Rapid detection of Pythium soft rot in ginger ( <i>Zingiber officinale</i> ) caused by <i>Pythium myriotylum</i> using LAMP

Pythium soft rot in ginger is caused by <i>Pythium myriotylum</i>, leading to significant reductions in ginger yields. In this study, a loop-mediated isothermal amplification (LAMP) detection method was established for <i>Py. myriotylum</i> in infected ginger plants and infested soil. The <i>Py. myriotylum</i> genome was compared with the genomes of seven additional oomycete species to obtain species-specific sequences. LAMP primer sets were designed and assessed at temperatures from 58°C to 68°C and tested for their specificity. Furthermore, the sensitivity of the screened primer set was evaluated, and the detection limit was 1 pg of <i>Py. myriotylum</i> genomic DNA (gDNA) in a 25-µl reaction volume. LAMP was used to detect <i>Py. myriotylum</i> zoospores in solution (1×10²/ml), vermiculite (1×10²/g), and soil (1×10⁴/g). A rapid and portable gDNA extraction method from ginger seedlings and rhizomes using solid-phase reversible immobilization beads for purification was adopted. The LAMP assay could reliably detect <i>Py. myriotylum</i> in artificially inoculated diseased ginger seedlings and rhizosphere vermiculite. The portable gDNA extraction method coupled with the LAMP assay facilitates point-of-care pathogen detection, requiring 2 to 3 hours for eight samples, greatly reducing the time needed for <i>Py. myriotylum</i> detection of Pythium soft rot in ginger.