Identification of three genes of Helicoverpa armigera single-enveloped nucleopolyhedrovirus and their application in determining HaSNPV-SA phylogeny

Baculovirus phylogeny based on a single gene may not accurately represent baculovirus relatedness. Phylogenetic trees might differ, depending on the genes used. An EcoRI genomic library of a single-capsid nucleopolyhedrovirus of Helicoverpa armigera (HaSNPV) was constructed with the plasmid pBluescript SK. Various clones ranging in size from 14 kb to 0.5 kb were constructed. Three important genes, late expression factor (lef) 8, chitinase and helicase, were identified during random sequence analysis of two clones (EcoRI-H and EcoRI-J), which formed part of the EcoRI plasmid library of the viral genome. Two of these genes, lef-8 and helicase, found to play a role in replication or expression, have been found in all baculovirus genomes sequenced to date. The genome sequence of Plutella xylostella granulovirus (PxGV) does not contain a chitinase gene. For each of the three genes, amino acid sequences were aligned with CLUSTAL X using default parameters and the Blosum matrix. Single nucleotide polymorphisms (SNPs) within each gene of the Helicoverpa SNPVs were tabulated. Trees were constructed using the neighbour-joining algorithm of CLUSTAL X and were rooted on Cydia pomonella GV. The resulting phylogenetic trees were visualized in Treeview. The two data sets of helicase and lef-8 yielded phylogenies supporting the separation of the Nucleopolyhedrovirus (NPV) and Granulovirus (GV) genera and further division of the NPVs into groups I and II. These two trees confirmed the placement of HaSNPV as a group II type NPV. The data set of chitinase incorrectly grouped the HaSNPV as part of the group I NPVs.