Visualization of halos in the epidermal cell wall of Allium cepa caused by Colletotrichum dematium f. circinans and Botrytis allii using fluorochromes

Abstract

Halos were detected with epifluorescence microscopy around penetration sites of Colletotrichum dematium f. circinans and Botrytis allii in onion epidermal cell walls as areas of less intense fluorescence or negatively stained areas in fluorescing cell walls following treatments with berberin sulphate and acridine orange but not with brilliant sulphaflavine (which stained the cell wall), ninhydrin, dansylchloride, or analine blue. Since pectin, pectic acid, avacil (microcrystaline cellulose super fine), filter paper, and Sephadex G-100–120 fluoresced with acridine orange and berberin sulphate, it was inferred that the halos were negatively stained or appeared as areas with less intense fluorescence because enzymes from these pathogens degraded cell wall pectin and cellulose at the point of penetration. Spores of both pathogens fluoresced when stained with brilliant sulphaflavine, acridine orange, ninhydrin, and dansylchloride. These stains and berberin sulphate caused germ tubes, appressoria, and primary infection mycelia to fluoresce. Nuclei in these fungal structures fluoresced when stained with acridine orange and brilliant sulphaflavine.