Development of a Real-Time Quantitative PCR Assay for Direct Detection and Quantification of the Root-Lesion Nematode Pratylenchus penetrans in Potato Roots

The root-lesion nematode, <i>Pratylenchus penetrans</i>, is a migratory endoparasite that attacks potato roots, causing necrotic lesions and yield losses of up to 73%. Traditional detection and quantification methods are labor-intensive, time-consuming, and require prior nematode extraction and taxonomic expertise. This study aimed to develop a SYBR Green-based real-time quantitative PCR (qPCR) assay for detecting and quantifying <i>P. penetrans</i> directly from potato root DNA extracts. Bovine serum albumin (BSA) was tested to neutralize qPCR inhibitors in root DNA extracts. The assay showed high specificity and sensitivity to <i>P. penetrans</i>, detecting 1.56 × 10^-2 of a single nematode in 0.2 g of roots. A standard curve based on artificial nematode inoculations demonstrated a strong linear relationship between Cq values and log-transformed nematode numbers (<i>R</i>² = 0.993) with high amplification efficiency. Assessment using root samples from two greenhouse experiments involving five potato cultivars showed strong correlations (<i>r</i> = 0.902 and 0.887) between qPCR estimates and microscopic nematode counts. This study presents a new qPCR assay specifically optimized for direct detection and quantification of <i>P. penetrans</i> within potato root tissues, offering enhanced sensitivity and applicability for rapid in planta diagnostics to facilitate effective management strategies.