Evaluation of methods for sterilizing boysenberry leaves for downy mildew infection studies

Abstract

Downy mildew of boysenberry, caused by Peronospora sparsa, is a major disease problem for New Zealand growers. Investigation of the biology and epidemiology of this biotrophic pathogen requires the production of sporangiospore inocula. Four leaf sterilisation treatments (70% ethanol, 10% bleach, sterile water, control) were assessed for their ability to reduce surface contaminants. After treatment, the leaf surfaces (six replicates) were pressed onto potato dextrose and nutrient agar plates and these were incubated at 20°C for 1 week. All treatments significantly (P<0.001) reduced both fungal and bacterial colonies compared with the untreated control. The numbers of fungal and bacterial colonies were higher on the adaxial than abaxial surfaces. To assess the effect of the treatments on susceptibility to P. sparsa infection the abaxial surface was inoculated with a 20 μl spore suspension droplet (2×104 spores/ml) and lesion development assessed after 1 month incubation at 20°C. The sterile water treatment did not affect the susceptibility of the leaves compared with the control, whereas ethanol and bleach altered susceptibility. Therefore, sterile water can be used to reduce contamination on boysenberry leaves before infection studies.