An improved real-time PCR system for broad-spectrum detection of Didymella bryoniae, the causal agent of gummy stem blight of cucurbits

Gummy stem blight is a major disease of cucurbit crops caused by Didymella bryoniae (anamorph Phoma cucurbitacearum). Previously, we used random amplified polymorphic DNA (RAPD) profiles and specific PCR primers to sort a worldwide collection of isolates of D. bryoniae into two major genotypes, RAPD Group (RG) I and RG II. A recently developed real-time PCR assay for D. bryoniae specific only for isolates with the RG I genotype has limitations in practical disease diagnosis. The objective of the present study was to develop a new set of primers and Tay Man probe in conjunction with real-time PCR to detect both genotypes. This new Didymella detection system generated strong Ct values to a worldwide collection of D. bryoniae including 105 RG I, 13 RG II and 7 unassigned isolates. This broad spectrum and specific real-time PCR system could be used in routine seed and plant health diagnosis for D. bryoniae.