Transmission of virus from serosal fluids and demonstration of antigen in neutrophils and mesothelial cells of cattle infected with bovine ephemeral fever virus

Following intravenous injection of bovine ephemeral fever (BEF) virus 6 cattle were autopsied after clinical disease became evident. Fluid from serosal cavities with serofibrinous inflammatory changes showed large increases in neutrophil numbers. BEF virus was detected for the first time in pericardial, thoracic and abdominal fluids. Virus was also detected in synovial fluids, confirming an earlier report of transmission with a synovial fluid sample. Using a direct fluorescent antibody technique, BEF virus antigen was identified for the first time in synovial, pericardial, thoracic and abdominal fluids, in synovial membranes and epicardium. In synovial membranes and epicardium, specific fluorescence was observed in two cell types, mesothelial cells and neutrophils. In the fluids, fluorescence was restricted to neutrophils, the predominant cell type. Specific fluorescence was observed in blood smears from only one animal although blood samples collected at autopsy from all animals contained infective virus.     

The indirect fluorescent antibody technique as a method for detecting antibodies in aborted fetuses.

In this investigation the indirect fluorescent antibody technique was used to titrate antibodies in bovine sera to parainfluenza 3, infectious bovine rhinotracheitis virus and bovine viral diarrhea virus. These results were compared to those determined on the same samples by hemagglutination inhibition for parainfluenza 3 virus and serum neutralization for bovine virus diarrhea and infectious bovine rhinotracheitis virus. The results of the serological methods agreed closely. The indirect fluorescent antibody technique is a rapid and sensitive method for detecting antibodies and the procedure lends itself to use in diagnostic laboratories. In addition to the above viruses the presence or absence of antibodies to bovine coronavirus and bovine adenovirus 3 were determined by the indirect fluorescent antibody technique in thoracic fluids from 100 aborted fetuses and 50 nonaborted fetuses. Results on these samples were not compared to hemagglutination inhibition or serum neutralization as the condition of fluid samples from aborted fetuses renders interpretation of such tests unreliable. Antibodies to one or more viruses were detected in 30 of the 100 aborted fetuses and in seven of the 50 nonaborted fetuses. Antibodies to more than one agent were detected in eleven of the 100 aborted and in one of the 50 nonaborted fetuses. Reasons for this occurrence and application of the test in determination of causes of abortion are discussed.     

Sero-survey on Aino, Akabane, Chuzan, bovine ephemeral fever and Japanese encephalitis virus of cattle and swine in Korea

Vector-borne arboviruses produce mild to severe symptoms in domestic animals. Bovine ephemeral fever (BEF), Akabane, Aino, and Chuzan virus have been primarily attributed to reproductive disorders or febrile diseases in cattle, and Japanese encephalitis virus (JEV) is mainly associated with reproductive failures in swine. We investigated antibody titers from domestic swine against four bovine arboviruses (BEF, Akabane, Aino, and Chuzan virus) and from cattle against JEV in Korea. While the positive rates for Akabane and BEF were 37.4% and 15.7%, the positive incidence of Chuzan and Aino were relatively low, with positive rates of 3.04% and 0.4%, respectively, based on a virus neutralization assay. Antibody titers against more than one virus were also frequently detected in domestic swine. The incidence of JEV was 51.3% among domestic cattle. In addition, one positive case was detected in the thoracic fluids from 35 aborted calves, based on the hemagglutination inhibition test. Our results indicate that swine are susceptible hosts of bovine arboviruses without showing clinical symptoms in a natural environment. Moreover, we confirmed that JEV could be associated with reproductive failure in pregnant cattle, as were other vector-borne bovine arboviruses assessed in this study.     

Outbreaks in Quebec pig farms of respiratory and reproductive problems associated with encephalomyocarditis virus.

Encephalomyocarditis virus (EMCV) was isolated from tissues of aborted fetuses and weaned and suckling piglets from 4 different pig farms in Quebec. The farms were experiencing reproductive failure in sows of different parities concomitant to respiratory problems in suckling and postweaning piglets. At necropsy, gross lesions were confined to the lung and consisted of pulmonary congestion and edema of various degrees. Lesions of multifocal interstitial to proliferative pneumonia were found in the lungs of these piglets. Bacteriologic examination of various tissues from necropsied pigs yielded no pathogens in most cases. No significant antibody titers against 3 swine viruses (transmissible gastroenteritis virus, porcine parvovirus, and swine influenza virus) and two bovine viruses (bovine viral diarrhea and infectious bovine rhinotracheitis viruses) were detected in the sera of convalescent pigs. The Quebec EMCV isolates were antigenically related to the reference ATCC-VR129 strain of EMCV, as demonstrated by indirect immunofluorescence, serum neutralization (SN), and Western immunoblotting. However, one of the Quebec isolates could be distinguish by SN. EMCV-specific SN antibody titers up to 1:12,800 were detected in thoracic and ascitis fluids of aborted fetuses and in sera of convalescent pigs. A possible pneumotropic EMCV variant in swine may exist.     

Prevalence of bovid herpesvirus-4 and its antibody in cattle in Minnesota

Serologic analyses and virus isolation studies were carried out to determine the role of bovid herpesvirus-4 (BHV-4) in infections in cattle, principally those of the reproductive tract. Serologic analyses were performed, using an indirect fluorescent antibody test on thoracic fluid specimens from aborted fetuses and on sera from 3 sources of adult cattle. Virus isolation was attempted from field cases of abortion, early embryo death, and postpartum vulvovaginitis/metritis, using uterine discharge and buffy coat preparations obtained from cows and tissues obtained from aborted fetuses. Of 420 fetal thoracic fluid specimens examined, 5 were positive for BHV-4 antibodies. Seventeen percent of adult cattle from 2 sources ie, clinically normal herds and abattoir cattle, were seropositive for BHV-4 antibodies. Cattle from a third source, 4 herds with high incidence of reproductive tract disorders, had a seroprevalence rate between 36 and 88%. Two isolates of BHV-4 were also obtained from this group. The overall incidence of BHV-4 antibodies in clinically normal cattle was higher than previously recognized, with relatively higher prevalence in herds having reproductive problems (chi 2 = 156.5, P less than 0.005). At least 10% of the BHV-4 antibody-positive sera did not have neutralizing antibody against bovine viral diarrhea virus and/or bovid herpesvirus-1, both important causes of bovine reproductive tract disorders.     

牛呼吸道疾病的病原学与防制研究进展

牛呼吸道疾病的出现多是由牛疱疹病毒1型(BHV-1)、牛呼吸道合包体病毒(BRSV)、牛病毒性腹泻病毒(BVDV)、牛副流感病毒3型(BPIV3)、牛冠状病毒(BCV)、牛支原体、出血性败血性巴氏杆菌等病原的单独或混合感染造成的。文章针对这几种病原的病原学及相应的防制研究进行了综述。     

Molecular cloning of complementary DNA from a pneumopathic strain of bovine viral diarrhea virus and its diagnostic application.

A pneumopathic strain of bovine viral diarrhea virus was grown in cell culture and purified. Genomic ribonucleic acid was extracted, polyadenylated at the 3' end, and copied into complementary DNA after oligo-dT priming. Complementary DNA was male double stranded and cloned into the pUC9 plasmid. Approximately 200 complementary DNA clones varying in length from 0.5 to 2.5 kilobases were obtained. Hybridization assays indicated that the sequences isolated were specific for bovine viral diarrhea virus and that at least 5.5 kilobases of bovine viral diarrhea virus genome was represented in the library of complementary DNA clones, the majority of which may have originated from the 3' end of the virus genome. One cloned complementary DNA sequence was used as a 32P-labelled hybridization probe for bovine viral diarrhea virus detection. The probe hybridized with all cytopathic and noncytopathic strains of bovine viral diarrhea virus tested and was 100 times more sensitive than infectivity assays for the detection of bovine viral diarrhea virus. Hybridization did not occur with nucleic acids from bovine coronavirus, bluetongue virus, bovine adenovirus or uninfected cell cultures. Native plasmid DNA sequences, labelled with 32P, did not hybridize with bovine viral diarrhea virus ribonucleic acid.     

Radiographic detection of thoracic lesions in adult cows: a retrospective study of 42 cases (1995-2002)

Medical records of 42 cows that underwent both thoracic radiographic and postmortem examinations within a period of 7 days were reviewed to develop an evaluation grid to interpret bovine thoracic radiographs and to determine the sensitivity and the specificity of thoracic radiographs for detection of thoracic lesions, based on postmortem examination. Most cows (64%) had clinical signs of respiratory disease, whereas 19% showed signs of cardiac problems. The sensitivity and specificity of radiographs for identifying cows with thoracic lesions were 94% and 50%, respectively. In this study, with a prevalence of thoracic lesions of 86%, the positive- and negative-predictive values were 92% and 57%, respectively. This study provides an evaluation grid that allows standardization of the reading of bovine thoracic radiographs and the identification of most thoracic lesions. Bovine thoracic radiographs are useful in detecting thoracic lesions in cows.     

牛副流感病毒3型RT—PCR检测方法的建立

参照GenBank中公布的牛副流感病毒3型（Bovineparainfluenzavirus3,BPIV-3）全基因序列,针对BPIV-3特异性NP蛋白保守基因设计一对引物,建立了BPIV-3的RT—PCR诊断方法。其最佳扩增退火温度为58．1℃,引物浓度为1．0μmol／L。采用该方法扩增BPIV-3参考病毒．能扩增出425bp预期大小的特异性片段,而扩增牛病毒性腹泻／黏膜病病毒、牛传染性鼻气管炎病毒、牛合胞体病毒、猪瘟病毒以及牛支原体、大肠埃希氏菌、牛巴氏杆菌和沙门氏菌等常见病毒和细菌均呈阴性结果。对参考病毒进行梯度稀释检测,结果证明该法检测BPIV-3的灵敏度可达10^-3FCID50／0．1mL。     

A syncytium regression test to detect antibodies to bovine syncytial virus.

A test to detect antibodies to bovine syncytial virus was developed from the observation that syncytia in monolayer cell cultures infected with bovine syncytial virus regress and disappear in the presence of bovine syncytial virus antibodies. The test is useful in monitoring the presence of bovine syncytial virus and bovine syncytial virus antibodies in cattle used in studies on bovine leukosis.     

Susceptibility of bovine macrophage and tracheal-ring cultures to bovine viruses.

Cultures of macrophages initiated from peripheral blood monocytes and organ cultures of tracheal rings were tested for their susceptibility to bovine viruses. With several notable exceptions, viruses cytopathogenic for bovine embryonic lung cultures were cytopathogenic for macrophages. Although cowpox virus replicated in macrophages, pseudocowpox did not, and although pseudorabies virus replicated within macrophages, infectious bovine rhinotracheitis and DN-599 herpesviruses did not. Bluetongue virus established an interesting relationship with macrophages. Whereas bluetongue virus was initially cytopathogenic for macrophages, it lost its cytopathogenicity on repeated passage, although it was capable of continued replication in macrophages. When subsequently passaged onto bovine embryonic lung cultures, it regained its cytopathogenicity. Parainfluenza-3, bovine viral diarrhea, and infectious bovine rhinotracheitis viruses readily destroyed ciliary activity in tracheal-ring cultures, as contrasted with the inability of bovine respiratory syncytial virus to destroy ciliary activity, even though bovine respiratory syncytial virus was able to replicate within ciliated epithelial cells of tracheal rings.     

BRS virus, PI3 virus and BHV1 infections of young stock on self-contained dairy farms: epidemiological and clinical findings

The role of bovine respiratory syncytial virus, parainfluenza type 3 virus and bovine herpesvirus 1 as disease agents in 28 groups of young cattle on 19 dairy farms which raised their own replacements was investigated. Bovine respiratory syncytial virus infections occurred in 27, parainfluenza type 3 virus infections in all and bovine herpesvirus 1 infections in three of the 28 groups. Some infections were accompanied by clinical signs while others were entirely subclinical. Clinical respiratory disease was observed on 25 occasions in 20 of the groups. Respiratory disease was associated with a bovine respiratory syncytial virus infection on 15 occasions with parainfluenza type 3 virus infection in four cases and with bovine herpesvirus 1 infection in two cases. In four cases there was no association between the respiratory disease and any of the four virus infections. Bovine respiratory syncytial virus infections caused more serious respiratory problems than parainfluenza type 3 virus infections.     

In vitro interferon production by bovine tissues: effects of hydrocortisone.

The effect of hydrocortisone on bovine interferon production in vitro was studied. Infectious bovine rhinotracheitis virus was used as an inducer. Interferon was assayed by the plaque-reduction method in bovine fetal kidney cultures, using vesicular stomatitis virus as challenge virus. Hydrocortisone decreased interferon production in bovine fetal spleen and peripheral blood leukocyte cultures. Hydrocortisone did not decrease interferon production by bovine alveolar macrophages, in 1 experiment. Properties of viral inhibitors were those of interferon.     

The association between serological titers in infectious bovine rhinotracheitis virus, bovine virus diarrhea virus, parainfluenza-3 virus, respiratory syncytial virus and treatment for respiratory disease in Ontario feedlot calves.

A seroepidemiological study of the association between antibody titers to infectious bovine rhinotracheitis, parainfluenza-3, bovine virus diarrhea and bovine respiratory syncytial viruses, and treatment for bovine respiratory disease was conducted. A total of 322 calves from five different groups were bled on arrival, then one month later all cases (cattle treated for bovine respiratory disease) were rebled together with an equal number of controls (cattle not treated for any disease). Titers to these viruses varied significantly from group to group. Based on seroconversion, infectious bovine rhinotracheitis virus was active in 4.4%, bovine virus diarrhea virus in 24%, parainfluenza-3 virus in 69.5% and bovine respiratory syncytial virus in 71.3% of the cattle. Cattle with low titers to infectious bovine rhinotracheitis and/or bovine respiratory syncytial viruses on arrival, were at increased risk of subsequent treatment for bovine respiratory disease. Treated cattle also had significantly greater increases to parainfluenza-3 and/or bovine virus diarrhea viruses than control calves. Treatment rates varied considerably from group to group and were not strongly correlated with weight gain in the postarrival period.     

Genetic comparison of ovine and bovine pestiviruses

Viral RNA oligonucleotide fingerprinting was used to compare genetic relationship among pestiviruses originating from ovine or bovine host species. Ovine pestiviruses, including reference border disease virus and 2 border disease isolates originating from natural pestivirus infections of sheep, appeared to have a more distant genetic relationship among themselves than with certain bovine pestiviruses. A closer genetic relatedness was evident between border disease virus and 3 noncytopathic bovine pestiviruses, including Draper bovine viral diarrhea virus (BVDV), a BVDV isolate that originated from aborted bovine fetuses, and a virus that was isolated from the serum of a calf that had a chronic BVDV infection. Four noncytopathic bovine viruses, including Draper BVDV and 3 field isolates, were closely related. Reference Oregon C24V BVDV, a cytopathic virus, was closely related to only 1 of the 7 noncytopathic viruses in this study.     

Detection of bovine leukemia virus by syncytium assay.

When various indicator cells, including virus transformed and nontransformed cells, were cocultivated with bovine leukemia virus-producing cells, strong positive syncytia formation was found in transformed cells one day after cocultivation. The results of comparison of bovine leukemia virus antibody titers and the detection of bovine leukemia by the syncytium assay showed 89% of serologically positive cows were positive for bovine leukemia virus, whereas no reactors were found in serologically negative cows. However, the frequency of bovine leukemia virus detection differed according to the difference of incubation periods in the syncytium assay. Therefore, it is important to choose the appropriate indicator cell and culture conditions for the detection of bovine leukemia virus in the syncytium assay.     

Prevalence of antibodies to seven viruses in a flock of ewes in Minnesota

Blood samples were collected from a flock of healthy ewes at a University of Minnesota research station. Sera from these blood samples were tested for antibodies against 7 viruses, using 3 tests (eg, virus-neutralization test for bovine viral diarrhea virus, infectious bovine rhinotracheitis virus, bovine adenovirus type 3, and bovine respiratory syncytial virus; hemagglutination inhibition test for parainfluenza virus type 3; and agar-gel immunodiffusion test for lentivirus of ovine progressive interstitial pneumonia and bluetongue virus). The number of seropositive ewes for each antibody type were 1 of 377 (0.3%) for bovine viral diarrhea virus, 2 of 377 (0.5%) for infectious bovine rhinotracheitis virus, 29 of 378 (7.6%) for bovine adenovirus type 3, 200 of 378 (52.5%) for bovine respiratory syncytial virus, 273 of 373 (71.7%) for parainfluenza virus type 3, and 210 of 379 (55%) for ovine progressive pneumonia virus. All ewes were seronegative for bluetongue virus antibodies.     

Spread of bovine syncytial virus in a dairy herd over a two year period

During a two year period the spread of bovine syncytial virus was monitored in a closed herd of 50 to 100 milking cows. Out of a nucleus of 49 nonpregnant and pregnant heifers, six were found to be infected with bovine syncytial virus. Virus was detected only in the progeny of infected cows and not in the progeny of uninfected animals. Nineteen progeny of the bovine syncytial virus infected cows were studied in detail and virus was isolated from only four. Horizontal spread of the virus did not occur.     

Eradication of bovine leukemia virus infection in commercial dairy herds using the agar gel immunodiffusion test.

Demands for bovine leukemia virus test negative breeding cattle and for semen from bovine leukemia virus test negative bulls by several countries have encouraged the eradication of bovine leukemia virus infection from selected herds in Canada. This project was undertaken to evaluate the suitability of the agar gel immunodiffusion test, standardized to detect anti-bovine leukemia virus glycoprotein antibodies, for eradication of bovine leukemia virus from commercial dairy herds. Of nine participating herds, the prevalence rate of bovine leukemia virus infection was low (less than 10%) in three, medium (11-30%) in four and high (greater than 30%) in two. The herds were tested by the agar gel immunodiffusion test, reactors were removed and the herds were then retested at regular intervals. The results indicate that it is possible to eliminate bovine leukemia virus infection from the herds after two to three cycles of agar gel immunodiffusion tests and prompt removal of the reactors.     

Production of cattle immunotolerant to bovine viral diarrhea virus.

Inoculation of bovine virus diarrhea virus into 58 to 125 day old fetuses of bovine virus diarrhea virus seropositive pregnant cows, or inoculation of bovine virus diarrhea virus into seronegative cows 42 to 114 days pregnant, may produce clinically normal calves which are persistently infected with the specific isolate of bovine virus diarrhea virus yet seronegative to the homologous and heterologous isolates. Reinoculation of these persistently infected cattle with their homologous isolate produced no neutralizing antibody response to bovine virus diarrhea virus. These persistently infected cattle were immunocompetent as they developed neutralizing serotiters to infectious bovine rhinotracheitis, parainfluenza-3 viruses and agglutinating serotiters to Pasteurella hemolytica .