Effect Of Antibodies Against Peritrophic Membrane Proteins On Baculovirus Infectivity

The peritrophic membrane (PM) is a noncellular inner-tube-like lining of the insect midgut that consists of chitin microfibrils associated with proteins, glycoproteins, and proteoglycans.It acts as a semipermeable membrane that regulates the passage of molecules between the midgut epithelium and lumen and may serve as a barrier to pathogenic microorganisms. Many studies demonstrated that physical or chemical factors such as chitinase, enhancin and optical brightener(or fluorescent brightener) could disrupt the PM and significantly enhanced the infectivity of baculo- virus. Therefore, the fact that it magnified the pores in the PM and disrupted the structural integrity of the PM could enhance the infectivity of virus. On the contray,we could have the opposite result,if the pores in the PM were jammed or decreased. Based on this understanding,we used the peritrophic membrane proteins dissociated from the PM of Spodoptera exigua by optical brightener to preparation antibodies which were applied to study the effect of antibodies against peritrophic membrane proteins on baculovirus infectivity.1% Fluorescent brightener 28 (M2R) was used to dissociate peritrophic membrane proteins from the peritrophic membrane of Spodoptera exigua in vitro. Antiserum were obtained by injecting healthy male rabbits weighted 2³ kilogram with the PM proteins.First, the antiserum and four proteinase inhibitors (0.5ug/mLLeupeptin, lug/mLPepstatin,lmmol/LEDTA,lmmol/LPMSF)were applied to the surface of artificial diet to feed the test larvae. Then we inoculated instars of 3¹⁶s(16h after third instar) and 4°s (early fourth instar)with AcMNPV-hsp70/lacZ of the concentration of 1.55x10⁷ PIBs/ml in presence of 1%(end concentration) M2R at various time intervals (0h,2h, 4h,6h), supposing control groups (namely in the absence of antibody and FB-28).The result indicated that with the time going, the death rate of the virus to the larvae gradually reduced after the instars of 3¹⁶s(16h after third instar) were fed the antiserum, but the mortality of two groups without feeding anti-serum generally was maintained at 37.8% and about 80%. When we test to the instars of 4°s (early fourth instar), with time postponing, the death rate of the virus to the larvae all gradually reduced, but the mortality of two groups feeding anti-serum was lower than that of two groups without feeding anti-serum.When Spodoptera exigua of end third instar larvae fed antiserum 4h were infected with virus , the LC50 value was 6.19xlO9 PIBs/ml. The LC50 value was 1.56xlO7 PIBs/ml. when Spodoptera exigua of end third instar larvae were infected at a single dosage of virus. The LC50 value increased 397-fold contrast between the two treatments. When Spodoptera exigua of end third instar larvae were infected with virus in the presence of FB-28, the LC50 value was 4.11xlO4 PIBs/ml. The LC50 value was 1.04xl06 PIBs/ml when Spodoptera exigua of end third instar larvae fed antiserum 4h were infected with virus in the presence of FB-28. The LC50 value increased 25-fold , when Spodoptera exigua were fed antiserum.According to the findings above, it could be concluded that: when Spodoptera exigua of end third instar larvae were fed antiserum, antiserum may react with peritrophic membrane proteins to form antigen-antibody compound which jammed the pores of peritrophic membrane. With the time going, the close degree would become much higher than before. On the one hand, the fact that antibodies led to jam the pores reduced the permeability of the virus invading peritrophic membrane. On the other hand, it probably affected fluorescent brightener to contact with peritrophic membrane proteins, thereby, preventing fluorescent brightener from dissociating peritrophic membrane proteins. Then, it caused the enhancing effects of fluorescent brightener to reduce.