Development of a new LAMP assay for the fast diagnosis of Gnomoniopsis castaneae

Gnomoniopsis castaneae (syn. Gnomoniopsis smitholgivyi) is an emerging fungal pathogen that causes nut rot in sweet chestnut (Castanea sativa). This gnomoniaceous fungus is also reported to be associated with cankers on twigs as well as with gall and leaf necroses. Nut rot disease was observed in northern Italy since the second half of the 19th century, but only in the last ten years the incidence and severity of nut rot was associated with outbreaks by G. castaneae. If, on the one hand, very little is still known about the epidemiology of the disease, the almost ubiquitous distribution of the fungus in chestnut groves and its endophytic occurrence in chestnut organs (shoots, foliage, fruits) make it difficult to find valid solutions for disease management and control. The possibility of a fast and accurate diagnosis could represent a valuable tool to monitor pathogen occurrence both in pre- and in post-harvest conditions. Among the strategies today available to counter the disease, molecular detection tools are the most effective for disease diagnosis with high sensitivity and specificity. However, molecular detection often requires a well-equipped laboratory to be applied, and this limits the speed and user-friendliness of the diagnosis. The aim of this work was to develop a new LAMP-based tool to be applied directly on site for the early detection of G. castaneae. The assay, optimized on the portable instrument Genie III (Optigene, UK) and based on the Ef1-a target region, revealed the ability to recognize the pathogen with a high level of specificity and sensitivity in about 20 minutes. Application of this method in chestnut orchards and in the subsequent processing steps of the fruit and derived products (e.g. chestnut flour) might provide a new, intriguing perspective for disease prevention and control directly in the field and in the chestnut processing chain.