In vitro binding of <i>Bacillus thuringiensis</i> var. <i>israelensis</i> individual toxins to midgut cells of <i>Anopheles gambiae</i> larvae (Diptera: Culicidae)

Abstract

Midguts from Anopheles gambiae fourth instars were dissected and processed for immuno-light microscopy. Cloned insecticidal crystal proteins (ICPs) from Bacillus thuringiensis var. israelensis (Bti) were individually expressed in crystal-negative strains of Bacillus thuringiensis. Tissue sections of A. gambiae were incubated in vitro with each solubilized and trypsin-activated ICP. Immunodetection of CryIVA, CryIVB, CryIVD and CytA toxins on sections was performed using purified rabbit IgG directed against Bti ICPs, in combination with an anti-rabbit IgG/peroxidase. CryIVA, CryIVB, CryIVD and CytA toxins were detected on the apical brush border of midgut cells, in the gastric caecae and posterior stomach. CytA was also detected, to a lesser extent, on microvilli of anterior stomach cells.