Cis-and trans-4,6 dihydroxymelleins: new phytotoxins of Phomopsis helianthi

Phomopsis helianthi Munt-Cvet et al. (perfect stage Diaporthe helianthi Munt-Cvet et al.) is an important fungal parasite causing a destructive disease known as "stem canker" of sunflowers (Helianthus annuus L.). In some European regions, particularly France and the Balkans, This wilting disease has caused noticeable economic losses to sunflower crops whereas P.helianthi infections were sporadic in Italy. A study was therefore carried out with two Italian (I1 and I2) and two french (F1 and F2) Phomopsis isolates which were examined for morphological, temperature growth responses and pathogenicity. The isolates showed marked differences; particularly Italian isolates produced only a-conidia whereas French isolates produced b-conidia. The optimum temperature for growth for all isolates was 25°C even if Italian isolates grew better within a large range temperature (5-30°C) with respect to French isolates (15-25°C). In pathogenicity test and then in screening for resistance with fourteen sunflower varieties carried out with I1 and F1 isolates, the French isolate was significantly more virulent than I1 isolate. A preliminary screening of different solid culture media (sunflower seeds, rice and maize kernels) for the production of phytotoxic secondary metabolites by P.helianthi showed the best results with F1 isolate cultured on rice. The extract of this culture showed the highest phytotoxic activity (tissue necrosis and chlorosis) when tested with leaf puncture bioassay and the presence of two major compounds with UV spectral characteristic similar to phomozin when analyzed by reversed phase HPLC. The structure of the two compounds were established on the basis of the spectroscopic data as trans-4,6-dihydroxymellein and cis-4,6-dihydroxymellein, respectively. This is the first report on the production of mellein derivates by P.helianthi and the isolation of trans-4,6-dihydroxymellein as a fungal metabolite.