Use of Green Fluorescent Protein-Transgenic Strains to Study Pathogenic and Nonpathogenic Lifestyles inColletotrichum acutatum

Abstract

Colletotrichum acutatum, which causes anthracnose disease on strawberry, can also persist on several other plant species without causing disease symptoms. The genetic and molecular bases that determine pathogenic and nonpathogenic lifestyles in C. acutatum are unclear. We developed a transformation system for C. acutatum by electroporation of germinating conidia, and transgenic isolates that express the green fluorescent protein (GFP) were produced. Details of the pathogenic and nonpathogenic lifestyles of C. acutatum were determined by using GFP-transgenic isolates. Major differences between colonization-mediating processes of strawberry and of other plants were observed. On the main host, strawberry, the germinating conidia formed branched, thick hyphae, and large numbers of appressoria were produced that were essential for plant penetration. In strawberry, the fungus developed rapidly, filling the mesophyll with dense mycelium that invaded the cells and caused necrosis of the tissue. In nonpathogenic interactions on pepper, eggplant, and tomato, the conidia germinated, producing thin, straight germ tubes. Appressoria were produced but failed to germinate and penetrate leaf tissue, resulting in epiphytic growth without invasion of the plant. Penetration of the plant occurred only several days after inoculation and was restricted to the intercellular spaces of the first cell layers of infected tissue without causing any visible damage. Much of the new fungal biomass continued to develop on the surface of inoculated organs in the nonpathogenic interaction. The differences in fungal development on strawberry compared with the other plant species suggest that signal molecules, which may be present only in strawberry, trigger appressorial germination and penetration of the primary host.

Ultrastructure of the early stages of Colletotrichum acutatum infection of strawberry tissues

“…The presence of subcuticular, intramural, and intra-or inter-cellular hyphae in leaf tissues disagrees with results of Leandro et al (2001), who did not detect host penetration of strawberry leaves by C. acutatum. However, our observations agree with those of Horowitz et al (2002), who detected the fungus inside strawberry leaves.…”

“…Microcyclic conidiation or conidiogenesis was observed on both adaxial and abaxial surfaces of attached strawberry leaves. This process has already been described in C. capsici on cowpea leaves (Latunde-Dada et al 1999) and in C. acutatum on detached (Leandro et al 2001) and attached (Horowitz et al (2002) strawberry leaves, which may have implications on disease epidemics. However, on strawberry petioles, conidiogenic structures were not found.…”

“…Howard et al (1991) reported that high turgor pressure in the appressorium is essential for mechanical penetration. Horowitz et al (2002) observed collapsed appressoria after penetration, which they related to losses of turgor pressure in these structures.…”

“…Leandro et al (2001) reported the germination and the formation of secondary conidia of this pathogen on the surface of detached strawberry leaves. Horowitz et al (2002) studied the development of C. acutatum on strawberry and other plant species by using transgenic isolates. Curry et al (2002) studied the histopathology of C. acutatum and C. fragariae in strawberry stolons and petioles and described the process of infection, showing, at the ultrastructural level, stages of invasion of these pathogens with hyphae under host cuticle and in anticlinal host wall.…”

Infection Process of Olive Fruits by Colletotrichum acutatum and the Protective Role of the Cuticle and Epidermis

“…Talhinhas et al, (2005) with molecular and phenotypic assays reported that the occurrence of C. acutatum in Portuguese olive orchards was higher (>97%) than C. gloeosporioides (<3%). The C. acutatum J. H. Simmonds is one of the most pathogenic species, which causes anthracnose and blight in agriculturally important hosts including many dicotyledonous plants such as strawberry, apple, citrus, and stone fruits (Horowitz et al, 2002;Peres et al, 2005).…”