The expression strategy of parvoviruses of the Densovirus genus has as yet not been reported. Clones were obtained from the densonucleosis virus of Galleria mellonella (GmDNV) that yielded infectious virus upon transfection into LD652 cells. Its genome was found to be the longest (6,039 nucleotides [nt]), with the largest inverted terminal repeats (ITRs) (550 nt) among all parvoviruses. The distal 136 nt could be folded into hairpins with flop or flip sequence orientations. In contrast to vertebrate parvoviruses, the gene cassettes for the nonstructural (NS) and structural (VP) proteins were found on the 5 halves of the opposite strands. The transcripts for both cassettes started 23 nt downstream of the ITRs. The TATA boxes, as well as all upstream promoter elements, were localized in the ITRs and, therefore, identical for the NS and VP transcripts. These transcripts overlapped for 60 nt at the 3 ends (antisense RNAs) at 50 m.u. The NS cassette consisted of three genes of which NS2 was contained completely within NS1 but from a different reading frame. Most of the NS transcripts were spliced to remove the upstream NS3, allowing leaky scanning translation of NS1 and NS2, similar to the genes of RNA-6 of influenza B virus. NS3 could be translated from the unspliced transcript. The VP transcript was not spliced and generated four VPs by a leaky scanning mechanism. The 5-untranslated region of the VP transcript was only 5 nt long. Despite the transcription and translation strategies being radically different from those of vertebrate parvoviruses, the capsid was found to have phospholipase A 2 activity, a feature thus far unique for parvoviruses. Densonucleosis viruses (densoviruses or DNVs) are parvoviruses of invertebrates (30).Thus far, about 25 DNVs have been isolated from different species of insects belonging to at least six insect orders, shrimps, and possibly crabs (6, 13, 27). All characterized DNVs package complementary, linear single strands of the DNA into separate virions, as do several parvoviruses of vertebrates (e.g., B19 and AAV [11]), and replicate autonomously. DNVs have little sequence identity with the vertebrate parvoviruses (9), and, consequently, they have been classified as a separate subfamily, the Densovirinae, in the family of Parvoviridae (34). Further studies indicated that there are at least three distinct DNV groups, one genus (Densovirus), exemplified by Junonia coenia DNV (JcDNV) (12, 19), a second genus (Brevidensovirus) by the Aedes (1, 8) and shrimp (27) DNVs, and a third genus (Iteravirus) by the Casphalia DNV (14) and Bombyx mori DNV type 1 (22). Most DNVs remain unclassified, since few are characterized with respect to their molecular biology.The DNV from the greater waxmoth, Galleria mellonella (GmDNV), was the first DNV to be isolated (25). Its particles yield four clearly distinct polypeptide bands upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) (between 90 and 45 kDa and relative amounts increasing in the order (from least to greatest) VP1, VP3, VP...

Bombyx mori densovirus (BmDNV-1), on the basis of the previously reported genome sequence, constitutes by itself a separate genus (Iteravirus) within the Densovirinae subfamily of parvoviruses. Inconsistencies in the genome organization, however, necessitated its reassessment. The genome sequence of new clones was determined and resulted in a completely different genome organization. The corrected sequence also contained conserved sequence motifs found in other parvoviruses. Some amino acids in the highly conserved domain in the unique region of VP1 were shared by critical amino acids in the catalytic site and Ca 2M -binding loop of secreted phospholipase A 2 , such as from snake and bee venoms. Expression of this domain and determination of enzyme activity demonstrated that capsids have a phospholipase A 2 activity thus far unknown to occur in viruses. This viral phospholipase A 2 , which is required shortly after entry into the cell, showed a substrate preference for phosphatidylethanolamine and phosphatidylcholine over phosphatidylinositol.