Differential responses of citrus calli and protoplasts to culture filtrate and toxin of Phoma tracheiphila.

Nucellar calli from two citrus cultivars with known tolerance to mal secco disease were chosen as experimental material, to test the pathogen's response to culture filtrate (CF) and partially purified toxin (PPT). The response of the two calli to the CF was in reverse order to the known response of the two cultivars to natural and artificial inoculations with Phoma tracheiphila. HPLC analysis of P. tracheiphila CF indicated the presence of a relatively high level of indole acetic acid (IAA). The response of the two calli and protoplasts derived from these calli to increasing amounts of IAA in the culture media was in the same order as that of calli and protoplasts to CF. In contrast, the responses to PPT of calli and protoplasts from these two types confirmed the relative tolerance of 'Femminello' lemon and 'Tarocco' orange trees to mal secco disease.

Nucellar callus of 'Femminello' lemon, selected for tolerance toPhoma tracheiphila toxin, shows enhanced release of chitinase and glucanase into the culture medium.

Phoma tracheiphila is the causative agent of the disease mal secco. Citrus cultivars differ substantially in respect to their sensitivity to the pathogenP. tracheiphila and its toxin. Some cultivars (e.g., 'Femminello' lemon) are inherently sensitive while others (e.g., 'Tarocco' orange) are tolerant. Cell lines derived from nucellar tissue of 'Femminello', 'Tarocco' and a cell line selected for tolerance to the fungal toxin ('Femminello-S') were used to study host-pathogen interaction. Our results showed that calli or conditioned media of 'Tarocco' and 'Femminello-S' inhibited the size of co-cultivated fungal colonies when compared to 'Femminello'. In addition, conditioned medium of 'Tarocco' as well as 'FemminelloS', but not 'Femminello', promoted bursting of hyphal tips. A ten-fold increase in chitinase and glucanase enzymatic activity, as evaluated by radiometric assay and laminarin hydrolysis respectively, was detected in 'Femminello-S' extracellular extracts as compared to 'Femminello'. An increase in chitinase was also shown by immunoblot analysis. Our findings suggest a positive correlation between the presence of chitinase and glucanase in the conditioned media of the cultured cells and the tolerance of those cells toP. tracheiphila toxin.