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.     

Prevention of bovine respiratory syncytial virus infection and clinical disease by vaccination

A double blind field trial was carried out with a live attenuated bovine respiratory syncytial virus vaccine. The trial involved 530 calves, two to 10 months old, on 27 dairy farms, where respiratory problems due to bovine respiratory syncytial virus infections had been observed during the preceding year. In 17 herds either all calves were vaccinated (nine groups) or all calves received a placebo (eight groups). In 10 herds half the number of calves were vaccinated and the other half kept as non-vaccinated controls. Calves were vaccinated intramuscularly twice with an interval of four to five weeks. These groups were under regular clinical observation and animals were tested periodically for antibodies to bovine respiratory syncytial virus and parainfluenza type 3 virus. Serological examination indicated that no bovine respiratory syncytial virus infection had occurred prior to the first vaccination in August. Vaccination did not cause adverse reactions. Low concentrations of neutralising and complement fixing antibodies were induced by vaccination and a sharp increase of antibody titres was observed after natural infection of vaccinated animals. Infections with bovine respiratory syncytial virus occurred in six out of eight non-vaccinated groups, in nine out of 10 partly vaccinated groups and in only two out of nine completely vaccinated groups. Virus infection in completely vaccinated groups was significantly reduced compared with partly vaccinated and non-vaccinated groups. The incidence of bovine respiratory syncytial virus lower respiratory disease was significantly reduced in completely vaccinated groups compared to non-vaccinated groups. Generally only mild signs of upper respiratory disease were present in completely vaccinated groups after bovine respiratory syncytial virus infection.(ABSTRACT TRUNCATED AT 250 WORDS)     

Prevalence of antibodies to bovine respiratory syncytial virus, bovine viral diarrhea virus, bovine herpesvirus-1, and bovine parainfluenza-3 virus in sheep and goats in Quebec

Serum samples were collected from 1,075 clinically normal sheep and goats from 77 flocks in 7 agricultural regions of Quebec from June to August 1982. Sheep and goats were tested for antibodies to bovine respiratory syncytial virus, bovine viral diarrhea virus, and bovine herpes-virus-1 by the indirect fluorescent antibody technique and for parainfluenza-3 virus by the hemagglutination inhibition test. The prevalence of antibodies in animals to respiratory syncytial virus was 31%; to bovine viral diarrhea virus, 22.2%; to bovine herpesvirus-1, 10.8%; and to parainfluenza-3 virus, 23.2%. Antibodies prevailed in similar proportions in young (less than 1 year) and adult (greater than 1 year) animals.     

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.     

Differences in fluorescent antibody staining of bovine respiratory syncytial virus-infected cells by ovine and bovine sera

In indirect fluorescent antibody tests in which sera from cattle and sheep with respiratory disease problems were used to stain foetal bovine lung cells infected with a bovine respiratory syncytial virus strain, differences were noted in the pattern of fluorescence produced by some sheep sera and that produced by positive bovine sera. In serum neutralisation tests, also using a bovine respiratory syncytial virus strain, 4 of 7 sera giving this atypical pattern of fluorescence had very low neutralising antibody titres (highest 1/4), and 3 were negative. It is suggested that two related but antigenically distinguishable respiratory syncytial virus types are present in sheep, one of which is similar to bovine strains.     

Studies on the occurrence and significance of bovine respiratory syncytial virus in Saskatchewan.

Seventy-six percent of 49 blood samples from Saskatchewan cattle had serum antibodies against bovine respiratory syncytial virus. Experimental infection of one week and seven month old calves with bovine respiratory syncytial virus (Iowa strain) caused transient fever, mucopurulent nasal discharge and coughing but no macroscopic or microscopic lesions attributable to bovine respiratory syncytial virus.     

Indirect haemagglutination test for the detection and assay of antibody to bovine respiratory syncytial virus

An indirect haemagglutination (IHA) test was used for the rapid assay of antibody to bovine respiratory syncytial virus. Antigens for the sensitisation of formalised tanned erythrocytes were prepared by treatment of virus infected cells with non-ionic detergent. A close serological relationship was shown by the IHA test between the strain of bovine respiratory syncytial virus used and the A2 strain of human respiratory syncytial virus. The IHA test was sensitive and reproducible. A linear correlation was demonstrated between antibody titres obtained by the IHA test and the serum neutralisation test. Titres obtained by the IHA test were approximately 60 times greater than serum neutralisation titres. Serum samples from 803 two-year-old heifers in 48 herds in England were examined by the IHA test. Ninety-four per cent of the animals had antibody to respiratory syncytial virus. Examination of paired serum samples from outbreaks of respiratory disease by the IHA test showed that respiratory syncytial virus was associated with seven out of 15 outbreaks.     

Characterization and identification of a bovine respiratory syncytial virus isolated from young calves.

Two identical viruses designated 371 and 375 were recovered from nasal secretions of 2 of 7 calves in a beef cow-calf herd in which calves (45 to 105 days of age) had signs of acute respiratory tract disease. The cytopathic, morphologic and physico-chemical characteristics of the isolates were those of bovine respiratory syncytial virus. Although a humoral antibody response to bovine respiratory syncytial virus was not observed, it was concluded that this virus probably had a part in the respiratory tract disease in these calves.     

Clinical signs following experimental lungworm infection and natural bovine respiratory syncytial virus infection in calves

Similar clinical signs have been reported in calves infected either by Dictyocaulus viviparus or bovine respiratory syncytial virus. Three experiments were carried out to establish the clinical picture and the course of the disease in animals with these infections. The clinical signs of calves infected with lungworm included coughing, nasal discharge, tachypnoea, abdominal breathing and pyrexia, and auscultation of their lungs revealed increased bronchial sounds. Similar signs were also observed after infection with bovine respiratory syncytial virus, but the signs were more acute and resolved more rapidly than in animals infected with lungworm larvae. Calves infected with lungworm had more serious clinical signs after infection with bovine respiratory syncytial virus than calves, which were not infected with lungworm.     

Application of a modified indirect fluorescent antibody test to the detection of antibodies to bovine respiratory syncytial virus in Ontario cattle.

A modified indirect fluorescent antibody test for the detection of serum antibodies to bovine respiratory syncytial virus was developed. The test made use of Terasaki plastic microtiter plates in which bovine respiratory syncytial virus (Saskatchewan strain) infected Georgia bovine kidney cells were grown and fixed in situ by a modified acetone fixation procedure. Evans blue dye was used as a counterstain to reduce nonspecific fluorescence. In a study of 986 field sera from a geographically broad cross-section of mature Ontario cattle, 95% of the samples were found to be positive at or above a 1:2 dilution. No seronegative regions, counties or herds were identified. When representative samples covering a range of indirect fluorescent antibody titers were further examined by a microtiter virus neutralization assay, a significant agreement was found between the two tests. Up to a fourfold decrease in titer was observed when antigen coated plates were stored at -70 degrees C for four months. The modified indirect fluorescent antibody test for bovine respiratory syncytial virus antibody detection proved to be a rapid, practical procedure for use in the diagnostic laboratory. This study confirms that bovine respiratory syncytial virus is widespread in the Ontario cattle population and that most mature cattle can be assumed to have been exposed to this virus.     

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.     

Effect of recombinant DNA-derived bovine and human interferons on replication of bovine herpesvirus-1, parainfluenza-3, and respiratory syncytial viruses

Antiviral effects of recombinant DNA-derived bovine (Bo) and human (Hu) interferons (IFN) on the replication of bovine herpesvirus-1, parainfluenza-3, and respiratory syncytial viruses were studied. Bovine monolayer cultures were treated with recombinant DNA-produced Bo IFN-alpha 1, Bo IFN-beta 2, Hu IFN-alpha A, or Hu IFN-alpha A/D and then challenge exposed with bovine herpesvirus-1, bovine parainfluenza-3 virus, bovine respiratory syncytial virus, or vesicular stomatitis virus. Treatment with each IFN reduced the viral yield for each of these viruses, compared with that of control cultures.     

The detection of bovine respiratory syncytial virus in formalin fixed bovine lung with commercially available monoclonal antibodies and avidin biotin complex immunohistochemistry.

Eight commercially available monoclonal antibodies directed against respiratory syncytial virus antigens were tested for ability to detect bovine respiratory syncytial virus (BRSV) antigen in formalin fixed, paraffin embedded bovine lung using avidin-biotin complex immunohistochemical staining. Monoclonal antibodies from clone 18B2 purchased from Biosoft, Paris, France and those from clone 8G12 purchased from the Department of Veterinary Sciences, University of Nebraska, Lincoln, Nebraska stained BRSV antigen in infected bovine lung with acceptable background staining of uninfected tissues. This method offers advantages over other techniques for BRSV diagnosis in that fresh tissue is not required and all reagents may be purchased commercially.     

Experimental infection of calves with bovine respiratory syncytial virus (Quebec strain).

An experiment was designed to evaluate the clinical, haematological, viral and serological aspects of bovine respiratory syncytial virus infection in calves. Eleven calves were inoculated intranasally with bovine respiratory syncytial virus (Quebec strain) in aerosol. Clinical, haemotological and serological responses of the calves and virus shedding were monitored. The experimentally infected animals manifested moderate to severe signs of respiratory disease. The parameters used to evaluate the severity of the disease included ocular discharge, conjunctivitis, lung sounds, nasal discharge, pyrexia and leukopenia. The animals were scored accordingly (scale infected 70.8-148.5, control 22-29.3). Highest disease scores were observed between day 6-9 after infection. Virological and serological assessment demonstrated that the observed clinical picture was due to bovine respiratory syncytial virus infection.     

Serologic studies of bovine respiratory syncytial virus in Minnesota cattle

Serum antibody titers to bovine respiratory syncytial virus were determined for 559 cattle. Serum samples were obtained through the Minnesota State-Federal Brucellosis Laboratory and were collected over a 1-year period. Results of this study revealed an antibody prevalence of 65.5% to bovine respiratory syncytial virus. The distribution of antibody titers is presented, as well as analysis of titers based on breed, sex, and age of the cattle.     

Association of bovine viral diarrhea virus with multiple viral infections in bovine respiratory disease outbreaks

We investigated eleven outbreaks of naturally occurring bovine respiratory diseases in calves and adult animals in the St-Hyacinthe area of Quebec. Specific antibodies to bovine herpesvirus-1, bovine viral diarrhea virus, respiratory syncytial virus, parainfluenza type 3 virus, reovirus type 3, and serotypes 1 to 7 of bovine adenovirus were found in paired sera from diseased animals. Several bovine viruses with respiratory tropism were involved concomitantly in herds during an outbreak of bovine respiratory disease. In addition, concomitant fourfold rises of antibody titers were frequently observed to two or more viral agents in seroconverted calves (61%) or adult animals (38%). Bovine viral diarrhea virus was found to be the most frequent viral agent associated with multiple viral infection in calves only (92%).     

Clinical and pathological observations on spontaneous bovine respiratory syncytial virus infections in calves

A clinical, pathological and microbiological study was made of acute spontaneous bovine respiratory syncytial virus infection in calves. Characteristic features were atelectatic areas, which had an exudative and, or, necrotising bronchiolitis with syncytial giant cells in the epithelial lining and in the lumina of these bronchioles. Restricted to these areas both bronchiolar and alveolar immunofluorescence for bovine respiratory syncytial virus were seen. Complications were severe interstitial oedema, interstitial emphysema and a catarrhal or fibrinous pneumonia due to secondary bacterial invasion.     

Association of bovine respiratory syncytial virus with atypical interstitial pneumonia in feedlot cattle

Thirty-three cattle with fatal respiratory tract disease were examined for gross and histologic lesions and for the presence of viral and bacterial agents in the lungs. Fifteen cattle had lesions characteristic of atypical interstitial pneumonia (AIP), and 18 had other respiratory tract diseases, including infectious bovine rhinotracheitis, shipping fever pneumonia, bronchopneumonia, pulmonary abscess, and edema of the trachea. Gross necropsy findings in the cattle with AIP were uncollapsed and emphysematous lungs; histopathologic findings included interstitial edema, thickening of alveolar walls, hyaline membrane formation, and hyperplasia of type-II pneumonocytes. The infective agents found in the lungs of the 33 cattle included bovine respiratory syncytial virus, bovine herpesvirus type 1, Pasteurella sp, mycoplasmas, and Corynebacterium pyogenes. Bovine respiratory syncytial virus was detected by use of immunofluorescence and immunoperoxidase on lung tissue sections; bovine herpesvirus type 1 was detected by these techniques and by isolation of the virus. Bovine respiratory syncytial virus was significantly (P = 0.01) associated with lesions of AIP (11 of 15), compared with those of other respiratory tract diseases (5 of 18).     

Replication of parainfluenza type-3 virus and bovine respiratory syncytial virus in isolated bovine type-II alveolar epithelial cells.

The objectives of our research were to determine whether bovine pulmonary type-II alveolar epithelial cells could be isolated from bovine lung and maintained in tissue culture and to determine whether isolated bovine type-II alveolar epithelial cells would support productive viral replication of bovine parainfluenza type-3 virus and bovine respiratory syncytial virus. Type-II alveolar epithelial cells were isolated from lungs of 4- to 7-day-old male Holstein calves by enzymatic dissociation of pulmonary tissue with trypsin and by separation of cells with the use of filtration and centrifugation on continuous Percoll gradients. Cells were further separated by panning on IgG-coated plastic plates and by lectin binding. Isolated type-II alveolar cells were maintained on basement membrane-coated tissue cultured plates. In culture, type-II cells formed alveolar structures and maintained other cytologic features of type-II cells, including osmiophilic lamellar inclusions. Cell cultures were inoculated with and supported productive replication of bovine parainfluenza type-3 virus and bovine respiratory syncytial virus. This was determined by recovery of infectious viruses from inoculated cell cultures and by identification of viral structures in type-II alveolar epithelial cells by transmission electron microscopy.     

Effect of 2-deoxy-D-glucose and glucosamine on bovine respiratory syncytial virus

2-Deoxy-D-glucose (2-dG) and glucosamine reversibly inhibited the replication and cytopathic effects of bovine respiratory syncytial virus in bovine turbinate cell cultures. The inhibitors were effective when added up to 12 hours after cell cultures were inoculated. Their effectiveness decreased as the time between inoculation of cells and drug treatment lengthened. The sugars did not inactivate the virus directly, and inoculation of drug-treated cells in drug-free medium produced normal yield of virus. Mannose fully reversed the inhibitory effects of 2-dG, but was ineffective for glucosamine. Protein synthesis was necessary for production and release of infective virus after removal of 2-dG and glucosamine blocks. Electron microscopic studies revealed a large amount of bovine respiratory syncytial virus production with characteristic spike-like projections on the viral envelopes in the absence of these inhibitors. There was a drastic reduction in the number of mature virions produced in inhibitor-treated bovine turbinate cells, and the virions lacked the characteristic surface projections.