| Peritrophic membrane (Peritrophic membrane/Peritrophic matrix, PM), is a kind of an acellular membrane structure which is secreted by the midgut epitheliums in the long-term evolutionary process of the insects. Peritrophic membrane is composed of glycoproteins and chitin which is similar to the mucous secretions of the vertebrate digestive tract. It is the first physical barrier to protect the epithelial cells of insect midgut from food particles abrasion and pathogenic microorganisms, when pathogenic microorganisms enter in the body of the insect through midgut. Research suggests that the rate of virus infection and mortality of insect will increase while the peritrophic membrane is destroyed. As a model of the Lepidoptera, there are plenty of studies and researches on the silkworm physiology and biochemistry and pathology. However, even with the great development in the genomics research in past decade, there are few researches on the peritrophic membrane. BmNPV(Bombyx mori nuclear polyhedrosis virus) is the main disease virus in silkworm rearing which causes a variety of Bombyx mori larvae disease and leads great economic loss in sericulture. To increase the resistance of silkworm to BmNPV and to breed the diesase-resistant variety is the effective measure to protect the infection of BmNPV. Studies on the relationship between the peritrophic membrane of the silkworm larvae and the BmNPV has significant theoretical and practical meaning. This thesis discusses the interrelationship between peritrophic membrane and BmNPV by using of morphology, histological and scanning electron microscope to observe the changes of the midgut and PM from BmNPV Per os and uninfected silkworm larvae. The main results are as follows:1. Investgate the mortality rate from9strain silkworm larvare by different concentrations BmNPV Per os. SM011has the highest LC50, LC50>1.00x109PBIs/ml, SM016has the lowest LC50, LC50is5.2x106PBIs/ml. Other strains have a medial mortality concentration LC50between1.68x107to1.19x108. It turns out that SM016showed susceptible to BmNPV virus, SM011showed resistant to BmNPV virus.2. Using varible concentrations of FB-28and virus to feed4th instar larvae from4strains, showing that FB-28promotes the virus infection to the larvaeS. Holes and gaps are clearly observed on the peritrophic membrane (PM) by scanning electron microscope, which verifies that feeding of FB-28destroyed the peritrophic membrane of silkworm larvae, but the damage could be temporarily recovered.The damage on the peritrophic membrane (PM) structure and integrality may be the main reason that increases virus infection.3. Midguts from three silkworm strain larvae were observed by paraffin dissection method. The results showed that the peritrophic membrane (PM) of normal silkworm larvae is a kind of long flexible tubular acellular net-membrane structure with varible thickness, smooth surface. The peritrophic membrane (PM) almost covers the whole inner surface along the midgut, with a multilayer structure.Otherwise the outer space of PM, the secretion substance quantity, the peritrophic structure tightness and the thickness are discrepancy among the investgated strains which showed stronger virus resistance larvae have more secretion substance in midgut and tighter and the thicker PM.4. Paraffin dissection showed that different strain larvae midgut appear different damages after BmNPV per os infection. Midgut epithelium of Sm011with stonger virus resistant showed no cell rupture after72hours per os, only a few nucleus inflations were observed. But Sm018and Sm013strain larvae with low resistance showed midgut epithelium were destroyed badly with nucleus sheded after72hours Per os. After96hours Per os, all three strain larvae midguts were discovered severe damages with nucleus sheded outside of the cell. This discrepancy of nucleus sheding may be related to the signaling, response and self-repairing capability of the midgut epithelium.5. Surface structure of silkworm larve peritrophic membrane (PM) was observed by scanning electron microscopy after Per os BmNPV. The results showed that the normal larve peritrophic membrane (PM) surface without infection is a smooth and integrityfiber network, with no hole and gap on it. After2-3hours infected per os by BmNPV, obvious changes occured on the PM surface, it becomed loose, irregular and a a variety of hole appearing. After6hours Per os, the surface recovered to a regular fiber network structure, no any holes exist. In the early stage of virus infection, the damage time and repairing time occurred on PM is different among different strains Damage appearing on PM of SM018strain is after2-3hours Per os, and the repairing time is after6hours Per os. However, damage time appearing on SM011strain is after2hours and3hours per os respectively. This result is in correspondence with strins virus resistant,i.e, SM018is susceptivity and SM011is more resistant to BmNPV.6. Holes and heterogenous materials apprearing on peritrophic membrane after24hours per os, at48hours holes becomed bigger. After72hours Per os, the damage on PM surface becoming much more serious. The time is different between different silkworm strains,for SM011with strong resistance, the hole appears after48hours Per os, but for SM018, the time is24hours Per os. The results suggest that epithelium responses to the virus, as well as their capability of epithelium renewing, may be different among different strains.The results verified the existence of resistances discrepancy to BmNPV among silkworm strains. We found that BmNPV Per os altered the host midgut epithelium and peritrophic membrane (PM) structure at the early and late stage of virus infection.And the influence varies among strains. This may be related to midgut epithelium response, cellular immune response and cell renewing. The results enrich our understanding to the interaction between BmNPV and silkworm. |
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