Bucyrus Strain Research Articles

Ultrastructure and immuno-cytochemistry of BHK-21 cells infected with a modified Bucyrus strain of equine arteritis virus.

SummaryMorphogenesis of a modified Bucyrus strain of equine arteritis virus (EAV) in BHK-21 cells was studied. Bacillary tubules were first detected in the cytoplasm 8 h after infection, and mature virions 79 to 122 nm in diameter, 101 nm on average, were mostly observed in the cisternae of the rough endoplasmic reticulum (RER) at 12 h or later. They had isometrical cores and morphological subunits in the outer layer. Budding occurred from the RER and the outer nuclear membrane, but not from the cell surface. Structural linkage was detected between the tubule and the virus core. Aberrant strands were occasionally demonstrated within the nucleus 12 h after infection, and immunofluorescence and immunogold labeling revealed viral antigen also in the nucleus.

Use of the serum neutralisation test for equine viral arteritis with different virus strains

Serum cross neutralisation tests were conducted with a recent American isolate (84KY-A1) and a European isolate, (Wroclaw-2) and compared with the prototype and modified viruses of the Bucyrus strain of equine arteritis virus by using virus specific immune horse sera. The modified Bucyrus strain was neutralised and showed high neutralisation titres with all the immune sera. The prototype Bucyrus strain was also substantially neutralised, followed by the 84KY-A1 strain. As a result of the tests with the modified Bucyrus strain as the antigen, 20 seropositive horses were discovered among home-bred horses which had no previous record of clinical equine viral arteritis. Heat inactivation of the sera at 62 degrees C for 30 minutes caused the disappearance of all but the most positive reactions. When the prototype Bucyrus strain was used instead of the modified virus, no positive reactions were detectable. A serological survey, using the Bucyrus strain in a microtitre neutralisation test, was conducted with 1656 horse sera collected between 1988 and 1990 in Japan. The test disclosed only eight foreign-bred horses positive to the virus; they had been imported as competition horses. No circumstantial evidence of an effect of such horses as a source of infection for horse populations free of the virus was obtained.

Histopathological and Immunofluorescent Studies on the Uterus of Aborted Mares Experimentally Infected with Equine Arteritis Virus.

Two pregnant mares, at 5 and 6 months gestation were inoculated intranasally with the Bucyrus strain of equine arteritis virus (EAV). Both mares aborted at 1 1 days after the inoculation. Multi focal and necrotizing myometritis was found histopathologically in the uterus. Large numbers o f mononuclear cells had infiltrated the smooth muscle and interstitium o f the myometrium. The smooth muscle cells showed signs o f degeneration with sarcoplasmic vacuolation, and necrosis with acidophilic sarcoplasm and pyknosis or karyorrhexis. EAV antigens were detected in the smooth muscle cells o f the myometrium. These findings suggested that a replication o f EAV had occurred in the smooth muscle cells o f the uterus, resulting in the myometritis.

Studies on Equine Viral Arteritis : I. Characterization of the Virus and Trial Survey on Antibody with Vero Cell Cultures

Two strains, the Bibuna and the Bucyrus, of equine arteritis virus (EAV) were tested for propagation in various cell cultures. They were grown in primary cell cultures of horse, rabbit, hamster, cat and pig kidney, and in established cell cultures of baby hamster kidney (BHK-21/13), hamster lung (HmLu), African green monkey kidney (Vero) and Cynomolgus monkey kidney (JINET). Cytopathic changes were comparatively common to all cells, and characterized by rounding and vacuolation of the cells and by increased density of the cytoplasm. Most affected cells were detached from the glass wall within 7 days after inoculation. The infectivity titers of the primary cell culture fluids were 104 to 106 TCID50/0.2 ml, and those of the established cell culture fluids 106 to 108 TCID50/0.2 ml. Both strains were able to replicate in the presence or absence of 5-iodo-2-deoxyuridine (IUDR). They were not stabilized by cation at 50°C, but were highly sensitive to ether and sodium deoxycholate, and moderately sensitive to heat and acid. The Bibuna strain was more sensitive to a high concentration of trypsin than the Bucyrus strain. Each virus formed distinct piadues 2 to, 3 mm in diameter on the monolayer of Vero cells. Serum samples were collected from 107 horses in Japan in 1972 and 1978 and tested for EAV antibody by neutralization tests in monolayer cell culture tubes or plaque reduction neutralization tests in Vero cells. No antibody, however, was detected from any sample so far.

Temporal Distribution of Equine Arteritis Virus in Respiratory Mucosa, Tissues and Body Fluids of Horses Infected by Inhalation

SUMMARY Horses exposed to aerosols containing the virulent Bucyrus strain of equine arteritis virus (EAV) developed a high febrile response, lymphopenia and, usually, severe signs of disease. Virus was recovered from the serum for as long as the nth day after exposure, from the nasopharynx up to the 14th day and from the huffy coat up to the 19th or terminal day of the experiment. Quantitative determinations of the viral content of various tissues and fluids collected from 18 horses serially sacrificed or succumbing to infection from 1 to 19 days after exposure indicated that the cells initially infected were those of the lung, followed by the bronchial lymph nodes from which the virus was disseminated throughout the body by the circulatory system. Virus was generally present throughout the body by the 3rd day and persisted in high concentrations, especially in the lymph nodes and organs for 9 days after which it diminished, except in kidney and urine, where it remained at a high level until the 19th or terminal day of the experiment.

Equine arteritis virus: ferritin-tagging and determination of ribonucleic acid core.

Equine arteritis virus (EAV), Bucyrus strain, has been identified in infected cultures of equine dermis cells by both direct and indirect ferritin-tagging. Furthermore, differential cytological staining in both glycolmethacrylate and Epon embedded thin sections shows that EAV has a ribonucleic acid (RNA) core.

Buoyant density studies on equine arteritis virus.

The buoyant density of the Bucyrus strain of equine arteritis virus was studied by isodensity centrifugation on three types of gradients. The distribution patterns of infectivity were dependent on the type of gradient used. In sucrose gradients a single peak of infectivity at densities approximating 1.17 g/ml was obtained. From this gradient the total recovery of infectious virus varied between 80 and 90%. In cesium chloride gradients a rather broad band appeared in the form of a “saddled” peak with a range of densities from 1.180 to 1.215 g/ml, the total recovery being 90%. In potassium tartrate (KT) gradients two peaks were formed, one at 1.17 and another at 1.24 g/ml; the total recovery was 50%. Recentrifugation of fractions from KT gradient on sucrose gradients resulted in a changed distribution of infectivity for particles of higher density (1.24 g/ml).

A plaque assay of equine arteritis virus in BHK-21 cells.

The Bucyrus strain of equine arteritis virus (EAV) was grown in BHK-21, primary rabbit kidney and primary horse testicle cells. Viral yields from these cells produced distinct plaques in BHK-21 cells. The relationship between virus concentration and plaque count was linear. Addition of 50 μg/ml DEAE-Dextran to the diluent of EAV gave a maximal number of plaques. Differences in end-point titers of infectivity indicated that BHK-21 cells were as sensitive to EAV as rabbit kidney or horse testicle cells.