A rapid real-time PCR assay for detecting Microdochium paspali causing sparse leaf patch on seashore paspalum and in environmental samples.

ABSTRACT

BACKGROUND: Sparse leaf patch (SLP) is one of the most significant diseases affecting seashore paspalum (Paspalum vaginatum Sw.), caused by Microdochium paspali. Fast and accurate detection of this pathogen is crucial for effective disease management. However, conventional culture-based methods are time-consuming and often compromised by the presence of other saprophytic or endophytic fungi. RESULTS: In this study, we developed a real-time fluorescent quantitative (q)PCR method based on the internal transcribed spacer (ITS) region of the ribosomal RNA gene to rapidly detect and quantify M. paspali. The qPCR assay demonstrated the ability to detect all 12 tested isolates of M. paspali, with no cross-reactions observed when tested against 30 isolates of other fungal pathogens from turfgrass samples. The detection limit of the qPCR method was as low as 3.65 × 102 copies µL-1 of M. paspali genomic DNA, and the entire detection process could be completed within 1 h. The fluorescence signal was detectable in the leaf tissues of seashore paspalum without apparent disease symptoms as early as 24 h postinoculation with M. paspali. Moreover, the qPCR method successfully detected M. paspali in both asymptomatic and symptomatic turfgrass samples, including leaf, stem, root and rhizosphere soil, indicating that this assay can significantly enhance the detection of M. paspali. CONCLUSION: The study developed a rapid real-time qPCR assay for the detection of M. paspali causing SLP on seashore paspalum and in environmental samples, which has important implications for early warning and management of SLP. © 2024 Society of Chemical Industry.