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The complete genome sequence was determined for an apparent alphabaculovirus isolated from larval cadavers of the brown tussock moth, Olene mendosa Hübner, collected during an epizootic in Coimbatore, India. The genome was determined to be a circular 142,291 bp molecule, and 147 ORFs and nine homologous regions were annotated for the sequence. Analysis of the sequence confirmed that this virus, Olene mendosa nucleopolyhedrovirus (OlmeNPV), was a member of genus Alphabaculovirus in family Baculoviridae. Phylogenies inferred from nucleotide and amino acid alignments indicated that OlmeNPV was part of a group of viruses that infect moths of genus Lymantria, suggesting that OlmeNPV may have shifted hosts from a Lymantria species to an ancestral Olene species at some point during its evolutionary history. OlmeNPV was most closely related to Lymantria xylina multiple nucleopolyhedrovirus isolate 5 (LyxyMNPV-5). The genomes of OlmeNPV and LyxyMNPV-5 were distinguished not only by differences in ORF content, but by a 27 kbp region of the genome that is inverted in LyxyMNPV-5 relative to OlmeNPV. Pairwise nucleotide distances between OlmeNPV and other Lymantria spp. alphabaculoviruses indicate that OlmeNPV represents a new baculovirus species.

A baculovirus isolate from a USDA Forest Service collection was characterized by electron microscopy and analysis of its genome sequence. The isolate, formerly referred to as Pseudoletia (Mythimna) sp. nucleopolyhedrovirus #7 (MyspNPV#7), was determined by barcoding PCR to derive from the host species Mythimna unipuncta (true armyworm) and was renamed Mythimna unipuncta nucleopolyhedrovirus #7 (MyunNPV#7). The occlusion bodies (OBs) and virions exhibited a size and morphology typical for OBs produced by the species of genus Alphabaculovirus, with occlusion-derived virions consisting of 2-5 nucleocapsids within a single envelope. The MyunNPV#7 genome was determined to be 148,482 bp with a 48.58% G+C nucleotide distribution. A total of 159 ORFs of 150 bp or larger were annotated in the genome sequence, including the 38 core genes of family Baculoviridae. The genome contained six homologous repeat regions (hrs) consisting of multiple copies of a 34-bp imperfect palindrome. Phylogenetic inference from concatenated baculovirus core gene amino acid sequence alignments placed MyunNPV#7 with group II alphabaculoviruses isolated from other armyworm and cutworm host species of lepidopteran family Noctuidae. MyunNPV#7 could be distinguished from other viruses in this group on the basis of differences in gene content and order. Pairwise nucleotide distances suggested that MyunNPV#7 represents a distinct species in Alphabaculovirus. The MyunNPV#7 genome was found to contain two copies of the late expression factor-7 (lef-7) gene, a feature not reported for any other baculovirus genome to date. Both copies of lef-7 encoded an F-box domain, which is required for the function of LEF-7 in baculovirus DNA replication.

A PCR-based method was used to identify and distinguish among 40 uncharacterized nucleopolyhedrovirus (NPV) isolates from larvae of the moth Spodoptera frugiperda that were part of an insect virus collection. Phylogenetic analysis was carried out with sequences amplified from two strongly conserved loci (polh and lef-8) from the 40 isolates in the collection and from eight previously studied S. frugiperda NPV (SfMNPV) isolates. To further distinguish these isolates, analysis was also carried out with sequences from two less-conserved loci, hr4 and hr5. Phylogenetic inference from the sequence data could distinguish among several of the individual isolates and between different groups of isolates from Georgia (USA) and Colombia, South America. A stronger degree of bootstrap support for the phylogenetic trees was obtained with the hr4 and hr5 homologous repeat sequences. Sequencing and phylogenetic analysis detected a relatively high degree of larva-to-larva sequence divergence occurring among isolates of SfMNPV collected from the same field in Missouri, USA. Restriction endonuclease analysis of viral DNA from larvae infected with five isolates from Georgia, Missouri, Louisiana, Florida (USA), and Colombia allowed for further comparison with other previously reported isolates of SfMNPV. Bioassays with these five geographically distinct isolates detected minor differences in virulence. This study highlights the use of PCR to rapidly distinguish and characterize large numbers of historical baculovirus isolates from the same host using minimal quantities of material, and the use of sequences from homologous repeat regions to distinguish closely related isolates of the same NPV species.