Chitinase-gold complex used to localize chitin ultrastructurally in tomato root cells infected byFusarium oxysporum f. sp.radicislycopersici, compared with a chitin specific gold-conjugated lectin

Abstract

A cytochemical technique for the ultrastructural localization of chitin in tomato root cells infected by Fusarium oxysporum f. sp. radicis-lycopersici is reported. Chitinase was complexed to colloidal gold and thin sections were incubated with the enzyme-gold complex. This technique yielded a more uniform distribution of gold particles over the fungus wall, compared to that obtained with the lectin-gold technique. Both techniques revealed no labelling of the fungus cytoplasm, except for organelles resembling Woronin bodies. No significant labelling of either healthy or infected root cells was seen except for the secondary walls of vessels and, occasionally, that of adjoining parenchyma cells. The importance of this technique in studying the development of the pathogen within host cells is discussed.

Secondary Cell-Wall-Specific Glycoprotein(s) from French Bean Hypocotyls

“…The isolation and purification of three novel cell-wall glycoproteins were based on the observation that WGA binds to secondary cell walls of xylem vessels and phloem fibers when used as a probe in ultrastructural studies (Chamberland et al, 1985;OConnell and Ride, 1990). Using WGA and anti-WGA antibodies, a detection system not used previously, we confirmed these observations.…”

“…In earlier works in which the occurrence of the gold deposition over secondary cell walls was reported (Chamberland et al, 1985;Benhamou and Asselin, 1989;Hogetsu, 1990;OConnell and Ride, 19901, mainly WGA or chitinase conjugates were used. Here the use of the WGA plus anti-WGA antibodies system, a new method with greatly increased sensitivity, is reported.…”

“…Vol. 108, 1995 deposition was also found in both healthy and infected plants over lignified secondary cell walls, mainly in xylem vessels, and this labeling was completely inhibited by N,N',N"-triacetylchitotriose (Chamberland et al, 1985;O'Connell and Ride, 1990). Two attempts to explain the observed binding have been made without firm conclusions (Benhamou and Asselin, 1989;Hogetsu, 1990).…”

Enzyme‐gold electron microscopic cytochemistry: A new affinity approach for the ultrastructural localization of macromolecules

“…The technique appears to be based on the enzyme-substrate interaction since 1) the enzyme-gold complexes display their enzymatic activities (Bendayan and Gisiger, unpublished results) and 2) incubations performed in conditions noncompatible with the enzymatic reaction (either at low temperature, wrong pH values, denaturation of the enzymes, or presence of inhibitors) result in an absence of specific labeling. The technique has now been applied by other groups confirming our results and extending its application (Raska et al, 1983;Danscher and Norgaard, 1983;Wan et al, 1983Chamberland et al, 1985. It has also been adapted for light microscope cyto- Fig.…”

“…After incubation with the xylanase-gold complex, the xylans proteins were specifically revealed over the secondary walls of this plant tissue while the primary ones and the cell cytoplasm were devoid of labeling (Vian et al, 1983) (Fig.20). (Chamberland;.…”