Quantification of the rice false smut pathogen Ustilaginoidea virens from soil in Japan using real-time PCR

Abstract

Ustilaginoidea virens is the causal agent of false smut disease of rice. In this study, we developed a real-time polymerase chain reaction (PCR) assay to clarify the relationship between false smut occurrence on rice and quantification of U. virens from soil in Japan. The method here described is sensitive, detecting less than 50 fg of pathogen DNA, and specific to the nuclear ribosomal DNA for U. virens when tested across 27 rice-pathogenic fungi and bacteria, 26 other fungi and bacteria and four plant species. As few as eight chlamydospores of U. virens per gram soil were detected when added to sterilized Gley and Ando soils. The real-time PCR assay for the soil samples was at least 100-fold more sensitive than the conventional and nested-PCR assays tested. By quantification of U. virens with real-time PCR using DNA extracted from naturally contaminated Gley soils and visual assessment of the disease in agricultural fields, a linear correlation between cycle threshold (CT) values and the number of false smut balls was revealed. Therefore, this specific quantitative assay could be a useful tool for optimization of disease control strategies, and for studying the ecology of U. virens.