Immunocytochemical localization ofBacillus thuringiensisCryl toxins in the midguts of three forest insects andBombyx mori

Abstract

Light microscopy was used to investigate the relation between toxicity, cytopathological effects, and in vivo binding of Bacillus thuringiensis CrylA(b) and CrylE toxin proteins in larvae of Lymantria dispar, Choristoneura fumiferana, Actebia fennica, and Bombyx mori. These target insects were selected for their contrasting susceptibility to the two toxins. Lymantria dispar is susceptible to CrylA(b), B. mori is susceptible to CrylE, C. fumiferana is susceptible to both, and A. fennica is not susceptible to either. In the susceptible species, both toxins caused typical pathological changes in midgut epithelial cells, including disruption and shedding of the brush border membrane, vacuolization of the cytoplasm, and swelling of the cells and their nuclei, followed by disintegration and release of cytoplasmic content into the lumen. In the highly resistant A. fennica, no cell damage was observed, but the midguts of toxin-fed larvae had a shrunken appearance. Immunohistochemical staining of midgut sections from toxin-fed larvae revealed that the toxins bound to the microvilli of the midgut epithelial cells of susceptible species only, with the exception of B. mori. In this species, the CrylA(b) toxin bound to the apical microvilli without causing cell damage or larval death. In vivo binding of toxins is thus not always correlated with larval toxicity. Accumulation of the toxins at the peritrophic membrane depended on both toxin and insect species and was not correlated with larval toxicity.Key words: Bacillus thuringiensis, Cryl toxins, forest Lepidoptera, immunocytochemical localization, cytopathology.