Use of Fusarium oxysporum f. sp. dianthi transformed with marker genes to follow colonization of carnation roots

A highly pathogenic F. oxysporum f. sp. dianthi (Fod) strain has been transformed with genes coding for the Green Fluorescent Protein (GFP) and the Red Fluorescent Protein (DsRedFP). Expression of GFP and DsRedFP resulted in bright green or red cytoplasmic fluorescence of mycelium when observed under fluorescent light. Stable transformants, selected on the basis of brightness and stability of fluorescence, were inoculated on roots of a partially resistant cultivar of carnation. Root and vessel colonization was evaluated, at regular times, by fluorescent microscopic observation of plant tissue sections. Autofluorescence of the carnation root was very high at the emission wavelength of the DsRed-Express, so that it was not possible to distinguish infecting hyphae within plant tissues. GFP transformed Fod was easily visualized on and within root tissues. The fungus invades the root tip and its growth seems to proceed randomly in the sub-apical zone. The hyphae are confined within the vascular cylinder by the endodermis cells beginning from the zone of differentiation of vascular tissues and are able to grow inside vessels.