Isolation and study of biological properties of non-oncogenic Marek's disease herpesviruses in chickens. 2. Characterization in vivo]

In a previous paper (Jurajda and Halouzka, 1992) the in vitro isolation of two chicken herpesviruses of Marek's disease (M and K strains) was described and results of their characterization were presented. The present paper deals with the in vivo characterization of both isolates: pathogenicity and immunosuppressive characteristics of isolates were observed in a five-week test period, along with the development and production dynamics of antibodies and viral antigen in the feathers of experimentally infected chickens of the Brown Leghorn breed. A technique of double immunodiffusion in agar gel according to Ouchterlony, modified by Woernl (1966), was used to determine the presence of antibodies to Marek's disease virus (MDV) in blood serum and of precipitating MDV-antigen in feather quills of tested chickens. Isolate multiplication and titration were performed in a system of chicken embryonal fibroblasts (CEF) (Jurajda et al., 1984). Chickens were infected i.m. with three virus doses - 10(2) to 10(4) PFU per chicken while the dose 10(4) corresponded to the titre of 10,800 PFU/0.2 ml for M isolate and to the titre of 8,600 PFU/0.2 ml for K isolate. The nature and rate of regressive changes in lymphatic organs were determined according to criteria described by Halouzka and Jurajda (1991). The results are summarized in Tabs. I and II. Neither of the isolates evoked clinical or pathomorphological macroscopic symptoms of the disease. M isolate induced microscopic MD-specific changes in the peripheral nerves (of C type) and only moderate and transient signs of immunosuppression in 11% of infected chickens.(ABSTRACT TRUNCATED AT 250 WORDS)     

Cyclophosphamide-induced amelioration of Marek's disease in Marek's disease-susceptible chickens.

Bursa- and thymus-dependent functions were examined in Marek's disease (MD)-susceptible normal chickens and in chickens treated with 5 and 16 mg of cyclophosphamide (CY) at the time of hatching. Chickens not exposed to Marek's disease virus (MDV) and treated with CY temporarily lost mitogenic response to concanavalin A but regained full response after 5 weeks. Bursa-dependent functions, such as presence of germinal centers in spleen and cecal tonsils, morphologic features of bursa, and sheep red blood cell antibody response were completely lost in chickens treated with 16 mg of CY and only partly retained in chickens treated with 5 mg of CY. In chickens exposed to MDV, the degree of thymus-dependent spleen cell mitogenic response was directly related to frequency and severity of MD. Chickens treated with 16 mg of CY had a mild mitogenic depression and low frequency and severity of MD lesions, whereas those treated with 5 mg of CY and those not treated had marked mitogenic depression and high frequency and severity of MD. Suppressions of bursa- and thymus-dependent functions by MDV alone were also evident when comparing MDV-exposed and nonexposed chickens. The results also indicate that presence of small, residual amounts of humoral factor(s) may enhance MDV oncogenesis.     

Immunological aspects of Marek's disease in chickens with maternal antibodies]

The dynamics of the production of immunoprecipitation antibodies to Marek's disease virus was studied in the serum of chickens with maternal antibodies in relation to the occurrence of the immunoprecipitation antigens of Marek's disease virus in feather follicles. One-day-old chickens were infected by the contact method with Marek's disease virus. The first occurrence of immunoprecipitation antigen was detected on the 14th day after infection and this occurrence persisted throughout the experiment, i. e. until the 112th day after infection. The antibodies were first detected the 28th day after infection and their titre kept rising until the 98th day after infection. Immunoprecipitation antibodies and antigens of Marek's disease virus were detected in some tumorously changed kidneys. Immunoelectrophoretic examination revealed in the same kidneys immunoglobulins of the class IgY, IgA and beta-globulin. The slowest-migrating fraction of IgY, together with IgA, beta-globulin and C-reactive protein were detected in the skin extracts from infected poultry. Indirect haemagglutination enabled the detection of the presence of haemagglutination antibodies in rabbit immunoglobulin to the skin antigen of Marek's disease virus, and in avian immunoglobulin to the same virus. Haemagglutination antigen was revealed in the extract from tumorously changed kidneys.     

Pathogenesis of Marek's disease in chickens with maternal antibody.

Histopathological lesions were studied in chickens with maternal antibody against MD virus at 14-day intervals after their exposure by contact to HPRS-16 of MD virus in the first-day of life. The first lesions occured 14 days after infection in the liver, kidney, heart, brain and the sciatic nerves. Relatively, the most expressive changes were observed between 56th and 70th day after infection. Lesions were characterized as light to heavy infiltrations formed by immature and mature pleomorphic mononuclear cells. Among these cells myeloid type cells occured too. In some cases expressive tumorous lesions with intensive mitosis were observed mainly in the liver. Intranuclear inclusion bodies were also observed mainly among the proliferating epithelial cells of the proventriculus. 14-days after infection changes in the brain were classified as nonpurulenta encephalitis. Gross lesions characteristic of MD occured the most frequently in the gonads, liver and the kidney. In one case 56 days after infection herpesvirus-like particles were observed by electron microscopy in the tumorous gonads and liver. They measured about 200 to 260 nm in diameter. Degraded and/or aberrant incompleted virus-like particles occured the most often and completed (enveloped) ones with electrondense nucleoids were observed occasionally. It was concluded that MD in chickens with materanal antibody against MD virus shows monophasis progress.     

Marek's disease in field chickens: correlation between incidence of Marek's disease and nuclear-inclusion formation in the feather-follicle epithelium

The present study confirmed that Marek's disease (MD)-associated nuclear-inclusion (NI) formation in the feather-follicle epithelium (FFE) is related to mortality from MD; it also presented useful data on the epidemiology of MD in HVT-vaccinated field chickens. Incidence of NI formation in the FFE of chickens on six rearing farms varied greatly by age and flock, but most of the field flocks showed biphasic peaks of incidence of NI in chickens consisting of a small peak at an early age (usually at 2-4 weeks of age) and a large peak between 13 and 16 weeks of age. MD tended to occur in chickens over 20 weeks old, and almost all MD-affected chickens showed NI formation persistently in the FFE, usually between 13 and 20 weeks of age. Chickens that were healthy at the end of observation showed either transient NI formation, usually between 13 and 16 weeks of age, or no detectable NI formation. Incidence of NI formation in the FFE of chickens at 19-20 weeks of age was related to mortality from MD: chickens with NI formation had a MD mortality rate of 66.7%, whereas chickens without NI formation had MD mortality of only 0.8%.     

The effect of cyadox and virginiamycin on Marek's disease in chickens

Chickens infected with highly virulent Marek's disease virus were administered in the course of 56-day fattening cyadox at a dose of 20 mg per kg feed mixture (MC group). and virginiamycin at a dose of 10 mg per kg feed mixture (MV group). The weight gains on the 56th day and dressing percentage were highest, in comparison with the control group (711.11 g and 60.62%), in the virginiamycin-treated group. An investigation into the basic chemical composition of meat revealed lower contents of dry matter and proteins in the virginiamycin-treated group (26.9% and 22.76%) if compared with the control group of chickens which were also infected with Marek's disease virus. A biometrical examination of the organs showed a statistically significant decrease in heart weight in the cyadox-treated group, increase in liver and heart weight and intestine shortening in the virginiamycin-treated group, in comparison with the control group. Pathologico-anatomical changes typical of Marek's disease were recorded in a randomly selected number of chickens in 33.3% in the control group, in 28.5% in the cyadox-treated group and in 50.0% in the virginiamycin-treated group.