• BMB Reports
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• v.40 no.1
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• pp.65-71
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• 2007

The Helicoverpa armigera nucleopolyhedrovirus (HearNPV) ORF80 (ha80) has 765 bp encoding a protein with approximately 254 amino acids and a predicted molecular weight of 30.8 kDa. Homologues of ha80 are found in most baculovirus sequences, including those from lepidopteran NPVs, lepidopteran granuloviruses (GVs), hymenopteran baculoviruses, and one dipteran baculovirus, yet their functions remain unclear. In this study we characterized ha80, and showed that it was transcribed late in infected host cells (HzAM1). The product of ha80 was a 31 kDa protein that was not a structural protein of budded virus (BV) or occlusion-derived virus (ODV) particles. Ha80 was first detected in the cytoplasm of infected HzAM1 cells at 12 h p.i., and was observed in the nucleus at later stages of infection, suggesting that it may be involved in transporting viral proteins into the host cell nucleus or play its roles in the nucleus.

• BMB Reports
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• v.45 no.12
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• pp.730-735
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• 2012

p13 gene was first described in Leucania separata multinuclear polyhedrosis virus (Ls-p13) several years ago, but the function of P13 protein has not been experimentally investigated to date. In this article, we indicated that the expression of p13 from Heliothis armigera single nucleocapsid nucleopolyhedrovirus (Ha-p13) was regulated by both early and late promoter. Luciferase assay demonstrated that the activity of Ha-p13 promoter with hr4 enhancer was more than 100 times in heterologous Sf9 cells than that in nature host Hz-AM1 cells. Both Ls-P13 and Ha-P13 are transmembrane proteins. Confocal microscopic analysis showed that both mainly located in the cytoplasm membrane at 48 h. Results of RNA interference indicated that Ha-p13 was a killing-associated gene for host insects H. armigera. The AcMNPV acquired the mentioned killing activity and markedly accelerate the killing rate when expressing Ls-p13. In conclusion, p13 is a killing associated gene in both homologous and heterologous nucleopolyhedrovirus.

• International Journal of Industrial Entomology and Biomaterials
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• v.4 no.2
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• pp.137-141
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• 2002

An experimental study was undertaken to quantify the effects of insect hormones on the replication of nucleopolyhedrovirus (NPV). The results demonstrated that TCID/ sub 50/ at 72 h post-infection (hpi) rose systematically from 0.55$\times$10$^{8}$ /m1, for untreated cells, up to 1.67$\times$10$^{8}$ / ml at 3$\mu$g/ml, then dropped down to 1.45$\times$10$^{8}$ /m1 at 4 $\mu$g/ml, by adding ecdysone to the culture medium for Bm-N cells infected with a wild-type Bambyx mori. nucleopolyhedrovirus (BmNPV). The optimum enhancement of about 3 times on budded virus (BV) titer at 72 hpi was given at 3 $\mu$g/ml of ecdysone. While the polyhedra number had no obvious variation within the range of concentrations from 0 to 4 $\mu$g/ml. By addition of juvenile hormone analogue (JHA) into the media with this concentration range, the BmNPV TCID/ sub 50/ and polyhedra number at 72 hpi did not show significant changes. Also, the addition of either 3 $\mu$g/ml of ecdysone or 3 $\mu$g/ml of JHA to the culture media did not appear to affect the TCID/ sub 50/ and polyhedra number significantly in infected Sf-21 cells with the autographa californica nucleopolyhedrovirus (AcMNPV).

• International Journal of Industrial Entomology and Biomaterials
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• v.23 no.1
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• pp.123-128
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• 2011

1-Deoxynojirimycin (DNJ) is an alkaloid that is found at relatively high concentrations in mulberry leaf and tissues of the silkworm, $Bombyx$ $mori$. DNJ is a well known inhibitor of ${\alpha}$-glucosidase, an enzyme that is involved in the early stages of the $N$-linked glycoprotein synthesis pathway. ${\alpha}$-Glucosidase activity in the cell extract from $B.$ $mori$-derived Bm5 cells showed approximately 40-fold less sensitivity to DNJ than ${\alpha}$-glucosidase activity in the cell extract from $Spodoptera$ $frugiperda$-derived Sf9 cells. The replication of $B.$ $mori$ nucleopolyhedrovirus (BmNPV) was not inhibited when it was propagated in BmN cells that were grown in medium containing up to 10 mM DNJ. In contrast, the replication of $Autographa$ $californica$ multiple NPV (AcMNPV) was reduced by 67% when it was propagated in Sf9 cells that were grown in medium containing 10 mM DNJ. The viability of Bm5 and Sf9 cells that were grown in medium containing up to 10 mM DNJ was not affected. Our results suggested that the reduced replication of AcMNPV was the result of the higher sensitivity of ${\alpha}$-glucosidase activity in Sf9 cells to DNJ.

• Journal of Microbiology
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• v.45 no.2
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• pp.133-138
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• 2007

This study addresses the susceptibility of Spodoptera frugiperda (Sf9 and Sf21), Trichoplusia ni (Hi5), and S. exigua (Se301) cells to the Bombyx mori nucleopolyhedrovirus (BmNPV). Although these cells have classically been considered nonpermissive to BmNPV, the cytopathic effect, an increase in viral yield, and viral DNA synthesis by BmNPV were observed in Sf9, Sf21, and Hi5 cells, but not in Se301 cells. Very late gene expression by BmNPV in these cell lines was also detected via ${\beta}-galactosidase$ expression under the control of the polyhedrin promoter. Sf9 cells were most susceptible to BmNPV in all respects, followed by Sf21 and Hi5 cells in decreasing order, while the Se301 cells evidenced no distinct viral replication. This particular difference in viral susceptibility in each of the cell lines can be utilized for our understanding of the mechanisms underlying the host specificity of NPVs.

• Journal of Microbiology
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• v.44 no.1
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• pp.77-82
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• 2006

The tea looper caterpillar, Ectropis obliqua, is one of the major pests of tea bushes. E. obliqua single-nucleocapsid nucleopolyhedrovirus (EcobSNPV) has been used as a commercial pesticide for biocontrol of this insect. However only limited genetic analysis for this important virus has been done up to now. EcobSNPV was characterized in this study. Electron microscopy analysis of the occlusion body showed polyhedra of 0.7 to $1.7\;{\mu}m$ in diameter containing a single nucleocapsid per envelope of the virion. A 15.5 kb genomic fragment containing EcoRI-L, EcoRI-N and HindIII-F fragments, was sequenced. Analysis of the sequence revealed that the fragment contained eleven potential open reading frames (ORFs): lef-1, egt, 38.7k, rrl, polyhedrin, orfl629, pk-1, hoar and homologues to Spodoptera exigua multicapsid NPV (SeMNPV) ORFs 15, 28, and 29. Gene arrangement and phylogeny analysis suggest that EcobSNPV is closely related to the previously described Group II NPV. Bioassays on lethal concentration $(LC_{50}\;and\;LC_{90})$ and lethal time $(LT_{50}\;and\;LT({90})$ were conducted to test the susceptibility of E. obliqua larvae to the virus.

• International Journal of Industrial Entomology and Biomaterials
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• v.15 no.2
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• pp.131-135
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• 2007

Open reading frame 4 of Bombyx mori nucleopolyhedrovirus (BmNPV), designated as Bm4, is a gene whose function is completely unknown. With the recently developed BmNPV bacmid and a modified pFastBac1 whose polyhedrin promoter was replaced with ie1 promoter, a recombinant bacmid expressing Bm4-EGFP fusion protein under the control of ie1 promoter in BmN cells was successfully constructed. The result not only showed that the polyhedrin promoter can be replaced efficiently with other promoters to direct the expression of foreign gene in BmN cells by using Bac-to-Bac/BmNPV baculovirus expression system but also laid the foundation for rescue experiment of Bm4 deletion mutant due to the ability of ie1 promoter to direct gene expression throughout the infection cycle.

• BMB Reports
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• v.38 no.3
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• pp.354-359
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• 2005

Open reading frame 29 (ha29) is a gene specific for Helicoverpa armigera single-nucleocapsid nucleopolyhedrovirus (HearSNPV). Sequence analyses showed that the transcription factor Tfb2 motif, bromodomain and Half-A-TPR (HAT) repeat were present at aa 66-82, 4-76, 55-90 of the Ha29 protein respectively. The product of Ha29 was detected in HearSNPV-infected HzAM1 cells at 3 h post-infection. Western blot analysis using a polyclonal antibody produced by immunizing a rabbit with purified GST-Ha29 fusion protein indicates that Ha29 is an early gene. The size of Ha29 product in infected HzAM1 cells was about 25 kDa, which was larger than the presumed size of 20.4 kDa. Tunicamycin treatment of HearSNPV-infected HzAM1 cells suggested that the Ha29 protein is N-glycosylated. Fluorescent confocal laser scanning microscope examination, and Western blot analysis of purified budded virus (BVs), occlusion-derived virus (ODVs), cell nuclear and cytoplasmic fraction, showed that the Ha29 protein was localized in the nucleus. Our results suggested that ha29 of HearSNPV encodes a non-structurally functional protein that may be associated with virus gene transcription in Helicoverpa hosts.

• International Journal of Industrial Entomology and Biomaterials
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• v.22 no.2
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• pp.75-82
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• 2011

To elucidate the novelty of Hyphantria cunea nucleopolyhedrovirus (HcNPV) isolated in Korea, polyhedrin and inhibitor of apoptosis (iap) gene structures were determined and analyzed. The analysis of HcNPV polyhedrin showed a little difference with 97.6% at the nucleotide level but no difference at the amino acid level when compared with that of previously reported H. cunea NPV (HycuNPV). On the other hand, iap genes showed variable differences with 89.0-99.6% nucleotide and 90.0-99.6% amino acid sequence identities. Especially, the 5' and 3' non-coding flanking sequences of iap1 gene had lower degree of identity with those of HycuNPV. Although the phylogenetic analyses using polyhedrin and iap genes showed that HcNPV is closely related with HycuNPV, we could provide that HcNPV is a novel isolate having novel gene structures.

• BMB Reports
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• v.39 no.3
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• pp.263-269
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• 2006

In the genome of Helicoverpa armigera single-nucleocapsid nucleopolyhedrovirus, open reading frame 39 (Ha39) is the only gene predicted to encode an RNA recognition protein. Computer analysis revealed that Ha39 homologues were found in 15 NPVs, but not in GVs. Its transcripts were detected from 3 through 72 hours post infection (h p.i.) using RT-PCR and Northern blot analysis. The protein was detected in infected-cell lysates from 6 h p.i. Western blot assay of ODV and BV preparations revealed that Ha39 encodes a structural protein associated with BVs. Additionally, immunofluorescence microscopy demonstrated that the protein was present within cytoplasm in virus-infected cells, but not in the nuclear region.