Use of sem x-ray microanalysis for the study of fungicide mode of action: preliminary application on Erysiphe necator

The knowledge of fungicide mode of action (MoA) provides useful information in modern plant disease management, for the targeted use of the fungicides and implementation of anti-resistance strategies. Nowadays, the MoA is well-defined for a restricted number of fungicides. The study of fungicide MoA requires the integration of microscopy and bio-molecular tools, which may be more difficult to apply when the target organism is a biotrophic fungus. In this work, we used the SEM X-ray microanalysis in order to find differences in the chemical composition of Erysiphe necator chasmothecia sprayed with meptyldinocap. SEM X-ray is a scanning electron microscope able to detect chemical elements by their scattered X-ray emissions from the specimen. The principal component analysis performed on element energy emissions recorded on control and treated chasmothecia, revealed a relevant association between Ca, P and K. The function obtained by linear discriminant analysis made it possible to correctly classify treated and untreated samples considering Ca and Al energy emissions as the most significant variables. Noteworthy, two way ANOVA performed on the semi-quantitative Ca percentage estimation, showed that relative amount of Ca was higher in treated chasmothecia and increased significantly from 24 to 48 h after treatment. These results, together with the negative correlation observed between Ca and Al (r=-0,693) and S and P (r=-0,612), suggest a pH change in treated chasmothecia wall and the occurrence of an oxidative stress inside the chasmothecia which involves the Ca-dependent signal transduction pathway. These preliminary hypotheses need confirmation through further studies