• International Journal of Industrial Entomology and Biomaterials
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• v.29 no.2
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• pp.145-152
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• 2014

The variation in the level of immune response related gene expression in silkworm, Bombyx mori following infection with Bombyx mori nucleopolyhedrovirus (BmNPV) was analyzed at different time intervals. The occlusion bodies of BmNPV orally inoculated to the two most divergent silkworm races viz., Sarupat (resistant to BmNPV infection) and CSR2 (susceptible to BmNPV infection) were subjected to oral BmNPV inoculation. The expression profile of gp 41 gene of BmNPV in the Sarupat and CSR2 races revealed that the virus could invade the midguts of both susceptible and resistant races. However, its multiplication was significantly less in the midgut of resistant race, while, in the susceptible race, the viral multiplication reached maximum level within 12 h. These findings indicate that potential host genes are involved in the inhibition of viral multiplication within larval midgut. The immune response genes arylphorin, cathepsin B, gloverin, lebocin, serpin, Hsp 19.9, Hsp 20.1, Hsp 20.4, Hsp 20.8, Hsp 21.4, Hsp 23.7, Hsp 40, Hsp 70, Hsp90 revealed differential level of expression on NPV infection. The gloverin, serpin, Hsp 23.7 and Hsp 40 genes are significantly up-regulated in the resistant race after NPV infection. The early up-regulation of these genes suggests that these genes could play an important role in baculovirus resistance in the silkworm, B. mori.

• International Journal of Industrial Entomology and Biomaterials
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• v.13 no.1
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• pp.31-35
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• 2006

The ubiquitin conjugating enzyme 2 (E2) is core component of ubiquitin proteasome pathway (UPP) which represents a selective mechanism for intracellular proteolysis in eukaryotic cells. The E2 has been implicated in the intracellular transfer of ubiquitin to target protein. We show here the involvement of E2 in antiviral immune of Bombyx mori to Bombyx mori nuclear polyhedrosis virus (BmNPV). In this study, mRNA fluorescent differential display PCR (FDD-PCR) was performed with BmNPV highly resistant silkworm strain NB and susceptible silkworm strain 306. At 24 h post BmNPV infection, FDD-PCR with the arbitrary primer AP34 showed that one cDNA band was down-regulated in the midgut of resistant strain, but highly expressed in susceptible strain. The deduced amino acid sequence of this cDNA clone share 99% identity with the recently published B. mori ubiquitin conjugating enzyme E2 (Genbank NO: DQ311351). Fluorescent quantitative PCR corroborated down regulation of E2 in resistant strain. We there conclude that BmNPV infection evokes strong response of susceptible strain including activation of UPP. BmNPV may evolve escape mechanisms that manipulate the UPP in order to persist in the infected host. In addition, the identification of down-regulation of E2 in resistant strain, as well as structure data, are essential to understanding how UPP operates in silkworm antiviral immune to BmNPV disease.

• International Journal of Industrial Entomology and Biomaterials
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• v.13 no.2
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• pp.85-95
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• 2006

Seveval Chinese and Japanese varieties with good characters were used in the breeding. After 5 years (15 generations), a pair of robust and high quality silkworm variety with NPV resistance was bred by means of a combination of crossing and pedigree selection complemented by the selection of NPV resistance. The variety was identified jointly nationwide in 2003 and 2004, and appraised by National Mulberry and Silkworm Appraising Committee. Results are as follows: its cocooning rate is over 93%, shell rate 23-25%, filament length 1200-1300 meters, reelability 75-88%, Length of non-broken cocoon filament 900-1100 meters, raw silk rate 17-19%, neatness 95-97 points, and cocoon crop, cocoon shell weight and raw silk weight per 10000 larvae is higher than those of the control variety by 7-10%, 14-19% and 14-18%, respectively. The variety is not only robust, resistant to high temperature and NPV, easy to rear, uniform in hatching, molting and maturing, but also lays more eggs, and its fecundity is high. It is suitable to rear in the Yangtze River Basin, the Yellow River basin and the Pearl River basin of China.

• International Journal of Industrial Entomology and Biomaterials
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• v.3 no.2
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• pp.187-190
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• 2001

Inheritance pattern of resistance to Bombyx mori nuclear polyhedrosis virus (BmNPV) was studied in an Indian silkworm stock TX by single back-cross test method. The resistant parent ［TX］, susceptible parent ［HM］, their Fl, F2, and Fl progeny back-crossed to TX [BC(R)] and HM [BC(S)] were inoculated per os with a fixed concentration of BmNPV($0.5{\times}10^{th} PIB/ml$) on the first day of second stadium. The cumulative mortality was recorded until day $10^{\times}$ post-inoculation. The results show that the resistance to BmNPV in TX fellow mono Mendelian inheritance pattern. The resistance dominated over the susceptibility at Fl. At F2, the resistant and susceptible offspring segregated in 3:1 ratio whereas at BC(S), the resistant and susceptible offspring segregated in 1:1 ratio. The response of BC(R) was more or less like the resistant parent TX which confirms the involvement of a major dominant gene conferring resistance to BmNPV in TX. The possible mechanism of inheritance of resistance in TX is discussed.

• Journal of Microbiology and Biotechnology
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• v.4 no.3
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• pp.183-190
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• 1994

Cell-free extracts prepared from cultured insect cells, Spodoptera. frugiperda, were analyzed for activation of early gene transcription of an insect baculovirus, Autographa californica nuclear polyhedrosis virus (AcNPV). The template DNA used for in vitro transcription assays contained promoter sites for the baculovirus genes that have been classified as immediate early ($\alpha$) or early genes. These genes are located in the HindIII-K/Q region of the AcNPV genome. Nuclei isolated from the AcNPV-infected Spodoptera frugiperda cells were also used for in vitro transcription analysis by RNase-mapping the labeled RNA synthesized from in vitro run-on reaction in the isolated nuclei. The genes studied by this technique were p26 and pl0 genes which were classified as delayed early and late gene, respectively. We found that transcription of the genes from the HindIII-K region was accurately initiated and unique in the whole cell extract obtained from uninfected cells, although abundance of the in vitro transcripts was reverse to that of in vivo RNA. With isolated nuclei transcription of the p26 gene was inhibited by $\alpha$-amanitin suggesting that the p26 gene was transcribed by host RNA polymerase II. However, transcription of the pl0 gene in isolated nuclei was not inhibited by $\alpha$-amanitin, but rather stimulated by the inhibitor. We also found that the synthesis of $\alpha$-amanitin-resistant RNA polymerase was begun before 6 hr p.i., the time point at which the onset of viral DNA replication as well as the appearance of a-amanitin-resistant viral transcripts were detected. These studies give us strong evidence to support the previous data that early genes of AcNPV were transcribed by host RNA polymerease III, while transcription of late genes was mediated at least by a novel $\alpha$-amanitin-resistant RNA polymerase.

• 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.

• Journal of Sericultural and Entomological Science
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• v.27 no.2
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• pp.20-27
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• 1985

This study was carried out to investigate the histological changes after the infection of nuclear and cytoplasmic polyhedrosis viruses(NPV, CPV) and the resistance to the viruses in various varieties of the silkworm fed on artificial diet. The results obtained were as follows; Among four varieties of silkworm tested, Jam 107$\times$Jam 108 was more resistant than the other varieties tested and Jam 119$\times$Jam 120 was the most resistant to CPV. In case of peroral infection with NPV, Jam 107$\times$Jam 108 showed lower mortality than the remained varieties in low concentration (104/ml). However, all varieties showed high mortality as the concentration of viruses was increased. With infection of CPV, the varieties showed high mortality at the concentration of 107 and 108/ml, while Jam 119$\times$Jam 120 showed the lowest mortality at virus concentration of 104/ml. The fat bodies, epidermal cells and tracheal epithelial cells showed high susceptibility to NPV to break the cells completely and liberate the debris to the body cavity. The CPV infected only the cylindrical cells of mid-gut and formed polyhedrons. In some cells, CPV was liberated to gastral cavity. In the electrophoretic pattern of hemolymph protein of silkworm larvae infected with NPV, bands were dimmed and disappeared as symptom aggravated after infection. Electrophoretic pattern of homolymph proteins of silkworm larvae infected with CPV showed no numerical difference at the later stage of infection, and one or two bands was observed along with lowering the concentrations.

• Journal of Sericultural and Entomological Science
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• v.22 no.2
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• pp.8-12
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• 1981

Practical application of silkworm artificial diet is very desirable to save labour in sericultural industry as the problem of labour shortage is becoming serious in Korea. However, silkworms reared on the artificial diet are more susceptible to viruses than those reared on mulberry leaves because of the lower anti-viral activity of gut juice of silkworms grown on artificial diet compared with that of silkworms grown on mulberry leaves. In this study, authors investigated the varietal difference of silkworm larvae, Bombyx mori L., reared on artificial diet which contained 20 percent of dried mulberry powder, in the susceptibility to nuclear polyhedrosis virus (NPV). The results showed that there is no difference in susceptibility to NPV among tested varieties when high concentration of NPV was admitted to silkworm larvae, but varietal difference appeared in lower concentration admitted. Among 7 hybrids tested, Hansaeng 1${\times}$Hansaeng 2 was most resistant to NPV with an $LC_{50}$ of 2.7${\times}$10$\^$6/ and Jam 111${\times}$Jam 112 was also more resistant comparatively than other hybrids.

• 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.

• International Journal of Industrial Entomology and Biomaterials
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• v.13 no.2
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• pp.73-77
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• 2006

Combining ability and hybrid vigour analysis was carried out in hybrids between newly developed non-susceptible lines to BmDNV1 and popular bivoltine breeds for certain quantitative traits viz. Pupation rate, Cocoon yield, Cocoon weight, Cocoon shell weight and Cocoon shell ratio, Survival rate against BmIFV and BmNPV. General combining ability (GCA) effects revealed that among the lines CSR2DR was found good general combiner exhibiting significant GCA effects for six characters, out of seven traits evaluated. Among testers CSR28DR was found as good combiner exhibiting significant GCA effects for six traits. Out of 36 hybrids made between $resistant{\times}resistant,\;resistant{\times}susceptible\;and\;susceptible{\times}susceptible$ breeds, one hybrid $CSR21DR{\times}CSR28DR$ exhibited significant SCA effects for six traits. The selected hybrid $CSR21DR{\times}CSR28DR$ also exhibited significant positive heterosis and heterobeltiosis expressions for maximum traits and could be exploited as commercial silkworm hybrid resistant to important viral diseases.