One-hour loop-mediated isothermal amplification assay for the detection of quarantinable toxigenic Fusarium garmanirum

Abstract

Fusarium head blight (FHB) caused by several Fusarium species is one of the most serious diseases affecting wheat throughout the world. The in vitro production of the toxins deoxynivalenol, zearalenone fumonisin, T-2, and HT-2 was quantitvely evaluated in 8 different isolates of Fusarium species collected from feed samples. It was possible to detect zearalenone and the other mycotoxins in 100% and 50% of the isolates, respectively. In the present study, loop-mediated isothermal amplification method (LAMP) was designed for diagnosing Fusarium garmanirum infections and testing against feed samples, infested samples and pure cultures. The LAMP amplicon was directly visualized in the reaction tubes by the naked eye following the addition of calcein fluorescence. The LAMP products appeared as DNA marker pattern, with many bands of different sizes from 145 base pairs up to the loading well. Loop-LAMP procedure was used to detect genomic DNA of F. graminearum in fungal pure culture and in contaminated feed samples. In the future, this assay will support plant quarantine programs in Saudi Arabia and Gulf Cooperation Council states, to prevent the introduction of foreign FHB species.