Dexamethasone-induced recrudescence of malignant catarrhal fever and associated lymphosarcoma and granulomatous disease in a Formosan sika deer (Cervus nippon taiouanus)

Malignant catarrhal fever (MCF) was diagnosed in a 2-week-old Formosan sika deer. The fawn had been previously exposed to a clinically normal neonatal wildebeest calf from which alcelaphine herpesvirus-1 was isolated. Alcelaphine herpesvirus-1 was isolated from buffy coat leukocytes and nasal and ocular secretions of the fawn during the acute illness. The fawn clinically recovered after 3 weeks. Virus was not recovered from blood at this time. Dexamethasone, given 4 months after clinical recovery, resulted in reisolation of MCF virus from blood and recrudescence of clinical MCF. The deer was euthanatized. At necropsy, pathognomonic lesions of MCF, granulomatous disease, and malignant lymphoma were observed. Antibodies to bovine leukosis viral antigens were not detected in the serum. The epidemiologic and pathogenetic importance of the findings are discussed.     

Immunofluorescence of bovine virus diarrhea viral antigen in white blood cells from experimentally infected immunocompetent calves.

A study to evaluate the detection of bovine virus diarrhea viral antigen using immunofluorescence testing of white blood cells was conducted. Five colostrum-deprived calves were inoculated intravenously with a cytopathic strain of the virus. Lymphocyte and buffy coat smears were prepared daily for direct immunofluorescent staining for detection of antigen. Lymphocytes were separated from heparinized blood using a Ficoll density procedure. Buffy coat smears were prepared from centrifuged blood samples collected using ethylenediaminetetraacetic acid as an anticoagulant. Bovine viral diarrhea virus antigen was detected by immunofluorescence between 3 and 11 days postinfection in lymphocyte smears and 3 to 12 days postinfection in buffy coat smears. Isolation of virus from both lymphocytes and buffy coat preparations correlated with detection of immunofluorescence. Serum neutralizing antibody to bovine virus diarrhea virus was detected on day 10 postinfection. Buffy coat smears were as sensitive as lymphocyte smears for the detection of antigen by immunofluorescence. It appeared that immunofluorescent staining of white blood cells was an effective method of detecting bovine virus diarrhea viral antigen.     

Evidence that the sheep associated form of malignant catarrhal fever is caused by a herpes virus

Out of a cow, which was infected with the sheep form of malignant catarrhal fever (MCF), blood and spleen samples were inoculated into rabbits. From the spleen cells of an infected rabbit, which were cocultivated with bovine embryonic gingiva cells, a herpesvirus could be isolated. The isolate showed crossreactions with reference sera against the strain WC 11 (wildebeest form) in the SNT. An immunosuppressed heifer, which was infected with the isolate, contracted typical clinical symptoms of MCF. The isolate was named No. 732.     

Diagnostic investigation of bovine viral diarrhea infection in a Minnesota dairy herd

Bovine viral diarrhea (BVD) virus infection was diagnosed in neonatal calves with enteritis. Successful diagnostic procedures included direct immunofluorescence of frozen tissue sections, histopathology, and virus isolation. Virus isolation from buffy coats and serum was successful in detecting infected animals, whereas direct immunofluorescence of buffy coat samples was found to be less reliable. Virus was not isolated from any fecal samples. Booster vaccinations and the culling of animals shedding virus resulted in improved calf viability in this herd. It is suggested that procedures for the diagnosis of BVD virus infection should always be included in the diagnosis of neonatal calf enteritis.     

A serologic survey of viral infections in captive ungulates in Turkish zoos

Zoos and zoologic gardens make optimal environments for interspecies transmission of viral infections. There are seven zoos and several small zoologic collections in Turkey. This study aimed to determine the current status of viral infections in captive ungulates living in these environments. Blood samples were taken from 163 captive animals from two zoos. There were 39 Cameroon sheep (Ovis ammon f aries), 11 Barbary sheep (Ammotragus lervia), 57 pygmy goats (Capra hircus), 9 Angora goats (Capra hircus), 21 mountain goats (Capra aegagrus-aegagrus), 7 llamas (Lama glama), 8 Persian goitred gazelle (Gazella subgutturosa subgutturosa), 7 Caspian red deer (Cervus elaphus maral), 2 fallow deer (Dama dama), and 2 camels (Camelus dromedarius). Antibodies against bovine viral diarrhea virus (BVDV), bovine herpesvirus-1 (BHV-1), bovine adenoviruses (BAV-1 and -3), parainfluenzavirus 3 (PI-3), and bluetongue viruses (BTV-4 and -9) were investigated using the virus neutralization test, and malignant catarrhal fever (MCF) antibodies were screened by ELISA. All animals were negative for BVDV and BHV-1 antibodies. Seroprevalence of BAV-1, BAV-3, PI-3, BRSV, BT-4, BT-9, and MCF were detected as follows: 46.6%, 60.1%, 0.6%, 7.3%, 1.8%, 1.2%, and 51.6%, respectively. Seroprevalence of BAVs and MCF were more common than all other viruses (P < 0.0001). Ten sheep (37.0%), 48 goats (84.2), and 1 Ilama (14.2%) were the only species positive for MCF antibodies. Prevalence of BRSV and MCF antibodies were found to be significantly higher in goats than in sheep. BTV antibodies were detected both in Cameroon sheep and mountain goats and suggest that zoo animals are at risk for BTV in endemic regions.     

A comparative study of bovine herpesvirus 1247 and equine herpesvirus 1 in ponies.

The clinical and immunological response of ponies exposed to a bovine herpesvirus isolate and equine herpesvirus 1 were compared. Each virus was inoculated into two ponies by the intranasal route. One uninoculated pony was used with each group as a contact control. The four inoculated ponies developed a mild rhinitis with an increase in rectal temperature. Virus was recovered from nasal secretions collected from the four inoculated and one contact pony. All ponies developed a serum neutralizing antibody to each virus. The data show that the two viruses are similar.     

Cloning and characterization of a genomic probe for malignant catarrhal fever virus

A genomic probe specific for malignant catarrhal fever (MCF) virus was cloned by using purified viral DNA from MCF-virus strain WCll. Restriction endonuclease analysis of the purified viral DNA was used to identify the cloned viral genomic fragment. Dot blot hybridization by use of the genomic probe (pRP-5) indicated that the probe hybridized specifically with WCll-MCF virus, as well as with one other isolate of MCF-associated herpesvirus. Hybridization also was observed to a non-MCF virus strain of bovine herpesvirus.     

Immunoblotting analysis of the reaction of wildebeest, sheep and cattle sera with the structural antigens of alcelaphine herpesvirus-1 (malignant catarrhal fever virus)

Malignant catarrhal fever (MCF) is a disease of cattle and some other ruminants caused by alcelaphine herpesvirus-1 (AHV-1), a virus of wildebeest. The disease also occurs in the absence of wildebeest and is then thought to be caused by a viral agent harboured by the sheep. The structural proteins of AHV-1 have been used as antigens for the immunoblotting analysis of sera from wildebeest, sheep and cattle infected by either AHV-1 or the "sheep-associated" form of the disease. Wildebeest sera showed a uniform response reacting strongly with six polypeptides. Sheep sera also gave positive results but individual sera reacted with varying subsets of the antigens recognized by wildebeest. These results support the earlier suggestion that sheep harbour a herpesvirus related to AHV-1. A bovine serum from a case of MCF caused by AHV-1 also reacted only with a subset of the six wildebeest-reactive polypeptides. Sera from cattle affected with the "sheep-associated" form of the disease gave reactions in only two of the eight cases tested; both positive sera reacted to a few polypeptides only.     

Frequency of persistent bovine viral diarrhea virus infection in selected cattle herds

Sera and blood buffy coat samples were obtained from 3,157 cattle in 66 selected herds. Antibodies to bovine viral diarrhea (BVD) virus were detected in 89% of the serum samples by immunoprecipitation or virus-neutralization tests. Cytopathic or noncytopathic BVD viruses were isolated from blood buffy coat samples from 60 cattle in 6 herds. A second blood buffy coat sample was obtained from 54 of the 60 cattle 2 months after the initial sampling, and BVD virus was isolated again from each cow. The 54 cattle were considered persistently infected with BVD virus. The frequency of persistent infection was 1.7%.     

Susceptibility of a fetal tongue cell line derived from bighorn sheep to five serotypes of bluetongue virus and its potential for the isolation of viruses

A cell line (BHFTE) was derived from a tongue explant of a bighorn sheep fetus (Ovis canadensis nelsoni). The cells have been maintained through 23 serial passages, and the modal number of chromosomes was calculated to be 55. Monolayer cultures were shown to be susceptible to various viruses, including bluetongue virus (BTV). Of 5 BTV serotypes (2, 10, 11, 13, and 17) tested, each produced a cytopathic effect (CPE) on initial passage at 33 C. A field isolate (serotype 10) of BTV from a black-tailed deer (Odocoileus hemionus columbianus) in its second passage in Vero-M cells also produced CPE when inoculated into BHFTE cells. Antigens of BTV were demonstrated by direct immunofluorescence in the cytoplasm of BHFTE cells inoculated with homogenates of chicken embryos injected with clinical specimens from a domestic sheep and an Arabian oryx (Oryx gazella leucoryx). A suspension of BTV-infected gnats (Culicoides spp.) produced CPE and BTV-specific fluorescence on the first passage in cells inoculated with a suspension of blood from sheep experimentally infected with BTV. Additionally, selected bovine viruses induced CPE in the cells. The cell line, which is free of mycoplasma and bovine viral diarrhea virus contamination, may be useful in diagnostic medicine and research involving the ruminant species.     

Isolation of bovine herpesvirus III from diarrheic feces

A herpesvirus was isolated from the feces of a cow with diarrhea. The viral isolate was identified as a herpesvirus on the basis of morphology and chloroform sensitivity. It was serologically distinct from bovine Herpesvirus I (IBR) and II (mammillitis virus) as well as equine herpesviruses and pseudorabies virus. It was serologically related to members of the bovine Herpesvirus III group.Experimentally inoculated calves developed a transient fever but no other clinical signs or lesions. The virus could be re-isolated from the calves and significant levels of virus neutralizing antibodies were present in the serum 40 days after inoculation.     

Encephalomyocarditis virus infection of captive elephants.

Four African elephants at each of 2 widely separated zoologic gardens in Florida died following a fulminating illness. Tissue suspensions obtained from an elephant from each of the zoologic gardens were inoculated into newborn mice, 3- to 4-week-old mice, and buffalo green monkey and baby hamster kidney cell cultures. Encephalitis and myocarditis developed in the mice. The cell cultures were destroyed within 24 to 72 hours, and intracytoplasmic viral inclusions were observed in infected cells by electron microscopy. The viral agent was neutralized by known antiserum to encephalomyocarditis virus.     

Gamma herpesvirus carrier status of captive artiodactyls

Between 1998 and 2000, 103 individuals of 19 species of the order Artiodactyla at Whipsnade Wild Animal Park were tested for evidence of infection with gamma herpesviruses in order to distinguish between species which are susceptible to malignant catarrhal fever (MCF), caused by alcelaphine herpesvirus-1 (AlHV-1) of wildebeest (Connochaetes sp.) or ovine herpesvirus-2 (OvHV-2) of domestic sheep, and species which carry related viruses sub-clinically. Gamma herpesvirus DNA was detected in the known, or suspected, carrier species: roan antelope (Hippotragus equinus), scimitar-horned oryx (Oryx dammah), gemsbok (Oryx gazella), musk ox (Ovibos muschatus) and mouflon (Ovis musimon). In six other species: lowland anoa (Bubalus depressicornis) yak (Bos grunniens), sitatunga (Tragelaphus spekei), greater kudu (Tragelaphus strepsiceros), waterbuck (Kobus ellipsiprymnus) and Nile lechwe (Kobus megaceros), DNA was present in some newborn calves and over 30% of adults, strongly suggesting a carrier state. In contrast five Père David's deer (Elaphurus davidianus) and two swamp deer (Cervus duvauceli) died of MCF during the study. A virus isolated from scimitar-horned oryx calves produced cytopathic effects in scimitar-horned oryx kidney cell-culture and caused MCF in a rabbit.     

Transmission of caprine herpesvirus 2 in domestic goats

Caprine herpesvirus 2 (CpHV-2) is a recently recognized gammaherpesvirus that is endemic in domestic goats and has been observed to cause clinical malignant catarrhal fever (MCF) in certain species of deer. In this study, transmission of CpHV-2 in goats was examined. A total of 30 kids born to a CpHV-2 positive goat herd were selected and divided into two groups: group 1 (n=16) remained in the positive herd; group 2 (n=14) was separated from the herd at 1 week of age after obtaining colostrum. Peripheral blood samples from each kid were examined regularly by competitive ELISA for MCF viral antibody and by PCR for CpHV-2 DNA. Fifteen out of 16 goats (94%) that remained with the positive herd seroconverted and became PCR-positive for CpHV-2 by 10 months of age. In contrast, all kids (100%) that were separated from the positive herd at 1 week of age remained negative until termination of the experiment at 1 year of age. Additional transmission experiments revealed that all CpHV-2-free adult goats were susceptible to CpHV-2 or ovine herpesvirus 2 (OvHV-2) infection. The data indicate that the transmission pattern of CpHV-2 in goats is similar to the pattern of OvHV-2 in sheep and that CpHV-2-free goats can be established by early separation of kids from positive herds, which has significant implications for MCF control programs.     

Diagnosis of malignant catarrhal fever by PCR using formalin-fixed, paraffin-embedded tissues.

A previously described polymerase chain reaction (PCR) assay (amplification of a 238-bp fragment of ovine herpesvirus 2 [OHV-2] genomic DNA) for diagnosis of sheep-associated malignant catarrhal fever (MCF) was adapted for use on formalin-fixed, paraffin-embedded tissues. Variables affecting its use were examined. Archived tissues from cattle, white-tailed deer (Odocoileus virginianus), and bison (Bison bison) diagnosed with MCF by clinical signs or histologic lesions were obtained from 2 veterinary diagnostic laboratories. Tissues from healthy animals and from animals diagnosed with other common bovine viral diseases were examined as controls. A total of 86 blocks from 37 suspect MCF cases were examined. Forty-one blocks from healthy animals and animals with unrelated viral diseases were examined as controls. The assay was specific for sheep-associated MCF and did not yield false-positive signals from healthy animals or from cases of infectious bovine rhinotracheitis, bovine virus diarrhea, mucosal disease, or parainfluenza-3 virus infection. A wide variety of tissues were suitable substrates, including spleen, lymph node, intestine, brain, lung, and kidney. Extracted DNA provided a more suitable target than did unextracted tissue lysate. The highest levels of viral DNA were present in lymphoid organs and intestine, but the data indicate that in acute clinical cases, most organs contain sufficient viral DNA to serve as a suitable diagnostic specimen. Fixation of 0.5-cm3 blocks of tissue in 10% neutral buffered formalin was deleterious to the target DNA, and PCR signals progressively diminished after fixation for >45 days. Detection of genomic DNA of OHV-2 by PCR was successful for archived tissues that were 15 years old.     

Reproduction of mucosal disease with cytopathogenic bovine viral diarrhoea virus selected in vitro

Isolates of non-cytopathogenic bovine viral diarrhoea (BVD) virus from 18 persistently infected calves from one herd were compared by using monoclonal antibodies directed against the major viral glycoprotein gp53. All the isolates displayed an almost identical reaction pattern. Based on this antigenic analysis three cytopathogenic BVD and three non-cytopathogenic BVD viruses closely related to the non-cytopathogenic BVD herd isolate were selected. Six of the persistently infected calves were inoculated with a pool of the three closely related cytopathogenic BVD viruses and two with a pool of the three non-cytopathogenic BVD viruses. In addition three animals were infected with one closely related cytopathogenic BVD strain (Indiana) and two animals with the antigenetically different cytopathogenic BVD viral strain A1138/69. Regardless of the inoculation route all the animals superinfected with closely related cytopathogenic BVD viruses developed the characteristic lesions of mucosal disease within 14 days of infection. Animals which were inoculated with non-cytopathogenic BVD viruses which closely resembled the herd isolate, or with cytopathogenic BVD viruses which did not resemble the herd isolate did not develop any signs of disease. However, the latter group produced antibodies to the superinfecting virus.     

Bovine herpes virus expressing envelope protein (E2) of bovine viral diarrhea virus as a vaccine candidate.

The gene encoding the envelope protein (E2) of bovine viral diarrhea virus (BVDV) was expressed under the thymidine kinase (TK) promoter of Korean bovine herpesvirus 1 (BHV-1) isolate. Thymidine kinase negative (TK-) BHV-1 recombinants expressing E2 of BVDV were constructed and the expression of E2 was identified by immunofluorescence and Western blotting. Compared to wild type BHV-1, the recombinant BHV-1 had a delayed cytopathogenic effect in cells. The immunogenicity of the recombinant BHV-1 was examined in guinea pigs and cattle. Although an increase in body temperature was detected for a few days, the inoculated cattle returned to normal temperature with the development of neutralizing antibodies to BVDV.     

Ultrastructure of cellular changes in the replication of the alcelaphine herpesvirus-1 of malignant catarrhal fever

Cell cultures inoculated with 5 different viral isolates from 4 species of ruminants with clinical signs of malignant catarrhal fever (from the San Diego Wild Animal Park) were examined by electron microscopy. Each had the morphology of a herpesvirus (118 to 220 nm) and was icosahedral, and the nucleocapsid matured in the nucleus of the infected cell. Envelopment of budding occurred with each viral isolate at the nuclear and the plasma membranes. The virions egressed from the cell by budding from the plasma membrane or through channels of the Golgi apparatus or the endoplasmic reticulum. A proposed scheme for the morphogenesis of the herpesvirus of malignant catarrhal fever is presented.     

Malignant catarrhal fever. I. Response of American cattle to malignant catarrhal virus isolated in Kenya.

Fifty-three American cattle were inoculated with malignant catarrhal fever virus isolated from a wildebeest in Kenya. Three animals showed the mild form of the disease and recovered, and 47 showed the severe form of the disease. The other three did not react. Of the 47 cattle, 28 died, 16 were killed for the collection of specimens and three recovered. The incubation period for the 47 cattle ranged from 16 to 29 days and the course of the fatal disease for 28 cattle averaged three to 23 days. Virus titration of specimens from nine infected steers yielded a mean titer of 10(4)/TCID50 per gm for lymph nodes, 10(3) TCID50 per mL for buffy coats and 10(2.3) TCID50 per gm for spleens. Smaller amounts of virus were found in the liver, kidneys, adrenals and thyroids. Malignant catarrhal fever virus was also found in nasal secretions and saliva of viremic cattle. Viral infectivity was shown in bovine buffy coat cells stored at 4 degrees C for two days but was immediately destroyed upon freezing even when glycerine or dimethylsulfoxide was added. Viral particles were not found in infected animal tissues by electron microscopy. The disease was successfully transmitted in steers by intratracheal intubation and by aerosol inhalation but not by contact.