Development and validation of a LAMP-based method for rapid and reliable detection of , the causal agent of sugarcane leaf scald.

INTRODUCTION

(Xalb)-induced leaf scald (LS) is a significant bacterial disease affecting sugarcane and posing a global threat to the sugarcane industry. The presence of irregular symptoms makes traditional phenotypic detection difficult, and molecular methods necessitate costly equipment, labor, and extended sample-to-answer processing times.

METHODS

This study introduces an innovative rapid DNA isolation method requiring no reagents, combined with an isothermal amplification-based assay for efficient detection of DNA in sugarcane xylem sap, leaf tissue, and meristematic tissue samples. Sugarcane samples from infected plants were subjected to heat lysis, followed by loop-mediated isothermal amplification (LAMP)-based fluorescence and colorimetric quantification within a single microcentrifuge tube.

RESULTS

The method exhibited exceptional detection sensitivity (detecting as low as 1 cell/μL), reproducibility [with a standard deviation (SD) of <5% for = 3], and a broad linear dynamic range (10 pM to 1 aM or 10-10 copies/μL, = 0.99). Quantification of was accurately correlated with sugarcane cultivar disease ratings. Validation using qPCR showed 91-98% agreement. This assay also effectively determined optimal sampling times and plant parts by monitoring the progression of the disease over time.

DISCUSSION

This diagnostic assay holds significant potential as a commercial opportunity for a kit-based DNA extraction/purification-free molecular detection alternative. It can be adapted into a handheld device, enabling on-farm detection and quantification of the pathogen responsible for LS disease.