IMMUNOCYTOCHEMICAL ANALYSIS OF SPECIFIC BINDING OF BACILLUS-THURINGIENSIS INSECTICIDAL CRYSTAL PROTEINS TO LEPIDOPTERAN AND COLEOPTERAN MIDGUT MEMBRANES

The in vitro binding of four insecticidal crystal proteins (ICPs) from Bacillus thuringiensis to midgut tissue sections of two lepidopteran (Manduca sexta and Plutella xylostella) and one coleopteran (Leptinotarsa decemlineata) species was immunocytochemically analyzed. Monoclonal antibodies, highly specific against the different ICPs, were used. Light microscopy observations showed that the crystal proteins accumulated at the exposed peritrophic membrane of all lepidopteran insects and at the apical microvilli of sensitive lepidopteran insects. No accumulation was observed at the microvilli present in the goblet cell cavity or at the Malpighian tubules. For all lepidopteran insects the binding of the different ICPs to the apical microvilli of the midgut epithelium appears to be highly specific: only the ICPs which were toxic bound to the apical microvilli of midgut tissue sections. As an exception, the nontoxic CryIB bound to the apical microvilli of M. sexta. The binding of ICPs to the peritrophic membrane was not correlated to toxicity. For the first time binding of an ICP to a coleopteran insect larva has been shown. Only the coleopteran-specific toxin CryIIIA bound to the brushborder and to the peritrophic membrane. The binding is preferential to the apical microvilli in the posterior part of the midgut. These results suggest that the mechanism of ICP toxicity in coleopteran insects may be slightly different from that in Lepidopteran and/or Diptera. The present procedure is sensitive and rapid, can be applied to individual insects, and as such may prove very useful for analyzing the binding of different ICPs to small insects. © 1992.