Mechanisms contributing to the virus persistence in Aleutian disease.

In this review published results and further studies concerning the persistence of Aleutian disease virus (ADV) isolate SL3 are presented. By Southern blot and in situ hybridization with strand-specific RNA probes focal replication of ADV-DNA was demonstrated in spleen, mesenteric lymph nodes, sporadically in mononuclear cells of the peripheral blood and bone marrow cells. These findings further support the concept of the lymphotropism of ADV. All cell culture-adapted ADV strains appear to have a ts-defect. Our in vitro studies indicate that the ADV isolate G(orham) induced the synthesis of comparable amounts of viral replicative DNA and viral proteins VP1 and VP2 at the non-permissive temperature of 37 degrees C. However, the viral progeny DNA synthesis was about threefold less at 37 degrees C compared to the permissive temperature of 32 degrees C. These findings suggest that the reduced level of viral progeny DNA at 37 degrees C accounts for the reduced production of infectious ADV. Finally, we provided experimental evidence that the apparent lack of neutralizing antibodies in AD is due to the masking of critical viral epitopes by cellular phospholipids.

Aleutian disease virus in B and T lymphocytes from blood and spleen and in bone marrow cells from naturally infected mink.

In the nuclei of 4% of peripheral blood or spleen mononuclear cells (MNC), Aleutian disease virus(ADV)-specific antigens were found by a direct immunofluorescence test. The MNC were further fractionated by nylon wool, affinity chromatography using Staphylococcus aureus protein, or Percoll gradient techniques. ADV and specific antigens were detected in MNC fractions enriched in either the B or T lymphocytes. In the bone marrow, up to 40% antigen-positive cells were demonstrated over a period of 15 months. These findings were confirmed by the detection of infectious virus in the MNC of blood and spleen and in bone marrow cells. Adherent cells from mink and control cells from ADV-negative ferrets were negative in both tests. These findings indicate that ADV exhibits a lymphotropism and can persist in the B- and T-cell fractions from ADV-infected mink over a long period of time. Furthermore, co-cultivation of mink MNC and bone marrow cells with the CCC clone 81 cells was shown to be reproducible method for the detection of ADV in persistently infected mink.

Propagation of Aleutian disease parvovirus in cell line CCC clone 81.

Aleutian disease parvovirus (ADV), mutant Gorham of the Utah-1 strain, was grown and comparatively assayed in feline cell lines CRFK and CCC clone 81 at 31.8 degrees C. The maximum virus titres as determined by a fluorescent focus assay were found to be about 10(5) FFU/ml in CRFK at day 6 p. i. and 10(6) FFU/ml at day 4 p. i. in CCC clone 81 cells. Shifting of the incubation temperature from 31.8 to 37 degrees C led to a reduced virus production after three passages. The synchronization of the CCC clone 81 cells by 1 X 10(-3) M hydroxyurea followed by infection with low (less than or equal to 0.8) multiplicities of infection (MOI) did not significantly influence the virus titres. Several mammalian cell lines such as MiCl 1 (S+L-), Mv1-Lu, 64F3 clone 7 and FEF or fish cell lines such as BB and CHSE 114 developed abortive infections after inoculation with the temperature-sensitive mutant Gorham of the ADV strain Utah-1 (ADV-G). Three new isolates designated ADV-Sl1--ADV-Sl3 were isolated from spleen and blood lymphocytes and bone marrow cells of ADV-infected mink and were adapted to grow in CCC clone 81 cells at 31.8 degrees C with virus titres between 10(4) and 10(4.7) FFU/ml. ADV particle populations varying in their bouyant density between 1.32, 1.36 and 1.43 g/ml were isolated from infected cells and culture supernatants. By protein blotting and immunodetection two major protein components with apparent M. W. of 85 and 75 KD and three minor polypeptides of 33, 28.9 and 27.5 KD were detected.