A quantitative confined inoculation method for studies of pathogenicity of fungi on plants

A technique for inoculation with precise numbers of fungal spores on leaves and stems of plants was developed. The technique consisted of placing 1-μl drops with a fixed number of spores on the surface of leaves and stems, and covering each inoculum drop with a 10-μl drop of low-temperature gelling SeaPlaque agarose to fix the inoculum on the target site. With this technique single zoospores of Phytophthora capsici were able to cause local lesions on leaves and stems of peppers, and the size of the lesions directly correlated with the number of spores in the inoculum drops. Similar results were obtained when the technique was used to inoculate taro leaves with zoospores of Phytophthora colocasiae and black mustard leaves with Alternaria brassicae. This method has the advantages of being accurate and precise, and it is also easy to handle the inoculated plants. It may also be applicable to other pathogens.