Patulin [KLH]

Dissemination and survival of ascomycetes is through asexual spores. ThebrlAgene encodes a C2H2-type zinc-finger transcription factor, which is essential for asexual development.Penicillium expansumcauses blue mold disease and is the main source of patulin, a mycotoxin that contaminates apple-based food. AP. expansumPe Delta brlAdeficient strain was generated by homologous recombination. In vivo, suppression ofbrlAcompletely blocked the development of conidiophores that takes place after the formation of coremia/synnemata, a required step for the perforation of the apple epicarp. Metabolome analysis displayed that patulin production was enhanced bybrlAsuppression, explaining a higher in vivo aggressiveness compared to the wild type (WT) strain. No patulin was detected in the synnemata, suggesting that patulin biosynthesis stopped when the fungus exited the apple. In vitro transcriptome analysis of Pe Delta brlAunveiled an up-regulated biosynthetic gene cluster (PEXP_073960-PEXP_074060) that shares high similarity with the chaetoglobosin gene cluster ofChaetomium globosum. Metabolome analysis of Pe Delta brlAconfirmed these observations by unveiling a greater diversity of chaetoglobosin derivatives. We observed that chaetoglobosins A and C were found only in the synnemata, located outside of the apple, whereas other chaetoglobosins were detected in apple flesh, suggesting a spatial-temporal organization of the chaetoglobosin biosynthesis pathway.

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.