Occurrence of atypical myxomatosis in Central Europe: clinical and virological examinations.

An outbreak of the atypical form of myxomatosis struck a rabbit farm in Hungary. The animals had previously been vaccinated with a vaccine containing Shope rabbit fibroma virus strain. The disease appeared in winter when the presence of mosquitoes and fleas is not common. The virus was isolated from an eyelid specimen of a naturally infected rabbit. The surviving animals were observed for four weeks, blood samples were collected and, after euthanasia, organ specimens were also examined by morphological methods including pathology and electron microscopy. Serum samples were examined by virus neutralisation for antibodies. Genetic analysis of the isolated virus was carried out by polymerase chain reaction (PCR) and direct sequencing. The primers were designed on the basis of the major envelope gene (Env) of the Lausanne reference strain in the GenBank. The viral proteins were examined by SDS-PAGE. The isolated virus (ref. no.: BP04/2001) was able to infect the susceptible animals directly, by contact. The disease was characterised by respiratory symptoms of the upper tracheal tract, conjunctivitis and high mortality by the 11th-14th day. Aerogenic infection with strain BP04/2001 resulted in 100% morbidity among the susceptible animals. Sequencing of the amplified 400-bp-long DNA revealed 97% homology with the Env gene of the Lausanne strain, which proves that strain BP04/2001 is a variant of the Lausanne strain having been enzootic throughout Europe. The live vaccine strain used in Hungary against myxomatosis, which is also a Lausanne-derived strain, protected the animals. According to the protein analysis a protein of 200 kDa in size is not expressed in strain BP04/2001. This is the first report on atypical myxomatosis in Central Europe. The virus spreads by airborne transmission and may cause severe losses in the rabbit population.

Pathogenicity of a bovine viral diarrhoea virus strain in pregnant sows: short communication.

The biological properties of bovine viral diarrhoea virus (BVDV) strain Oregon C24V were studied after intranasal and subcutaneous infection of pregnant sows. This virus strain is widely used in Hungary for immunising cattle against bovine viral diarrhoea (BVD). Based upon the results of the clinical, gross pathological, histopathological and virological examinations it can be established that the given strain caused asymptomatic infection and serological conversion in sows that were in the second third of gestation. The virus caused clinically apparent disease in some of the piglets born at term, which indicates that it had crossed the placenta. More than half (57%) of the live-born piglets died within 60 days of birth. The sows and their progeny did not shed the virus. BVDV infection has great differential diagnostic importance in pigs, as classical swine fever (CSF) virus strains of reduced virulence cause similar clinical symptoms and gross and histopathological changes.

Pathogenesis of primary respiratory disease induced by isolates from a new genetic cluster of bovine viral diarrhea virus type I.

The pathogenesis of infection induced by cytopathogenic isolates from the newly identified genetic cluster Id of bovine viral diarrhea virus (BVDV) type I was studied in two experimental infections of previously seronegative, immunocompetent calves. Experiment 1 focused on the evaluation of clinical patterns, viremia, and serological responses. All infected calves in this experiment developed respiratory symptoms and seroconverted to BVDV positivity. Contact calves also contracted a respiratory tract infection following exposure to infected animals. Viremia was demonstrated between postinfection days 2 and 17, and the virus was detected in organ specimens of all but one each of the infected and contact calves. In experiment 2, the distribution of BVDV in various tissues of calves euthanized at defined days postinfection was studied. In two of these calves recurrent shedding of BVDV in nasal secretions was shown. BVDV was detected in various tissues of all infected calves throughout the experiment and also following seroconversion and the clearance of BVDV from the circulatory system. Despite the widespread distribution of the virus in various organs, significant tissue damage was found mainly in respiratory tract and lymphoid tissues. These experiments revealed that viruses from cluster Id of BVDV are able to induce primary respiratory disease in previously seronegative, immunocompetent calves. Contact transmission and virus recurrence, contrary to observations from acute experimental infections with noncytopathogenic BVDV, are likely to reflect differences in biological features of these cytopathogenic isolates. Virus shedding and its presence in tissues following peripheral clearance and in the presence of antibodies may have implications in the diagnosis, pathogenesis, and epidemiology of BVDV-induced syndromes in cattle.

Gene immunization of mice with plasmid DNA expressing rabies virus glycoprotein.

Gene immunization can be an effective vaccine strategy eliciting both humoral and cell-mediated immune responses. We constructed plasmid vectors expressing the full-length Vnukovo-32 rabies virus glycoprotein G under the control of CMV IE promoter and enhancer, adenovirus tripartite leader sequences and poly A signal of SV40. The gene vaccines were evaluated for the ability to elicit neutralizing antibodies and to protect BALB/c mice against lethal rabies virus challenge. First, mice were injected intramuscularly (i.m.) into the left hind leg and by the intradermoplantar (i.d.p.) route with equal amounts of plasmid DNA (0.25-0.1 mg). Two weeks later, immunization was boosted with an additional dose of the DNA. The immunized mice were challenged by intracerebral (i.c.) inoculation of CVS-27 (10-50 LD50) rabies virus. All mice produced anti-rabies virus neutralizing antibodies with a titre of > or = 1:45 after immunization with 0.1-0.4 mg of DNA. In challenge experiments, 83 to 91.6% protection was observed. These results confirm that a DNA vaccine could be a simple and effective solution for preventing the spread of rabies.

Studies on the pathogenicity of bovine herpesvirus type 5 in sheep.

Four Merino lambs were intranasally inoculated with bovine herpesvirus type 5 (BHV-5) reference strain N569. Two lambs were mock-inoculated as negative controls. The virus-inoculated animals developed apathy, inappetence, rhinitis, nasal, ocular and genital discharge, slight diarrhea and neurological disorders, like tremor and salivation. BHV-5 was isolated from the nasal discharge in two of the animals, while the polymerase chain reaction (PCR) detected the virus in all the infected lambs. Two lambs died on post infection day (PID) 13, while the other two infected animals were euthanized on PID 15 and 30. Gross pathological changes were not observed, however, histopathological examinations revealed diffuse nonsuppurative meningo-encephalitis in all infected animals. Viral antigen was detected by immunohistochemistry and viral nucleic acid was revealed by in situ hybridization in the brain of the two lambs, which died on PID 13. The virus was demonstrated by virus isolation and by PCR from different organs of all the infected animals. Slight rise of antibodies was observed in the infected animals from PID 15. The results show that BHV-5 is able to cross the species barrier and may establish infection in sheep.

Experimental infection of weaned calves and lambs with two strains of bovine adenovirus type 2.

The pathogenicity of two genetically different strains of bovine adenovirus type 2 (BAV-2) was tested in a double cross experimental infection of weaned lambs and calves. One of the virus strains was isolated from cattle and represented BAV-2 subgroup "A", while the other was isolated from sheep and belonged to subgroup "B" (Belák et al., 1983). The two virus strains caused infection both in lambs and calves; the viruses were repeatedly reisolated and definite seroconversion was found in all four groups. However, remarkable differences were observed in the pathogenicity: the most intensive virus multiplication and shedding, the most severe clinical symptoms and histopathological lesions were observed in the lamb group infected with the ovine isolate.

ISCOM of BHV-1 envelope glycoproteins protected calves against both disease and infection.

A subunit vaccine in the form of immunostimulating complex (iscom) was prepared to contain the envelope glycoproteins of bovine herpesvirus type 1 (BHV-1). This iscom preparation was tested in a vaccination experiment on 4-month-old calves seronegative to BHV-1. In this experiment, four groups with three animals per group were used. Two groups were vaccinated with the iscom preparation twice, four weeks apart, one group with 50 micrograms and the other with 100 micrograms per calf. The third group received a commercial inactivated whole-virus vaccine applying the same vaccination program. The fourth group served as control. Two weeks after the second vaccination, all the animals were challenge-infected intranasally with a virulent BHV-1 strain and four days later with a virulent Pasteurella multocida--this in order to mimic hard field conditions. When exposed to challenge infection, all the animals vaccinated with the iscom were fully protected, i.e., no virus could be recovered from their nasal secretions and no clinical symptoms were recorded. In contrast, the animals vaccinated with the commercial vaccine, responded to challenge with moderate fever and loss of appetite, and virus was isolated from the nasal secretions. The animals in the control group developed severe clinical symptoms. In the sera of iscom-vaccinated animals, the virus neutralization titers reached levels of 1/3500 or higher.

DNA restriction enzyme analysis of a bovine herpesvirus 1 strain isolated from encephalitis in Hungary.

The DNA of a bovine herpesvirus 1 (BHV-1) strain isolated from calf encephalitis in Hungary was analysed with restriction enzymes. The cleavage pattern of the encephalitis strain Na/67 differed from those of all the other Hungarian BHV-1 isolates investigated so far. The EcoRI and HindIII cleavage patterns of virus strain Na/67 were found to be similar to the patterns of two other encephalitis strains (N569 and A663 from Australia and Argentina, respectively) characterized earlier. Strain Na/67 is the first isolate in Europe which showed the restriction enzyme pattern of BHV-1.3 previously supposed to be characteristic of encephalitis strains.