• Journal of Sericultural and Entomological Science
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• v.40 no.2
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• pp.111-116
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• 1998

To construct transformed Bm5 cells, Autographa californica nuclear polyhedrosis virus (AcNPV)IE1 gene, an immediate early viral gene was firstly used in this study. AcNPV IE1 gene, which shares on 95.3% uncleotide sequence homology with Bombyx mori nuclear polyhedrosis virus (BmNPV) IE1 gene, was isolated and cloned into pBluescript. Neomycin gene from pKO-neo was inserted under the control of the IE1 promoter to yield pAcIE1-neo. The plasmid pAcIE1-neo was transfected into Bm5 or Sf9 cells, and neomycin-resistant cells were selected in TC100 medium containing 10% fetal bovine serum (FBS) and 1 mg/$m\ell$ G418 for two weeks. Individual clones were picked and each was amplified for further characterization. The genomic DNA from neomycin-resistnt cells was isolated and characterized by PCR using AcNPV IE1 gene-specific primers and by Southern blot analysis using neomycin gene probe. We concluded that AcNPV IE1 gene was functional in B. moridrived Bm5 cells as well as Spodaptera frugiperda-derived Sf9 cells to produce stably-transformed insect cells.

• Korean journal of applied entomology
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• v.59 no.4
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• pp.451-454
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• 2020

Sericulture is a main insect industry in Sangju (Kyungpook, Korea). This study reports an occurrence of a viral disease in the sericulture farms in 2020. More than 20% silkworm larvae (Bombyx mori) suffered diarrhea and melted tissues with pathogenic lethality at 4th or 5th instars. PCR diagnosis showed a positive response against B. mori nucleopolyhedrosis virus (BmNPV) infection. Tissue extract of the infected larvae was applied to healthy larvae by a leaf-dipping method and exhibited the same viral symptoms. The viral extract was used to be overlaid on Sf9 cells. The infected Sf9 cells showed polyhedra in the cytoplasm. These results indicate that the silkworm larvae reared in the sericulture farms in Sangju were infected with BmNPV.

• International Journal of Industrial Entomology and Biomaterials
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• v.12 no.2
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• pp.95-100
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• 2006

Central Sericultural Research and Training Institute, Mysore have evolved several highly productive bivoltine hybrids which can produce international grade raw silk. Among them $CSR2{\times}CSR4,\;CSR2{\times}CSR5,\;CSR3{\times}CSR6,\;CSR17{\times}CSR16,\;CSR18{\times}CSR19$ and $CSR12{\times}CSR6$ are being popularized in the field. There is a minimum difference in their economic characters but they appear to differ in survival. Though they are productive under high input management conditions, they are very susceptible to different diseases under normal rearing practices. No systematic attempts have been made to test their susceptibility status / resistance. Thus the present study is a modest attempt to screen the above six productive bivoltine hybrids to two important pathogens viz., Bombyx mori Nuclear Polyhedrosis Virus (BmNPV) and Bombyx mori Infectious Flacherie Virus (BmIFV) along with existing hybrid, $KA{\times}NB4D2$ to assess their susceptibility / resistance. The results shows that the productive hybrid $CSR2{\times}CSR4$ is the most resistant to BmNPV and it is suggested by its highest $LC_{50}$ value followed by $CSR12{\times}CSR6,\;KA{\times}NB4D2,\;CSR3{\times}CSR6,\;CSR17{\times}CSR16,\;CSR18{\times}CSR19,\;CSR2{\times}CSR5$. Based on the $LC_{50}$ value and $LT_{50}$ values for BmIFV, the hybrid $KA{\times}NB4D2$ was found to be the most resistant (1st position) one followed by $CSR3{\times}CSR6$ (2nd position) $CSR2{\times}CSR$ (3rd position) and $CSR12{\times}CSR6$ (4th position) $CSR17{\times}CSR16$, $CSR18{\times}CSR19$ (5th position) and $CSR2{\times}CSR5$ being the least. The response of 7 bivoltine hybrids to both the pathogens BmNPV and BmIFV indicates that, the hybrids $CSR2{\times}CSR4$, $CSR12{\times}CSR6$ and $KA{\times}NB4D2$ were found to be the most resistant when compared to others. Further, $KA{\times}NB4D2$ being less productive hybrid with a shell ratio of 20.08%, the other two hybrids $CSR2{\times}CSR4$ (Cocoon shell ratio, 21.44%) and $CSR12{\times}CSR6$ (cocoon shell ratio, 23.45%) can be considered to be most productive with superior quality cocoon and resistant to both BmNPV and BmIFV pathogens. The overall study indicated that the hybrid $CSR2{\times}CSR5$ is the most susceptible hybrid to both the pathogens.

• 한국생물공학회:학술대회논문집
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• 2003.04a
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• pp.315-319
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• 2003

A novel method is developed to yield virus titers in 10 h, is easy to .perform using 96-well plates, and applicable to both any Autographa californica nucleopolyhyderovirus (AcNPV) and Bombyx mori nucleopolyhedrovirus (BmNPV)-based recombinant baculovirus. This assay uses an antibody to a DNA-binding protein to detect the infected cells via immune-staining. The titer is determined by counting foci produced due to infection of virus under a fluorescent microscopy. The required incubation period was shortened considerably because infected cells expressed viral antigens at the post infection time of 4 h. Therefore, 10 hours were enough to estimate the virus titer including virus infection time, insect cell culture, and estimation of virus titer.

• International Journal of Industrial Entomology and Biomaterials
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• v.8 no.2
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• pp.169-174
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• 2004

Bombyx mori nucleopolyhedrovirus(BmNPV) ecdysteroid UDP-glucosyltransferase gene (egt) promoter fragments of different lengths were amplified from BmNPV ZJ-8 genomic DNA by PCR. Reporter plasmids pBmegt542-luc, pBmegt309-luc and pBmegtl59-luc with luciferase (lue) driven by egt promoters were constructed. Both in vitro and in vivo expressions showed that BmNPV egt promoter activity requires the transactivation of viral factor(s), and expression of luc was detected earliest at 24 hrs post infection (pi). BmNPV ZJ-8 homologous region 3 (hr3) increased the expression of luc by over 1,600-fold. Molting hormone of 1.0 - 2.0 $\mu\textrm{g}$/$m\ell$ can dramatically down regulate expression of luc. Juvenile hormone analogue of 0.5-2.0 ${\mu}g$/$m\ell$ increased expression of luc by 145.8% to 75.7%. Deletion assay revealed that the promoter fragment of 159 bp contains the basal promoter structure; Promoter fragments of 309 bp and 542 bp showed similar but much higher transcriptional activities than that of 159 bp, suggesting that nucleotide from -159 to -309 nt upstream the translation initiation site harbors the main cis-acting elements.

• International Journal of Industrial Entomology and Biomaterials
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• v.2 no.2
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• pp.149-153
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• 2001

The influence of baculovirus Bombyx mori Nuclear Polyhedrosis virus (BmNPV) infection on intermediary metabolic pathways in silkworm Bombyx mori L. was investigated. Studies revealed that NADP-linked malate dehydrogenase activity in hemolymph of infected silkworms at 96 hrs post infection (p.i.) with visible symptoms of infection was enhanced in comparison to healthy larvae of the same age. Also, NADP-dependent MDH activity was significantly lower in fat body cytosol of infected larvae at 96 hrs p.i. when compared to healthy larvae. Similarly, some biometabolic parameters like growth, protein content and cholesterol titer were observed to be influenced by baculovirus infection. While the growth of infected larvae was significantly retardedi protein content was also drastically reduced in both hemolymph and fat body tissues. Cholesterol titers however, was enhanced in infected larvae. The results observed herein point to a significant change in the normal biochemical and biometabolic pathways required for growth and development following BmNPV infection.

• Journal of Microbiology and Biotechnology
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• v.11 no.4
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• pp.585-591
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• 2001

To compare the p10 promoter activity of Bombyx mori nucleopolyhedrovirus (BmNPV)K1 and K4, recombinant viruses Bm101-LacZ and Bm104-LacZ with a lacZ gene under the control of each p10 promoter were constructed. The $\beta$-galactosidase activity due to Bm101-LacZ was about 5.5- and 1.1-fold higher than that due to Bm104-LacZ and BmK1-LacZ, respectively. expressing ${\beta}$-galactosidase under the control of a polyhedrin promoter. The recombinant virus BmK1-104LacZ with the same genome structure as Bm101-LacZ, except for a p10 promoter region, produced a similar ${\beta}$-galactosidase activity to that due to Bm104-LacZ and 5.5-fold lower than that due to Bm101-LacZ. The virus yield, expression level of polyhedrin, and polyhedra productivity for each recombinant virus was almost similar. These results suggested that the difference in the expression level of ${\beta}$-galactosidase resulted from a difference in the p10 promoter regions, and that an expression vector using the p10 promoter of BmNPV-K1 could be usefully exploited in the mass production of foreign proteins with silkworm larvae by means of oral ingestion.

• Korean journal of applied entomology
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• v.30 no.2
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• pp.144-149
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• 1991

Biochemical characteristics of Spodoptera uigua nuclear polyhedrosis virus (SeNPV) isolated in Jinju were studied. SeNPV contained a number of nucleocapsids within a viral envelope embeded in polyhedra. The polyhedral protein of SeNPV was composed of a single polypeptide with a M. W. of 30kd. Double-immunodiffusion test showed that the polyhedral protein of SeNPV had common antigenic determinants with SINPV and BmNPV. Virion proteins of SeNPV were resolved into 49 polypeptides by silver staining after SDS-PAGE. The approximate genome size of SeNPV by restriction endonuclease analysis was 1l0kb.

• 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 Sericultural and Entomological Science
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• v.40 no.1
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• pp.60-62
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• 1998

To understand expression of some early and late genes of Bombyx mori nuclear polyhedrosis virus (BmNPV) in the B. mori-derived BmN cell line, the transcripts were analyzed by polymerase chain reaction with synthetic primers. After infection, the transcript of early genes, which include p35, IE1 and helicase p143, was immediately detected in the infected cells. In addition, the transcript of late genes, which include p10 and polyhedrin, was also detected in just-infected cells. In conclusion, our results revealed that transcripts of early and late genes of BmNPV are immediately expressed from the cells after infection.