Methodologies for Plasmopara halstedii Research

Abstract The objective of this work was to find practical procedures to overcome methodological drawbacks encountered during studies on sunflower downy mildew. Techniques for recovering living isolates of Plasmopara halstedii from the field and for the preservation of infected leaf samples for further molecular analysis were developed. A Polymerase Chain Reaction (PCR)-based test for the detection of P. halstedii in sunflower leaves and a method to remove azoxystrobin from fungicide-treated seeds are proposed. In situ-inoculations of pre-germinated seeds allowed the recovery of living isolates from the field. Three sample-preservation methods were evaluated (silica, heating and lyophilization) resulting in high yield and quality of the DNA extract. It was detected the presence of the pathogen in symptomless leaves through PCR using molecular markers based on expressed sequence tags. A treatment using sodium hypochlorite is recommended for the removal of azoxystrobin from fungicide treated seeds.