Molecular comparisons of in vivo- and in vitro-derived strains of Aleutian disease of mink parvovirus

Abstract

DNA from one cell culture-adapted and two pathogenic strains of Aleutian disease of mink parvovirus (ADV) was molecularly cloned into the vectors pUC18 and pUC19. The DNA from the two pathogenic strains (ADV-Utah I and ADV-Pullman) was obtained from virus purified directly from the organs of infected mink, whereas the DNA from the nonpathogenic ADV-G was derived from cell culture material. The cloned segment from all three viruses represented a 3.55-kilobase-pair BamHI (15 map units) to Hindlll (88 map units) fragment. Detailed physical mapping studies indicated that all three viruses shared 29 of 46 restriction endonuclease recognition sites but that 6 sites unique to the pathogenic strains and 5 sites unique to ADV-G were clustered in the portion of the genome expected to code for structural proteins. Clones from all three viruses directed the synthesis of two ADV-specific polypeptides with molecular weights of approximately 57 and 34 kilodaltons. Both species reacted with sera from infected mink as well as with a monoclonal antibody specific for ADV structural proteins. Because production of these ADV antigens was detected in both pUC18 and pUC19 and was not influenced by isopropyl-lo-D-thiogalactopyranoside (IPTG) induction, their expression was not regulated by the lac promoter of the pUC vector, but presumably by promoterlike sequences found within the ADV DNA. The proteins specified by the clones of ADV-G were 2 to 3 kilodaltons smaller than those of the two pathogenic strains, although the DNA segments were identical in size. This difference in protein molecular weights may correlate with pathogenicity, because capsid proteins of pathogenic and nonpathogenic strains of ADV exhibit a similar difference.

Characterization of chimeric full-length molecular clones of Aleutian mink disease parvovirus (ADV): identification of a determinant governing replication of ADV in cell culture

“…The experiments indicated that a determinant governing the ability of ADV to replicate in CRFK cells mapped to a 531-bp EcoRI-EcoRV fragment spanning nt 2553 to 3084 (Fig. 1) (12,14,15). No construction containing this MU 54 to 65 piece from ADV-Utah 1 was replication competent in CRFK cells.…”

“…SDS-PAGE and immunoblot analysis. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting for ADV proteins were performed as detailed in previous publications (1,15,21,53), but the signal was developed with a chemiluminescent substrate (ECL; Amersham, Chicago, Ill.). This method amplified the sensitivity by at least a factor of 10 over standard chromogenic substrates.…”

Aleutian mink disease parvovirus infection of mink peritoneal macrophages and human macrophage cell lines

“…Membranes were incubated with a 1/100 dilution of ADVinfected adult mink serum and with a 1/4,000 dilution of peroxidase-labeled protein A (Boehringer Mannheim Biochemicals, Indianapolis, Ind.) and developed as previously described (8).…”

Nucleotide sequence analysis of Aleutian mink disease parvovirus shows that multiple virus types are present in infected mink

“…Newborn mink kits were inoculated with 106 adult mink 50% infective doses and killed 10 to 20 days later. Virions were partly purified from lung, liver, and spleen tissues from the mink kits by isopycnic centrifugation in CsCl gradients, and DNA was extracted by the sodium dodecyl sulfate-proteinase K procedure (4,14).…”