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POSTERS: Disease Detection and Diagnosis II

P-216 - A universal primer pair for real-time and simultaneous detection of six Solanaceae plants infected viroids

Tuesday, August 5, 2025
9:30 AM - 10:15 AM HST
Location: Exhibit Hall 3

    Presenting Author(s)

  • HC

    Ho-Hsiung Chang

    Assistant Professor
    Department of Plant Pathology, National Chung Hsing University
    Taichung, Taichung, Taiwan (Republic of China)

    • Author(s)

  • JH

    Jui-Chen Hung

    Department of Plant Pathology, National Chung Hsing University
    Taichung, Taichung, Taiwan (Republic of China)

  • FJ

    Fuh-Jyh Jan

    Professor
    Department of Plant Pathology, National Chung Hsing University
    Taichung, Taichung, Taiwan (Republic of China)

Abstract Text: Viroids cause significant damage to agriculture worldwide. Six pospiviroids, Columnea latent viroid, pepper chat fruit viroid, potato spindle tuber viroid, tomato apical stunt viroid, tomato chlorotic dwarf viroid, and tomato planta macho viroid, are listed as quarantine pathogens in many countries due to their being seed-transmissible nature and the severe damage they caused in fields. In this study, we aimed to develop a real-time, simultaneous detection method for these six viroids using a universal primer pair. Based on conserved regions of these viroids, two target sequences were identified: a long amplicon (187 bp) and a short amplicon (110 bp) for primer design. RT-qPCR was conducted on tobacco samples infected with viroids. The results indicated that the primer pair targeting the short amplicon demonstrated higher stability and specificity at a reaction temperature of 60°C. Sensitivity analysis using total tomato cDNA as the reaction background, mixed with viroid DNA constructs, showed that qPCR with this primer set could detect as few as 10,000 viroid molecules. In practical tests with artificially inoculated tomato samples, the detection system successfully identified viroids, even in samples that did not exhibit obvious disease symptoms. This system can also be applied to tomato seeds to detect the presence of viroids. Although sequencing of PCR products is still needed to confirm viroid species, the system developed in this study can simultaneously detect six seed-borne quarantne pospiviroids in Solanaceae plants, and seeds intended for import and export.