Development of molecular assays for detection of Stenocarpella maydis and Stenocarpella macrospora in corn

The causal agents of Diplodia ear rot are two species of the Stenocarpella genus, S. macrospora and S. maydis. In addition to ears, both pathogens can infect leaves and stalks, and both are present in most cornproduction regions around the world. It is difficult to visually distinguish between the two pathogens based on plant symptoms and fungal signs. To facilitate accurate and rapid pathogen identification, polymerase chain reaction (PCR) assays were developed for identification of each species. Species-specific primers of 18 to 20 nucleotides in length were designed, targeting a portion of the internal transcribed spacer (ITS) region of the fungal genome for conventional and real-time PCR assays. The conventional PCR method successfully amplified a single 1.7-kb and 800-bp fragment for each S. maydis and S. macrospora isolate, respectively. The real-time method was performed using SYBR green dye, and detection of each specific target pathogen was successfully obtained. In total, 82 S. maydis and 15 S. macrospora isolates were tested to evaluate the reproducibility of these primers. Both methods provide a rapid and specific tool for the detection of Stenocarpella spp.