acute and subacute, and in stationary outbreaks it is chronic and asymptomatic. In a number of farms, an immunizing subinfection is mainly recorded among birds.
PATHOGENESIS- consists of damage to lymphoid tissues, and first of all, lymphocytes of the bursa of Fabricius, spleen, and caecal glands of the blind processes are destroyed. The virus penetrates the digestive tract and after 24-48 hours is localized in the bursa of Fabricius, infecting B lymphocytes.
CLINICAL SIGNS- incubation period 1-2 days. Occurs in chickens under 3 weeks of age in the form of immunosuppression, which is manifested by increased sensitivity to bacterial infections.
It can occur in an acute form in the first 5-7 days after the disease in chickens aged 3 to 6 weeks. In case of low poultry resistance, mortality can reach 90%.
One of the first signs is diarrhea, with the release of yellow, liquefied droppings, or mucous-watery, white droppings, and feathering is impaired.
Then there is sudden apathy, trembling, signs of damage to the nervous system. The bird soon loses the ability to move and dies in a state of prostration.
Maximum mortality for 3-4 days from the beginning of the disease outbreak,
then the mortality rate decreases.
When the disease lasts 6-8 days, morbidity is 10-20% of birds, mortality is 1-15%.
Hematological changes are characterized by lymphopenia and erythrocytosis. Over 2 days of illness, the total number of leukocytes decreases, on the 5th day it increases and reaches a maximum on the 7th day after infection.
PATHOLOGANATOMICALCHANGES- the corpses are well-fed, but the muscles are dehydrated and pale, the goiter is empty, multiple pinpoint and striped hemorrhages are revealed, especially often under the skin of the thigh; the muscles are dark purple.
The bursa of Fabricius is greatly increased in volume, more than 2 times, and contains gelatin-like transudate; there are fibrinous deposits in the folds of the bursa, and in severe cases there is bloody fluid.
Swelling of the liver, necrotic foci, and atrophy of the spleen are noted. The pancreas is changed, nephrosis. In the final stage of the disease, swelling of the kidneys and atrophy of the bursa of Fabricius appear. Partial banded hemorrhages in the degenerated skeletal muscles of the myocardium, serous membranes, glandular stomach and intestines.
The most typical histological changes are necrosis
lymphoid elements of the bursa of Fabricius, thymus, spleen, renal degeneration.
DIAGNOSIS- infectious bursitis is a difficult-to-detect infection that spreads unnoticed, is masked by other diseases and physiological disorders, and only with a typical course is it relatively easily diagnosed based on clinical and pathological signs. They take into account the high percentage of morbidity, rapid spread and relapse within 5-7 days. The diagnosis can be confirmed by the detection of characteristic changes in the bursa of Fabricius.
For the final diagnosis, histological studies are carried out and a bioassay is performed by infecting 9-day-old chicken embryos on the chorioallantoic membrane. Embryos die within 3-5 days after infection.
The virus is identified in RN, RDP and ELISA.
DIFFERENTIAL DIAGNOSIS- exclude coccidiosis, poisoning, infectious bronchitis, hemorrhagic syndrome, mycoses, Newcastle disease.
TREATMENT- not developed.
IMMUNITY- use live and inactivated vaccines of the BG strain (Gumboro disease), IBD (infectious bursal disease), Winterfield-2512.
The first vaccine is administered twice at the age of 7-21 days with an interval of 10-14 days using the drinking method. Second time at the age of 110-120 days
once intramuscularly into the pectoral muscle area or into the thigh in a volume of 0.5 ml. Immunity occurs 14-21 days after vaccination and lasts up to a year.
In foreign practice, a vaccine made from a weakened strain of the infectious bursitis virus is used with drinking water and aerosolized. Among foreign vaccines, you can use Nobilis Gumboro D78 and 228E. An inactivated vaccine, Nobilis Gumboro inc., has also been developed.
PREVENTION AND CONTROL MEASURES- carry out general veterinary and sanitary measures to prevent the introduction of the pathogen into the farm.
The young animals of each technological batch are raised in isolation. The state of poultry resistance is controlled through targeted feeding and maintenance.
The air entering the poultry house is cleaned with filters and disinfected with ultraviolet rays.
When infectious bursitis appears, restrictions are introduced. Sick and suspicious birds are destroyed. Healthy people are vaccinated.
The premises are thoroughly disinfected with solutions of caustic soda, bleach (2-3%), and an aerosol of iodide preparations.
If the disease cannot be controlled by general veterinary and sanitary measures, the farm stops incubating eggs and carries out additional health measures.
There are no deadlines for lifting the restrictions; they are set by veterinarians, since it is difficult to get rid of this disease due to the rapid development of this disease as a permanent one.
Gumboro disease poses a serious threat to industrial poultry farming, causing high mortality and loss of productivity in both meat and egg chickens. The use of intermediate plus vaccines protects chickens from acute and subclinical forms of this disease. To study chicken blood sera for the presence of antibodies to the Gumboro disease virus, the ELISA method has been used for more than 15 years, which has a number of advantages, for example, it is fast and cheap, but its main advantage is that the method is reliable and standardized. This article presents the results of a study of broiler blood serum by ELISA during the growing period (from the 1st to the 45th day).
Protection throughout the growing period
One of the main objectives of vaccination against Gumboro disease is to protect chicks throughout the rearing period. Day-old chicks are protected from MA disease, which they received from chickens of the parent flock. Broiler chickens become sensitive to the Gumboro disease virus by 20 days of age, and egg-laying chickens by 30 days of age. The presence of MA in chickens at an early age prevents their vaccination, since MA neutralize the vaccine virus (Fig. 1).
It has been proven that when immunizing chickens with a high level of MA, the vaccine virus does not cause seroconversion. When vaccinating chickens with critical level MA (the level of MA that the vaccine strain can overcome) a good immune response is formed, preventing clinical and subclinical signs of Gumboro disease - mortality, hemorrhagic inflammation of the bursa, decreased productivity.
In chickens in the early period (Fig. 2), the average antibody titer decreases, which reaches its critical value in chickens at 12 days of age (559). The chickens were vaccinated the next day. During a serological study, the first antibodies were detected on the 4th day after vaccination. The average antibody titer reached its maximum value at 26 days of age (3338) and remained at a high level until the slaughter of the bird (3398). A field virus can also cause seroconversion, but in this case, along with the latter, clinical signs appear and chicken productivity decreases. When assessing antibody titers (Fig. 3), it is also very important to take into account their homogeneity. High uniformity indicates good quality of chicken vaccination.
Determining the date of vaccination
An equally important task of vaccinating chickens is to create a smooth transition from passive immunity (PA) to active immunity. However, in real conditions It is only possible to calculate the exact date of vaccination if the chicks come from the same supplier and from a homogeneous parent flock. Blood serum for serological testing is usually taken from chickens during the first three days of life.
Vaccination time is calculated using the Kohavena formula (for broiler chickens) or Deventor (calculation based on the decay period of antibodies).
These methods are often used in laboratories because they are included with the equipment software. The date of vaccination determined in this way must be confirmed by a veterinary specialist or manager, in accordance with the actual situation in the factory.
In this experiment, chickens were immunized with the Cevac® IBD L vaccine by feeding at 13 days of age (Ceva Santé Animal, Libourne, France). The average antibody titer in chickens before vaccination at 12 days of age was 559, with a larger number of samples (8) in the first group with antibody titers from 0 to 396 (Fig. 4). Coefficient of variation 97%. Chickens were immunized once with the intermediate plus vaccine Cevac® IBD L, since it overcomes the MA level at a dilution of 1:500.
Seroconversion and duration of antibody circulation
When studying blood serum samples from chickens at 26 days of age (2 weeks after vaccination), a high and uniform seroconversion was noted (Fig. 5), with a coefficient of variation of 32%. A positive titer of antibodies against Gumboro disease is considered to be from 1000 to 2000. In this study the average antibody titer was 3338 and all samples were positive. The results of an ELISA study depend on many conditions - type of kit, operator skills, external conditions for setting up the reaction (temperature, humidity). It is possible to compare research results only if they were performed simultaneously, by the same operator and using kits from the same manufacturer.
The homogeneity of antibody titers can be assessed by the coefficient of variation (see Fig. 5). The diagram shows blood serum samples, which are distributed into groups: from 0 to 395, from 396 to 999, from 1000 to 1999, etc. It is generally accepted that for statistically reliable results when performing ELISA, it is necessary to select from 18 to 23 (18 minimum) blood serum samples. Information in the form of a diagram allows you to evaluate the arrangement of titers by group (most samples in one group or samples distributed in several groups) and in relation to the average value (heterogeneous or homogeneous titers). When studying the blood sera of day-old chicks (see Fig. 3), most antibody titers were located in one group and were close to the average value.
Good uniformity and a high level of MA indicate a satisfactory state of health of the parent stock and the right program vaccination of chickens before laying using live and inactivated vaccines.
Many studies have noted a positive correlation between the virus neutralization reaction and the ELISA method. In addition, differences in mean titers between two groups of chickens vaccinated with different vaccines do not always indicate the immunogenicity of the vaccine. For example, the average antibody titer of 3000 in 25-day-old chickens immunized with vaccine A and 4500 in chickens vaccinated with vaccine B does not mean that immunization is better when using vaccine B. We can only say that chickens of both groups were successfully immunized. Differences in mean titer may be due to differences in kits or other conditions. The average titer value can be used to assess the quality of vaccination (bad or good), but not to compare different vaccines. With ELISA the most essential criterion To evaluate the results, the homogeneity of titers is used, which depends mainly on the quality of the vaccine, the conditions of its transportation and storage, and the technique of vaccinating chickens. To check the quality of vaccination, you can carry out additional research blood serum samples from 45-day-old chickens. In these studies, the average antibody titer in 45-day-old chickens was 3398 (Fig. 6) with a coefficient of variation of 23% (high homogeneity).
