Development and optimization of a group-specific loop-mediated isothermal amplification (LAMP) assay for the detection of patulin-producing Penicillium species

摘要

The mycotoxin patulin is a toxic fungal secondary metabolite occurring in food worldwide. Methods for rapid, simple and specific detection of patulin-producing fungi in food and feed are therefore urgently needed. In the current study, a loop-mediated isothermal amplification (LAMP) assay based on the isoepoxydon dehydrogenase (idh) gene of the patulin biosynthetic pathway was developed and optimized for the group-specific detection of patulin-producing Penicillium species. By testing purified DNA of 174 fungal strains representing 31 genera, the assay was demonstrated to be highly specific for the detection of patulin-producing species in Penicillium, Byssochlamys and Paecilomyces. The assay had a detection limit of 2.5 pg of purified genomic DNA of P. expansum per reaction. Moreover, the assay was demonstrated to detect patulin-producers when conidia were directly added to the master mix as template without any sample preparation. The applicability of the assay in food analyses was successfully tested on artificially contaminated grapes and apples requiring minimal sample preparation. A screening of grapes from the 2018 harvest from different locations in Germany revealed no presence of patulin-producers. The developed LAMP assay is a promising tool for rapid diagnosis in quality control applications in the food and beverage industry.