Modification of active immunization with live bovine herpesvirus 1 vaccine by passive viral antibody.

The study in BHV-1 naïve calves investigated the effect of intramuscularly (IM) administered BHV-1 neutralising bovine immunoglobulin on the efficacy of a live intranasally (IN) administered BHV-1 vaccine. Overall on daily basis there was between 40- and 5000-fold less vaccine virus shed by the passively immune calves compared with that shed by the naïve counterparts. The latter seroconverted to the vaccine whereas the virus neutralising (VN) antibody titres in the passively immune calves decreased after vaccination. Compared with unvaccinated naïve or passively immune calves, both groups of vaccinated calves shed significantly less challenge BHV-1 but the daily amount shed was significantly lower in vaccinated naïve calves. The latter were also significantly better protected against pyrexia following the IN BHV-1 challenge compared with vaccinated passively immune calves. Unlike vaccinated calves, clinical reaction to challenge in both unvaccinated groups also involved nasal discharge but the duration of both the nasal discharge and the severe pyrexia was significantly shorter in unvaccinated passively immune calves. Conclusions from the study are: (1) the circulating VN antibody is significantly protective against virus shedding and to alesser extent also against the febrile respiratory disease; (2) the passively immune calves are unlikely to seroconvert after an active infection and (3) the passive antibody does negatively affect vaccine efficacy.

Prevention of transplacental infection of bovine foetus by bovine viral diarrhoea virus through vaccination.

Results are presented for an experimental validation of the efficacy of an EU licensed, inactivated bovine viral diarrhoea virus vaccine (Bovilis BVDV). This study was designed to assess the quality of efficacy 6 months after a single course of vaccination (two intramuscular doses a month apart). A natural challenge at about 87 days of gestation by 3 persistently infected carrier heifers rapidly infected all experimental heifers. This resulted in transplacental BVDV infection of all 7 unvaccinated dams whereas 11 immunised dams produced 9 live-born calves and 2 aborted foetuses from which no BVDV could be recovered.

Isolation of highly cytolytic MDV strains from Germany and Spain.

Four serotype 1 Marek's disease virus (MDV) strains were isolated from Marek's disease (MD) outbreaks in Germany and Spain and their virulence was tested in SPF White Leghorn chickens. Two of the isolates from Germany (MR48/1-2 and /2-8) induced early mortality from cytolytic MD in 25% of the chickens whereas one isolate from Germany (MR48/3-16) caused early mortality in 58% of the inoculated chickens. Histological examination showed severe depletion of the lymphoid tissue of the bursa of Fabricius and the thymus, consistent with the cytolytic phase of MD. AH surviving chickens later developed acute MD in the form of multiple visceral tumours. Inoculation of SPF chickens with various doses of the isolate MR36 8/1C (Spain) caused early mortality in up to 69% of the chickens and acute MD in 92 to 100% of the chickens surviving the cytolytic phase of the disease.

Detection of lymphoproliferative disease virus by an enzyme-linked immunosorbent assay.

Hitherto, detection of lymphoproliferative disease virus (LPDV), a C-type retrovirus of turkeys, has proved difficult since no tissue culture or serological assay has been available. Development of serological tests has been hampered by the problems of raising virus-specific antisera. An indirect enzyme-linked immunosorbent assay (ELISA) is reported, using a viral antiserum raised with bromelain-digested virus. The assay specifically detected purified virus at a concentration of 250 ng/ml or greater. In an experiment to detect virus in plasma from turkeys over a period of 4 weeks following LPDV infection, ELISA results correlated closely with the viral reverse transcriptase activity. Both assays were of similar sensitivity and detected small amounts of virus in high-speed pellets of turkey plasma. Evidence is presented indicating that LPDV-infected or hyperimmunized turkeys do not produce readily detectable circulating viral antibodies. In reciprocal ELISA tests, using antibodies to group-specific antigens of other avian retrovirus groups (avian sarcoma-leukosis (ASLV) and reticuloendotheliosis (REV] no antigenic cross-reaction was found between LPDV, ASLV and REV.

Diagnosis of lymphoproliferative disease virus infection of turkeys by an indirect immunofluorescence test.

An indirect immunofluorescence (IIF) test for the detection of lympho-proliferative disease virus (LPDV) is described. Results presented show that smears of buffy coat cells and frozen sections of spleen, but not thymus, form suitable test materials for a rapid and specific diagnosis of LPDV infection of turkeys. Between 0.1 and 0.6% of buffy coat cells from infected birds were antigen positive, and spleens from 18/18 and thymuses from 11/18 birds respectively, tested 16 months after viral inoculation, were positive. Antisera containing group-specific antibodies to avian reticuloendotheliosis virus (REV) and avian sarcoma-leukosis virus (ASLV) did not cross-react with LPDV-infected buffy coat cells or spleens.

Production of virus-specific antisera to lymphoproliferative disease virus of turkeys.

Lymphoproliferative disease virus, a recently isolated C-type retro-virus, cannot yet be cultured in cells in vitro and rarely induces a high titre virus-specific antiserum. However, after digestion of purified virus with bromelain and rebanding the treated virus in sucrose gradients bald virions devoid of the viral spike glycoprotein and up to 20 host proteins were found to induce a high level virus-specific antibody response in rabbits and chickens.

Characterisation of lymphoproliferative disease virus of turkeys. Structural polypeptides of the C-type particles.

Lymphoproliferative disease virus of turkeys (LPDV), a C-type retrovirus, was shown to contain 3 major [32 kilodaltons (kd, p 32), 26 kd, 22/21 kd] and 2 minor (41 kd and 12 kd) polypeptides. Preliminary evidence suggests a glycoprotein of 76 kd (GP 76) and a major doublet polypeptide of 13.5/13 kd to be also of viral origin. Of these GP 76 was susceptible to bromelain action implying its surface location in the virion, while p 32, p 26 and p 13.5/13 were the main constituents of viral cores. p 13.5/13 bound an RNA probe, suggesting it to be the main constituent of viral ribonucleoprotein. p 22/21 was not cleaved by bromelain, and was absent in viral cores suggesting its intramembrane location between virion envelope and core. The polypeptide profile of LPDV is distinct from those of avian sarcoma-leukosis viruses and avian reticuloendotheliosis viruses.

Further studies on vertical transmission of reticuloendotheliosis virus in turkeys.

Turkey hens were inseminated with infected semen from REV tolerant turkey stags at weekly intervals for 6 weeks, and subsequently with semen from uninfected control stags for a further 6 weeks. Cell cultures prepared from eggs produced during the experimental period were assayed for REV by an indirect fluorescent antibody test. Poults hatched from such eggs were also assayed for REV at 1 day of age or maintained to 28 weeks of age to study lateral spread of virus to uninfected hatch mates. REV was detected in 27.5% of embryos from hens inseminated with REV infected semen and 8.7% of 1-day-old poults hatched were found to be viraemic. Following the change to insemination with uninfected semen, REV was detected in 0.7% of embryos examined. REV was isolated from the lower oviduct of two of 18 hens, 12 weeks after first insemination with infected semen. REV spread readily to uninfected poults maintained in contact with progeny of hens inseminated with REV-infected semen, in that 50% had neutralising antibody by 8 weeks of age. At 28 weeks of age the leukosis mortality in such poults was 16%.

A quantitative sequential study of viraemia and neutralising antibody to HVT and MDV in a commercial flock vaccinated with HVT.

Levels of viraemia due to the herpesvirus of turkeys (HVT) and neutralising antibodies to HVT and Marek's disease virus (MDV) were followed in a flock of commercial broiler breeders held in commercial premises and vaccinated at 1-day-old with HVT. The flock was sampled at 3, 5 and 8 weeks of age and then at 4-weekly intervals until 32 weeks of age. The mean viraemia titre was highest at 3 and 5 weeks and then fell to a low level by 16 weeks where it remained until 32 weeks of age. At all sampling periods there was great variability in individual viraemia titres and a significant proportion with no detectable viraemia. Neutralising antibody titres to both HVT and MDV were highest at 1 day old. The geometric median titres fell rapidly and only started to rise slowly between 8 and 12 weeks of age. The titre of individuals within a sample varied greatly and from 16 weeks of age between 3% and 43% had no neutralising antibody to MDV. An examination of the factors affecting the accuracy of the assay for HVT viraemia indicated that the bird effect was greatest suggesting the variability in viraemia was not due to errors in the assay. Results similar to the field study were obtained when the same strain of chicken was vaccinated under laboratory conditions indicating that faulty vaccination was not responsible for the variable response to vaccination. There was a positive correlation between HVT viraemia and neutralising antibody to HVT and MDV. Groups of 1-day-old progeny of HVT vaccinated parents and grandparents of three laying strains and one broiler strain derived from a number of farms were examined for levels of neutralising antibody to HVT. Both the geometric mean titres of each group and the titres of individuals within a group varied greatly. The strain of chicken had no effect on the neutralising antibody response to vaccination with HVT and probable field exposure to MDV but the farm of origin affected both the geometric mean titre and the variability in titre of neutralising antibody in individuals. It is suggested that the exceptionally high levels of neutralising antibody to HVT noted in a proportion of the 1-day-old chicks of the field study could be responsible for a poor response of some individuals to vaccination with HVT.

Experimental infection and vertical transmission of reticuloendotheliosis virus in the turkey.

Groups of turkey poults were infected with reticuloendotheliosis virus (REV) at 1 day of age, 4 weeks of age and by contact exposure from 1 day of age. Mortality from reticuloendotheliosis occurred in all three groups, but was at a lower level and later in life in turkeys inoculated at 4 weeks of age. Using a plaque neutralisation test, antibody to REV was detected in birds inoculated when 4 weeks old and in contact exposed birds, but not in turkeys inoculated at 1 day of age. Neutralising antibody persisted to 40 weeks of age and was present in individual birds which subsequently died from reticuloendotheliosis. REV viraemia was present in poults inoculated at 1 day of age and persisted throughout the 40 weeks of the experiment. A transient viraemia was also detected in individual contact exposed birds, but was not present in tests on turkeys inoculated at 4 weeks of age. Turkey hens from each group were brought into production and inseminated with turkey semen from uninfected male turkeys. REV was isolated from two of 20 eggs produced by viraemic turkey hens, but not from 92 eggs cultured from contact exposed hens or from 27 eggs from hens inoculated at 4 weeks of age.

A leukosis in turkeys associated with infection with reticuloendotheliosis virus.

The clinical features and lesions are described for an outbreak of a leukosis in an integrated turkey breeding organisation. Turkeys had diarrhoea between 8 and 12 weeks of age and from 15 weeks onwards leukosis lesions appeared. A greater than 20% mortality occurred in most affected flocks. The disease was characterised clinically by enlargement of the liver and to a lesser degree other organs. Microscopically the lesions consisted of proliferations of lymphoblastoid cells. Reticuloendotheliosis virus (REV) was isolated from ailing culled turkeys and from cell cultures prepared from embryonated eggs produced by a flock with this disease. In affected flocks a REV was also isolated from turkeys 8 to 12 weeks of age in which enteritis was the main clinical feature. Inoculation of 1-day-old turkeys with this isolate of REV produced a syndrome of enteritis followed by leukosis. Antibody to REV was detected in turkeys surviving experimental inoculation and in the two flocks of turkeys examined, one of which had experienced considerable leukosis mortality.

Lymphoproliferative disease of turkeys. II. Experimental transmission and aetiology.

The clinical and pathological features of a previously reported lympho-proliferative disease of turkeys were reproduced by inoculation of 4-week-old poults, and spread to in contact turkeys was demonstrated. From studies on the development of the lesions in inoculated turkeys, it was possible to assay the causative agent based on microscopic examination of early lesions. Poults were more susceptible to inoculation when 4 weeks old than to inoculation at 1 day old, and different strains of turkey varied in their susceptibility to inoculation. The causative agent did not appear to be antigenically related to avian leukosis virus, to Marek's disease virus or to reticuloendotheliosis virus. Preliminary characterisation studies indicated a virus with properties similar to those of oncoviruses.