CHARACTERIZATION OF CHIMERIC FULL-LENGTH MOLECULAR CLONES OF ALEUTIANMINK DISEASE PARVOVIRUS (ADV) - IDENTIFICATION OF A DETERMINANT GOVERNING REPLICATION OF ADV IN CELL-CULTURE

The ADV-G strain of Aleutian mink disease parvovirus (ADV) is nonpatho genic for mink but replicates permissively in cell culture, whereas th e ADV-Utah 1 strain is highly pathogenic for mink but replicates poorl y in cell culture. In order to relate these phenotypic differences to primary genomic features, we constructed a series of chimeric plasmids between a full-length replication-competent molecular clone of ADV-G and subgenomic clones of ADV-Utah 1 representing map units (MU) 15 to 88. After transfection of the plasmids into cell culture and serial pa ssage of cell lysates, we determined that substitution of several segm ents of the ADV-Utah 1 genome (MU 15 to 54 and 65 to 73) within an inf ectious ADV-G plasmid did not impair the ability of these constructs t o yield infectious virus in vitro. Like ADV-G, the viruses derived fro m these replication-competent clones caused neither detectable viremia 10 days after inoculation nor any evidence of Aleutian disease in adu lt mink. On the other hand, other chimeric plasmids were incapable of yielding infectious virus and were therefore replication defective in vitro. The MU 54 to 65 EcoRI-EcoRV fragment of ADV-Utah 1 was the mini mal segment capable of rendering ADV-G replication defective. Substitu tion of the ADV-G EcoRI-EcoRV fragment into a replication-defective cl one restored replication competence, indicating that this 0.53-kb port ion of the genome, wholly located within shared coding sequences for t he capsid proteins VP1 and VP2, contained a determinant that governs r eplication in cell culture. When cultures of cells were studied 5 days after transfection with replication-defective clones, rescue of dimer ic replicative form DNA and single-stranded progeny DNA could not be d emonstrated. This defect could not be complemented by cotransfection w ith a replication-competent construction.