Structure, secretion, protein characterization and effect of wheat germ agglutinin on the peritrophic membrane of larval European corn borer (Ostrinia nubilalis)

European corn borer (ECB, Ostrinia nubilalis) larval peritrophic membrane (PM) formation and structure and protein characteristics were examined as well as the effect wheat germ agglutinin (WGA) on PM structure and larval growth inhibition. PM secretion and formation occur primarily in the anterior region of the mesenteron as determined by light and electron microscopy. Nascent PM first became visible within and at the tips of the microvilli as fibrous linear chitin-containing structures as visualized by staining with gold-labeled WGA. No formed PM was visible at the foregut-midgut junction, but a thin single PM first appeared in the lumen between the stomodeal valves and the midgut epithelium. Just posterior to the stomodeal valves, multiple PMs were observed that became progressively thicker and more numerous in the mid and posterior regions of the mesenteron.; ECB larvae fed WGA in their diet for 72 h showed very little increase in body weight, whereas control larvae increased their weight nearly four fold. Light and transmission electron microscopy studies showed that the morphology of the PM changes within 24 h after ECB larvae have fed on the WGA diet. Whereas the PM of control larvae is a thin membranous structure in the anterior region of the midgut, the WGA-fed larvae secreted a multiple layered and unorganized PM that contained embedded food particles, bacteria and broken microvilli. Posteriorly, food particles and bacteria penetrated through the membrane to the brush border. Dark staining amorphous structures inundated the PM that intensely localized anti-WGA which showed they were WGA aggregations.; PM extracted with SDS contained multiple proteins ranging from 20 to over 200 kDa as determined by SDS-PAGE, some of which bound WGA. Western blotting demonstrated WGA-fed larvae partition the lectin from midgut epithelium by binding WGA to PM. Two major PM proteins were purified and N-terminal sequence analysis showed close similarity of one to trypsin-like proteases and the other to basic juvenile hormone sensitive proteins. Trypsin, chymotrypsin, and elastase-like proteases occur in the PM and activity remains after multiple washes. SDS at certain concentrations activates trypsin-like protease activity as well as solubilizing the enzyme from the PM.