| BmNPV(Bombyx mori L. Nucleopolyhedrovirus)is one of the most serious virus in sericultural production. Based on the incompleted statistics, about 70% losses in silkworm sericulture industry caused by the BmNPVdiseases. So far, the silkworm variety which can resistant to BmNPV is the most effective way for farmers. However, the mechanism of BmNPV disease resistance is still unclear. Recently, our research group bred a varity QiufengN, which carrying disease resistance genes and showing the highest resistance to BmNPV. The present study want to get some information about the profile of the resistance mechanism and some correlated genes using the high Bm NPV resistance variety QiufengN and the general variety Qiufeng. Based on the transcriptome data of fat body of these two variety, we cloned a 1512 bp cDNA sequence which encoded 504 amino acid. Blast analysis showed this gene very similarly to Bmtret gene, and belong to the trehalose coenzyme gene families.The gene's profile in the fat body of resistant varietyQiufengN an the general variety Qiufeng wer detected using fluorescence quantitative PCR. And the main results were listed as follows:1 Histomorphology observation on the blood, midgut, fatbody of QiufengN and Qiufeng after BmNPV infectionIn order to explore the histomorphology changes of BmNPV virus after infecting silkworm, the experiment were carried out on Qiufeng and QiufengN. The results showed that, compared to the Qiufeng, the blood in 60 h in QiufengN after BmNPV began to appear cloudy, which indicated the virus had a significant proliferation. In the midgut after feeding about 60 h compared to Qiufeng strain, rose slightly darker, erosion, atrophy of the state after 84 h. After about 48 hours after subcutaneous puncture BmNPV silkworm blood and control group began to appear cloudy, 60 H turbidity in the more obvious that virus multiplication near plateau, midgut in 72 hours and control group comparison began to appear yellow, erosion, after 96 h atrophy blackened, which can draw midgut tissues of virus immune defense is stronger than the blood. By optical microscope sectioning observation of silkworm has been around mouth added BmNPV 36 h after virus in the blood in the complete assembly of the first generation of; in the midgut, 24 h can be observed in the presence of polyhedrosis particles.2 Transcriptomic analysis of resistant and susceptible Bombyx mori strains providesinsights into the antiviral mechanismsThe resistance mechanisms of Bombyx mori against to BmNPV remain obscure. In the present study, RNA-Seq technology was applied to profile the transcriptome of QiufengN as well as Qiufeng strains and approximately 14.78 Gb clean data(more than 7.15 Gb for QiufengN and Qiufeng, respectively) were obtained from the two libraries obtained. A toal of 78,408 SNPs were identified in the Qiufeng strain of silkworm and 56,786 SNPs were identified in QiufengN strain. Besides, novel AS events were found in these 2 strains. In addition, 1,728 DEGs(242 up-regulated and 1,540 down-regulated) were identified in the QiufengN strain compared with the samples of Qiufeng strain. Analyses of GO categories revealed that these DEGs were mainly involved in GO terms related to membrane,metabolism, binding and catalytic activity, cellular processes, and organismal systems, such as oxidative phosphorylation and TCA cycle. Pathway analysis with KEGG annotation showed that the highest levels of gene representation were found in oxidative phosphorylation, Phagosome, TCA cycle, Arginine and proline metabolism, and Pyruvate metabolism. Additionally, COG analysis indicated that DEGs were associated with “Amino acid transport and metabolism” and “signal transduction mechanisms”.We identified that a rather larger number of DEGs were involved in some functional categories, indicating a series of major pathological changes in the silkworm following infection and several functions were related to the antiviral mechanisms of silkworm.Oxidative stress might be related to the defensive function of silkworm following BmNPV infection. In addition, many metabolic activities such as oxidative phosphorylation were affected and had biological relevance for the antiviral mechanisms of silkworm. Moreover,understanding the physiological functions of these DEGs involved in signaling pathway will require further characterization in order to determine their roles in immunity or other processes. These findings provide new research directions that may deepen our understanding of virus resistance of silkworm on the molecular level.3 Expression and effective analysis of trehalose coenzyme gene in midgut and fat Body of Bombyx mori induced by BmNPV?Trehalose transporter cDNA sequence was cloned using Bombyx mori strain Qiufeng N(resistance to BmNPV) and Qiufeng(Susceptible to BmNPV). The gene encodes 503 amino acids, which is predicted to have 12 transmembrane structures. To investigate the relationship between the tret gene and the mechanism of resistance to BmNPV, we usedreal-time fluorescence quantitative PCR to detect the relative expression levels in the larvae of silkworm's midgut and fat body after infection with BmNPV. The results showed that Qiufeng N and Qiufeng coding consistent sequence and expression level of tret gene was up-regulated at 24~48 h after Bm NPV infection, which indicated that the tret gene may involved in the immune response to BmNPV infection. |
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