Direct evidence of in vivo apoptosis of Spodoptera litura larvae was demonstrated by haemocoel inoculation with wild-type Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV) budded virus (BV). In sharp contrast to natural infection, cadavers did not melt, liquefy and melanize. Typical morphological changes of apoptosis in insect haemocytes post-infection, including blebbing of the cell surface, chromatin margination and condensation, vacuolization of the cytoplasm and formation of apoptotic bodies, were observed by light and electron microscopy. Total DNAs extracted from virus-infected haemocytes showed DNA ladders. Cleavage of chromatin DNA by endogenous endonucleases were detected in the cells of most tissues cells, including epithelial cells and fat body cells, using terminal dUTP nick end labelling assays. Virogenic stroma and viral nucleocapsids could be seen in the nuclei of a few haemocytes. Yields of BV and OV (occluded virus) produced from the infected S. litura larvae were much lower than from the infected S. exigua larvae. These data suggest that host apoptotic responses to virus infection reduce AcMNPV spread at the level of the organism and that apoptosis could be host-range limiting factor for baculovirus infections.

Spodoptera litura multicapsid nucleopolyhedrovirus (SpltMNPV) gp41 gene is 993 bp long and the protein encoded by this gene has 6–66% amino acid identities with other known baculovirus GP41 proteins. Slgp41 transcripts were detected from 12 to 96 h post-infection (p.i.) and the mRNA start site was mapped within a consensus baculovirus late promoter sequence (ATAAG). Western blot analysis of extracts from SpltMNPV-infected S. litura cells detected a 41 kDa protein, and this protein was present in the nucleus of infected cells from 12 to 96 h p.i., whereas in the cytoplasm from 24 to 96 h p.i. Structural localization confirmed that SlGP41 is associated with the envelope of occlusionderived virus (ODV). Lectin-binding assay showed that three lectins erythrina cristaglli lectin (ECL), lycopersicon esculentum lectin (LEL), and bandeiraea simlicifolia lectin (BSL) recognizing N-acetylglucosamine were specifically bound to SlGP41. It was proposed that SlGP41 is an O-glycoprotein.

The complete nucleotide sequence of Spodoptera litura nucleopolyhedrovirus (SpltMNPV) Uba256 gene, encoding ubiquitin fused to GP37 protein of 256 amino acids was determined. The first 76 amino acids of the SpltMNPV ubiquitin showed 78-88, 77 and 81-84% amino acid sequence identity to baculovirus, Melanoplus sanguinipes entomopoxvirus and eukaryotes ubiquitins, respectively. The deduced amino acid sequence of SpltMNPV GP37 protein was similar to other baculovirus GP37 proteins and to entomopoxvirus fusolin proteins. The GP37 protein also showed a distant similarity to Pseudaletia separata entomopoxvirus enhancing factor, bacterial chitinase B and chitinbinding protein 1, but the significance of this is unclear. The mRNA start site of Uba256 fusion gene was mapped within a consensus baculovirus late promoter sequence (ATAAG), commonly found for baculovirus late genes. Uba256 transcripts were present from 48 h p.i. and remained detectable until 72 h p.i. Western blot analysis of SpltMNPV-infected Sl-zsu-1 cells revealed that the intact Uba256 was processed to free ubiquitin and GP37 protein. Whereas expression Uba256 gene in Escherichia coli did not result in processing of the fusion protein. Tunicamycin treatment of SpltMNPV-infected cells confirmed that SpltMNPV GP37 protein is N-glycosylated. These findings provide additional information on the evolution of ubi genes and insight into genomic variation in baculoviruses.

The GP37 amino acid sequence of Spodoptera litura multicapsid nucleopolyhedrovirus (SpltMNPV) was compared with other baculovirus GP37, entomopoxvirus fusolin, the enhancing factor of Pseudaletia separata entomopoxvirus, and Alteromonas sp. chitin-binding protein 1. In these proteins, five 'conserved regions' previously reported constitute a chitin-binding domain. SpltMNPV GP37 effectively bound to purified crab shell chitin and the dissociation constant (Kd) for binding was 0.28 mM. Immunofluorescence analysis indicated that SpltMNPV GP37 was located in both cytoplasm and nucleus. Immunoblot analysis revealed that this protein was present in the envelopes of both occlusion body-derived virus and budded virus. Further analysis suggested that GP37 may bind to the chitin component of the peritrophic membrane of S. litura larvae.