In-vitro selection of Vitis vinifera 'Chardonnay' with Elsinoe ampelina culture filtrate is accompanied by fungal resistance and enhanced secretion of chitinase

Proembryogenic masses of grapevine (Vitis vinifera L.) 'Chardonnay' (clone 02Ch) were exposed to the culture filtrate of Elsinoe ampelina (deBary) She ar, the causal agent of anthracnose disease. After four or five cycles of r ecurrent in-vitro selection with medium containing 40% fungal culture filtr ate, putative resistant lines RC 1 and RC 2 respectively, were established. The selected lines inhibited the growth of E. ampelina and Fusarium oxyspo rium (Schlecht.) (isolated from watermelon) in a dual-culture assay and red uced the growth of mycelium on a conditioned-medium test, thus suggesting t he involvement of extracellular compounds in resistance. Sodium dodecyl sul fate-polyacrylamide (SDS-PAGE) gel electrophoresis of extracellular protein s from spent suspension-culture medium showed enhanced secretion of new pro teins by selected lines. A 36-kDa protein was immunodetected by a chitinase antiserum. This chitinase continued to express constitutively in different iated somatic embryos and also in the intercellular fluids of plants regene rated from the selected lines. Somatic embryos from selected lines grew uni nhibitedly in a medium containing 40% fungal culture filtrate, whereas non- selected (control) somatic embryos became necrotic and died within a few da ys. Plants regenerated from selected lines exhibited resistance to infectio n by E. ampelina in both greenhouse tests and detached leaf bioassays. Resu lts suggest that embryogenic cells can be selected for resistance following in-vitro selection, resulting in resistant plants. Whether or not resistan t cells pre-existed in the original embryogenic culture or were induced by the selection pressure could not be determined.