SENSITIVE STAINING OF FUNGAL EXTRACELLULAR MATRICES USING COLLOIDAL GOLD

A method is described for visualizing fungal extracellular matrices (ECMs), based on the binding of non-conjugated colloidal gold particles. Staining is permanent and is viewed either with epi-polarization microscopy or with bright field microscopy after silver enhancement with a physical developer. Colloidal gold bound to cell surfaces through ionic interactions with the free amino groups of basic proteins. Sensitivity of the stain was high: five other protein stains (Orange G, Light Green SF, Alkaline Fast Green FCF, fluorescein isothiocyanate and N-(7-dimethylamino-4-methyl-3-coumarinyl)-maleimide), failed to label extracellular material that was stained by colloidal gold. Colloidal gold was used to demonstrate ECMs around germlings of four plant pathogenic fungi attached to artificial and host surfaces. Large circular films of protein were detected around conidia of Colletotrichum gloeosporioides and Erysiphe graminis and around appressoria of Magnaporthe grisea. Colloidal gold also revealed fibrillar protein sheaths enveloping germ-tubes of C. gloeosporioides and Botrytis cinerea.