Abstract
To characterize the infection of Bombyx mori nucleopolyhedrovirus (BmNPV) in Spodoptera frugiperda (Sf9) cells, several relevant parameters to viral replication were examined. Sf9 cells showed no clear overall cytopathology and proliferated continuously following inoculation with BmNPV. However, careful examination under a microscope revealed a small number of cells with polyhedra. Also the immunoblot analysis showed significant but low levels of BmNPV polyhedrin expression in BmNPV-inoculated Sf9 cells. Examination with a recombinant BmNPV, BmΔpolh-lacZ that possessed lacZ driven by polyhedrin gene (polh) promoter, further indicated that the number of Sf9 cells expressing lacZ increased with the increase in viral multiplicities of inoculation. Slot-blot hybridization analysis detected low levels of viral DNA replication and northern blot analysis showed that transcription of an immediate early viral gene, ie-1, was restricted strikingly in BmNPV-inoculated Sf9 cell cultures, suggesting that early events leading to productive viral infection occurred in only a limited number of BmNPV-inoculated Sf9 cells. Comparative infectivity assay between inoculation of BmNPV budded virions (BVs) and transfection of BmNPV DNA revealed that infection efficiency was higher in the transfection than in the inoculation. Viral adsorption assay showed that BmNPV BVs were able to be adsorbed efficiently by Sf9 cells. These results suggest that BmNPV replication in majority of Sf9 cells is restricted mainly at the step prior to the expression of immediate early viral genes, that includes viral entry into the cells through endocytotic pathway and nucleocapsid uncoating in the nuclei.
