Quantitative Detection of Phytophthora infestans in Potato Leaves Using Nanopore Amplicon Sequencing

Potato late blight, caused by Phytophthora infestans, is a devastating disease affecting potato cultivation worldwide, causing significant yield losses. Early and accurate detection of P. infestans is crucial for efficient disease management. Molecular techniques, such as qPCR, are specific and reliable, while next-generation sequencing approaches can provide broader data and are rapidly improving in their detection and user-friendly potential. In this study, we tested Oxford Nanopore Technologies (ONT) amplicon sequencing as a quantitative method for detecting and monitoring P. infestans in artificially inoculated potato plants. The outcome of the infection was then compared between resistant (cv. Inova) and susceptible (cv. Agata) cultivars, and relative abundance of P. infestans reads obtained through ONT sequencing was validated via qPCR and visual symptom evaluation. By using an Internal Transcribed Spacer (ITS) metabarcoding approach, MinION device and a customized pipeline, Nanopore sequencing proved able to detect the presence of P. infestans at early and asymptomatic stages of the infection, showing a progressive increase of reads from 2 to 7 days post-infection. Furthermore, our approach successfully differentiated different levels of infection between susceptible and resistant cultivars, showing an abundance of reads mapping P. infestans correlated with qPCR quantification and symptomatology. This work highlights the feasibility of Nanopore sequencing as a diagnostic tool, complementing more traditional molecular methods.