Rapid and Sensitive Detection of Toxigenic Fusarium asiaticum Integrating Recombinase Polymerase Amplification, CRISPR/Cas12a, and Lateral Flow Techniques.

Fusarium head blight (FHB) is a global cereal disease caused by a complex of Fusarium species. Both Fusarium graminearum and F. asiaticum are the causal agents of FHB in China. F. asiaticum is the predominant species in the Middle-Lower Reaches of the Yangtze River (MLRYR) and southwest China. Therefore, detecting F. asiaticum in a timely manner is crucial for controlling the disease and preventing mycotoxins from entering the food chain. Here, we combined rapid genomic DNA extraction, recombinase polymerase amplification, Cas12a cleavage, and lateral flow detection techniques to develop a method for the rapid detection of F. asiaticum. The reaction conditions were optimized to provide a rapid, sensitive, and cost-effective method for F. asiaticum detection. The optimized method demonstrated exceptional specificity in detecting F. asiaticum while not detecting any of the 14 other Fusarium strains and 3 non-Fusarium species. Additionally, it could detect F. asiaticum DNA at concentrations as low as 20 ag/µL, allowing for the diagnosis of F. asiaticum infection in maize and wheat kernels even after 3 days of inoculation. The developed assay will provide an efficient and robust detection platform to accelerate plant pathogen detection.

Species Diversity and Chemotypes of Fusarium Species Associated With Maize Stalk Rot in Yunnan Province of Southwest China.

Maize stalk rot caused by Fusarium species is one of the most important fungal diseases of maize throughout the world. The disease is responsible for considerable yield losses and has also been associated with mycotoxin contamination of the crop. In this study, a survey of maize stalk rot was performed in seven locations of Yunnan Province in China during the cropping season of 2015 and 2016. Based on morphological and molecular characteristics, 204 isolates belonging to 12 Fusarium spp. from symptomatic stalks of maize were identified. Among the isolated strains, 83 were identified as Fusarium meridionale (40.5%), 46 as Fusarium boothii (22.5%), 34 as Fusarium temperatum (16.5%), 12 as Fusarium equiseti (5.9%), 10 as Fusarium asiaticum (4.9%), six as Fusarium proliferatum (3.0%), four as Fusarium verticillioides (2.0%), four as Fusarium incarnatum (2.0%), two as Fusarium avenaceum (1.0%), one as Fusarium cerealis (0.5%), one as Fusarium graminearum (0.5%), and one as Fusarium cortaderiae (0.5%). Fusarium cortaderiae was the first report on the causal agent of maize stalk rot disease in China. These isolates were divided into five chemotypes: nivalenol (NIV), deoxynivalenol (DON), beauvericin (BEA), zearalenone (ZEN), and fumonisin (FUM). Phylogenetic analysis based on partial sequences of the translation elongation factor 1α (TEF1-α) showed a high degree of interspecific polymorphisms among the isolates. Pathogenicity analysis on maize stalks indicated that all the 12 species of Fusarium were able to cause the disease symptoms with different aggressiveness. This study on population, pathogenicity, and toxigenic chemotypes of Fusarium species associated with maize stalk rot in Yunnan Province of southwest China, will help design an effective integrated control strategy for this disease.