A one‐step multiplex PCR assay for the detection and differentiation of four species of Clarireedia causing dollar spot on turfgrass

BACKGROUND Dollar spot (DS) is one of the most destructive and economically important diseases of cool‐ and warm‐season turfgrasses worldwide. A total of six species causing DS disease in the genus Clarireedia have been described, and four of them have been reported to be distributed countrywide in China. Identification of different species of Clarireedia is a prerequisite for the effective management of DS disease. RESULTS Here we report a novel polymerase chain reaction (PCR)‐based method for the detection and differentiation of the four species of Clarireedia associated with DS on turfgrass in China: C. jacksonii, C. paspali, C. monteithiana and C. hainanense. Species‐specific genes were identified for each species by comparative genomics analysis. Four primer pairs were designed and mixed to amplify species‐specific PCR fragments with differential sizes for the four species of Clarireedia in a single multiplex PCR assay. No PCR products were generated from the DNA templates of other common fungal pathogens associated with multiple turfgrass diseases. The multiplex PCR method developed can be used for the rapid and accurate detection and differentiation of the four species of Clarireedia from pure cultures as well as from infected turfgrass blades with DS symptoms. CONCLUSION The study developed a one‐step multiplex PCR assay for the detection and differentiation of four species of Clarireedia causing DS on turfgrass in China, which will have important implications for DS management in China and worldwide. © 2022 Society of Chemical Industry. A one‐step multiplex PCR assay for the detection and differentiation of the four species of Clarireedia associated with dollar spot on turfgrass in China: C. jacksonii, C. paspali, C. monteithiana and C. hainanense with species‐specific PCR fragments under differential sizes.