Detection and Identification of Six Monilinia spp. Causing Brown Rot Using TaqMan Real-time PCR from Pure Cultures and Infected Apple Fruit

Brown rot is a severe disease affecting stone and pome fruits. This disease was recently confirmed to be caused by the following six closely related species: Monilinia fructicola, Monilinia laxa, Monilinia fructigena, Monilia polystroma, Monilia mumecola, and Monilia yunnanensis. Because of differences in geographic distributions, some of these species are important quarantine pathogens in certain countries. In this study, we developed TaqMan real-time polymerase chain reaction (PCR) assays to detect and identify the six species. Primer pairs and probes were designed for M. fructicola, M. fructigena, M. laxa, and Monilia polystroma based on sequence differences in the laccase-2 genes. Additionally, based on sequence differences in the elongation factor genes, primer pairs and probes were designed for Monilia mumecola and M. yunnanensis. The real-time PCR assays were able to specifically identify the target pathogens, with detection limits of 10–100 fg DNA, which is equivalent to 1–7 conidia. The assays we...