A DNA Adsorption-Based Biosensor for Rapid Detection of Ratoon Stunting Disease in Sugarcane

Early and accurate detection of plant diseases is critical for ensuring global food security and agricultural resilience. Ratoon stunting disease (RSD), caused by the bacterium <i>Leifsonia xyli</i> subsp. <i>xyli</i> (<i>Lxx</i>), is among the most economically significant diseases of sugarcane worldwide. Its cryptic nature-characterized by an absence of visible symptoms-renders timely diagnosis particularly difficult, contributing to substantial undetected yield losses across major sugar-producing regions. Here, we report the development of a potential-induced electrochemical (EC) nanobiosensor platform for the rapid, low-cost, and field-deployable detection of <i>Lxx</i> DNA directly from crude sugarcane sap. This method eliminates the need for conventional nucleic acid extraction and thermal cycling by integrating the following: (i) a boiling lysis-based DNA release from xylem sap; (ii) sequence-specific magnetic bead-based purification of <i>Lxx</i> DNA using immobilized capture probes; and (iii) label-free electrochemical detection using a potential-driven DNA adsorption sensing platform. The biosensor shows exceptional analytical performance, achieving a detection limit of 10 cells/µL with a broad dynamic range spanning from 10⁵ to 1 copy/µL (r = 0.99) and high reproducibility (SD < 5%, n = 3). Field validation using genetically diverse sugarcane cultivars from an inoculated trial demonstrated a strong correlation between biosensor signals and known disease resistance ratings. Quantitative results from the EC biosensor also showed a robust correlation with qPCR data (r = 0.84, n = 10, <i>p</i> < 0.001), confirming diagnostic accuracy. This first-in-class EC nanobiosensor for RSD represents a major technological advance over existing methods by offering a cost-effective, equipment-free, and scalable solution suitable for on-site deployment by non-specialist users. Beyond sugarcane, the modular nature of this detection platform opens up opportunities for multiplexed detection of plant pathogens, making it a transformative tool for early disease surveillance, precision agriculture, and biosecurity monitoring. This work lays the foundation for the development of a universal point-of-care platform for managing plant and crop diseases, supporting sustainable agriculture and global food resilience in the face of climate and pathogen threats.