Detecting Phytophthora capsici (P. capsici) based solely on visual symptoms is challenging and often leads to misdiagnosis. Farmers frequently harvest seemingly healthy fruits, only for fruit rot to develop after shipping. Furthermore, other pathogens can mimic P. capsici symptoms on cucurbits and peppers, and plants may even suffer simultaneous attacks by multiple pathogens, complicating identification. Without timely and accurate diagnosis, P. capsici can spread rapidly, causing significant crop losses. Current diagnostic methods, including traditional microscopy-based culture techniques and polymerase chain reaction (PCR), are time-intensive and lack sensitivity for early-stage infections. This study introduces an optimized Oxford Nanopore Technology (ONT) genomic approach for rapid and precise detection of P. capsici in plant samples, both in laboratory and field settings. Designed for portability and capable of sequencing reads up to 100 kb, the ONT MinION device—smaller than a smartphone—provides a promising solution for in-field diagnostics. Plant tissue samples, symptomatic and non-symptomatic, were collected from cucurbit and pepper fields through collaborators during late summer and early fall. Total DNA was extracted using a magnetic bead-based kit (Primerdesign, Southampton, UK). Sequencing libraries were prepared using ONT’s 1D-cDNA sequencing kit, loaded onto a MinION 107 v9.5 Flow Cell, and analyzed using the Mk1B MinION device. Raw sequence reads in fast5 format were converted to fastq or fasta, with high-quality reads subjected to BLAST searches against the NCBI database for P. capsici identification. The deployment of ONT enables the generation of actionable genomic data in real-time, enhancing our understanding of P. capsici and its role in Phytophthora blight disease development in cucurbits. This technology represents a breakthrough in the rapid, field-based diagnosis of P. capsici, providing farmers with an efficient tool to mitigate crop losses. Keywords: Oxford Nanopore Sequencing Technology (ONT), Phytophthora capsici, Raw sequence reads, Phytophthora blight.
