Complete sequence, analysis and organization of the Orgyia leucostigma nucleopolyhedrovirus genome.

The complete genome of the Orgyia leucostigma nucleopolyhedrovirus (OrleNPV) isolated from the whitemarked tussock moth (Orgyia leucostigma, Lymantridae: Lepidoptera) was sequenced, analyzed, and compared to other baculovirus genomes. The size of the OrleNPV genome was 156,179 base pairs (bp) and had a G+C content of 39%. The genome encoded 135 putative open reading frames (ORFs), which occupied 79% of the entire genome sequence. Three inhibitor of apoptosis (ORFs 16, 43 and 63), and five baculovirus repeated ORFs (bro-a through bro-e) were interspersed in the OrleNPV genome. In addition to six direct repeat (drs), a common feature shared among most baculoviruses, OrleNPV genome contained three homologous regions (hrs) that are located in the latter half of the genome. The presence of an F-protein homologue and the results from phylogenetic analyses placed OrleNPV in the genus Alphabaculovirus, group II. Overall, OrleNPV appears to be most closely related to group II alphabaculoviruses Ectropis obliqua (EcobNPV), Apocheima cinerarium (ApciNPV), Euproctis pseudoconspersa (EupsNPV), and Clanis bilineata (ClbiNPV).

Sequence analysis of the Choristoneura occidentalis granulovirus genome.

The genome of the Choristoneura occidentalis granulovirus (ChocGV) isolated from the western spruce budworm, Choristoneura occidentalis, was sequenced completely. It was 104,710 bp long, with a 67.3% A+T content and contained 116 potential open reading frames (ORFs) covering 88.4% of the genome. Of these, 29 ORFs were conserved in all fully sequenced baculovirus genomes, 30 were GV-specific, 53 were present in some nucleopolyhedroviruses (NPVs) and/or GVs, three were common to ChocGV and Choristoneura fumiferana GV (ChfuGV) and one was so far unique. To date, ChocGV is the only GV identified that contains a homologue of the apoptosis inhibitor protein P35/P49, present in some group I NPVs. It is also the first GV without a Xestia c-nigrum GV ORF 26 homologue. Five homologous regions (hrs)/repeat regions, lacking typical NPV hr palindromes were identified. ChocGV hrs were similar to each other but not to other GV hrs. A 1.8 kb repeat region with a high A+T content (81%) and multiple repeats of 21-210 bp was found between choc36 and 37. This area resembled the non-homologous region origin of DNA replication (non-hr ori) identified in Cryptophlebia leucotreta GV (CrleGV) and Cydia pomonella GV (CpGV). Based on the mean amino acid identities of homologous proteins, ChocGV was closest to fully sequenced genomes CpGV (52.3%) and CrleGV (52.1%). The closest amino acid identity was to individual ORFs from the partially sequenced ChfuGV genome (97.2% in 38 ORFs). Phylogenetic analysis placed ChocGV in a clade with CrleGV and CpGV.

Genomic comparison of Neodiprion sertifer and Neodiprion lecontei nucleopolyhedroviruses and identification of potential hymenopteran baculovirus-specific open reading frames.

Genomic comparison of Neodiprion sertifer nucleopolyhedrovirus (NeseNPV) and Neodiprion lecontei nucleopolyhedrovirus (NeleNPV) showed that the hymenopteran baculoviruses had features in common and were distinct from other, fully sequenced lepidopteran and dipteran baculoviruses. Their genomes were small in size (86,462 and 81,755 bp, respectively), had low G+C contents (33.8 and 33.3 mol%, respectively) and contained fewer open reading frames (ORFs) (90 and 89, respectively) than other baculoviruses. They shared 69 ORFs (48.6% mean amino acid identity overall), 43 of which were previously identified baculovirus homologues. The remaining shared ORFs could be common to other baculoviruses, but low amino acid identities precluded identifying them as such. Some may also be unique to hymenopteran baculoviruses. These included a trypsin-like protease, a zinc-finger protein, regulator of chromosome condensation proteins, a densovirus capsid-like protein and a phosphotransferase. Structural analysis, the presence of conserved domains and phylogenetic studies suggested that some of these ORFs may be functional and could have been transferred horizontally from an insect host. ORFs found only in NeseNPV and NeleNPV may play a role in host specificity and/or tissue tropism, as hymenopteran baculoviruses are restricted to the midgut. The genomes were basically collinear, but contained non-syntenic regions (NSRs) with large numbers of repeats between their polyhedrin and dbp genes. They differed from each other in the number of ORFs and the G+C content of their NSRs and the presence of homologous regions in the NeseNPV genome. NeleNPV also had a short inversion relative to NeseNPV. NeseNPV contained 21 ORFs not found in NeleNPV and NeleNPV had 20 ORFs not found in NeseNPV.

Analysis of the Choristoneura fumiferana nucleopolyhedrovirus genome.

The double-stranded DNA genome of Choristoneura fumiferana nucleopolyhedrovirus (CfMNPV) was sequenced and analysed in the context of other group I nucleopolyhedroviruses (NPVs). The genome consists of 129,593 bp with a G + C content of 50.1 mol%. A total of 146 open reading frames (ORFs) of greater than 150 bp, and with no or minimal overlap were identified. In addition, five homologous regions were identified containing 7-10 repeats of a 36 bp imperfect palindromic core. Comparison with other completely sequenced baculovirus genomes revealed that 139 of the CfMNPV ORFs have homologues in at least one other baculovirus and seven ORFs are unique to CfMNPV. Of the 117 CfMNPV ORFs common to all group I NPVs, 12 are exclusive to group I NPVs. Overall, CfMNPV is most similar to Orgyia pseudotsugata MNPV based on gene content, arrangement and overall amino acid identity. Unlike other group I baculoviruses, however, CfMNPV encodes a viral enhancing factor (vef) and has two copies of p26.

Gene organization and sequencing of the Choristoneura fumiferana defective nucleopolyhedrovirus genome.

Two distinct nucleopolyhedrovirus species of the eastern spruce budworm, Choristoneura fumiferana, exist in a symbiont-like relationship. C. fumiferana defective nucleopolyhedrovirus (CfDEFNPV) only infects C. fumiferana larvae per os in the presence of C. fumiferana nucleopolyhedrovirus Ireland strain (CfMNPV), but is infective when injected into the haemolymph. CfDEFNPV synergizes CfMNPV in per os infections and CfMNPV is always the predominant progeny. This study was undertaken to report the genomic makeup and organization of CfDEFNPV in an attempt to identify its defect and understand its synergistic role. The genome was mapped, sequenced, characterized and compared to other baculoviruses. The CfDEFNPV genome was 131,160 nt long with 149 putative open reading frames (ORFs) and a G + C content of 45.8 mol%. Homologues of all 62 conserved lepidopteran baculovirus genes were found including those implicated in per os infectivity, p74, per os infectivity factor (pif) and pif-2. Although no obvious deletions were observed to explain the defect, two ORFs, Cfdef79 and Cfdef99 (inhibitor of apoptosis-4), contained potential deletions. Cfdef50 (late expression factor-10)/Cfdef51 (vp1054) and Cfdef76/Cfdef77 (telokin-like protein) had large overlaps and a potential homologue to ac105/he65 was split. Four baculovirus repeat ORFs were present, as were two unique genes, but no enhancins were identified. CfDEFNPV contained 13 homologous regions, each with one to five palindromes. Comparison with fully sequenced baculovirus genomes identified CfDEFNPV as a group I NPV with the closest average amino acid identity to Epiphyas postvittana NPV, followed by Orgyia pseudotsugata MNPV and CfMNPV, with its closest matches being to individual Anticarsia gemmatalis MNPV gene sequences.

Sequence and organization of the Neodiprion lecontei nucleopolyhedrovirus genome.

All fully sequenced baculovirus genomes, with the exception of the dipteran Culex nigripalpus nucleopolyhedrovirus (CuniNPV), have previously been from Lepidoptera. This study reports the sequencing and characterization of a hymenopteran baculovirus, Neodiprion lecontei nucleopolyhedrovirus (NeleNPV), from the redheaded pine sawfly. NeleNPV has the smallest genome so far published (81,755 bp) and has a GC content of only 33.3%. It contains 89 potential open reading frames, 43 with baculovirus homologues, 6 identified by conserved domains, and 1 with homology to a densovirus structural protein. Average amino acid identity of homologues ranged from 19.7% with CuniNPV to 24.9% with Spodoptera exigua nucleopolyhedrovirus. The conserved set of baculovirus genes has dropped to 29, since NeleNPV lacks an F protein homologue (ac23/ld130). NeleNPV contains 12 conserved lepidopteran baculovirus genes, including that for DNA binding protein, late expression factor 11 (lef-11), polyhedrin, occlusion derived virus envelope protein-18 (odv-e18), p40, and p45, but lacks 21 others, including lef-3, me53, immediate early gene-1, lef-6, pp31, odv-e66, few polyhedra 25k, odv-e25, protein kinase-1, fibroblast growth factor, and ubiquitin. The lack of identified baculovirus homologues may be due to difficulties in identification, differences in host-virus interactions, or other genes performing similar functions. Gene parity plots showed limited colinearity of NeleNPV with other baculoviruses, and phylogenetic analysis indicates that NeleNPV may have existed before the lepidopteran nucleopolyhedrovirus and granulovirus divergence. The creation of two new Baculoviridae genera to fit hymenopteran and dipteran baculoviruses may be necessary.