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<p><i>Foorkey</i> and <i>chirke</i> are the two major viral diseases impacting large cardamom productivity, caused by cardamom bushy dwarf virus (CBDV), a DNA virus, and large cardamom chirke virus (LCCV), an RNA virus, respectively. Relying solely on visible symptoms for diagnosing these infections is unreliable, as infected plants may not always exhibit clear symptoms. Therefore, rapid, accurate, and sensitive detection methods are crucial for identifying and propagating virus-free plants. Conventional laboratory-based techniques such as polymerase chain reaction (PCR) and real-time PCR, while effective, are time-consuming and require specialized equipment. In contrast, isothermal methods like recombinase polymerase amplification (RPA) offer faster results without the need for advanced laboratory infrastructure and are well suited for onsite diagnostics, especially when combined with lateral flow assay (LFA). In this study, an RPA-LFA assay was developed for detecting CBDV and an RT-RPA-LFA assay for LCCV in large cardamom, using TwistAmp reagents, labeled primers, and crude extracts from infected plants. Optimization of the assay using 1&#xa0;M betaine eliminated false positives by suppressing non-specific amplification. The optimized assays demonstrated either comparable (CBDV) or tenfold higher sensitivity (LCCV) than conventional PCR. The entire workflow could be completed within 35–45&#xa0;min, making them suitable for large-scale indexing of planting material. Both assays were successfully validated using field-collected samples of large cardamom that included asymptomatic and mixed infections.</p>

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Rapid on-site detection of cardamom bushy dwarf virus and large cardamom chirke virus using recombinase polymerase amplification coupled lateral flow assays

  • 期刊论文

  • 发表日期：

    2026年3月19日

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  • Pradeep Malavika,
  • Alangar Ishwara Bhat,
  • Malayathodi Greeshma

摘要

Foorkey and chirke are the two major viral diseases impacting large cardamom productivity, caused by cardamom bushy dwarf virus (CBDV), a DNA virus, and large cardamom chirke virus (LCCV), an RNA virus, respectively. Relying solely on visible symptoms for diagnosing these infections is unreliable, as infected plants may not always exhibit clear symptoms. Therefore, rapid, accurate, and sensitive detection methods are crucial for identifying and propagating virus-free plants. Conventional laboratory-based techniques such as polymerase chain reaction (PCR) and real-time PCR, while effective, are time-consuming and require specialized equipment. In contrast, isothermal methods like recombinase polymerase amplification (RPA) offer faster results without the need for advanced laboratory infrastructure and are well suited for onsite diagnostics, especially when combined with lateral flow assay (LFA). In this study, an RPA-LFA assay was developed for detecting CBDV and an RT-RPA-LFA assay for LCCV in large cardamom, using TwistAmp reagents, labeled primers, and crude extracts from infected plants. Optimization of the assay using 1 M betaine eliminated false positives by suppressing non-specific amplification. The optimized assays demonstrated either comparable (CBDV) or tenfold higher sensitivity (LCCV) than conventional PCR. The entire workflow could be completed within 35–45 min, making them suitable for large-scale indexing of planting material. Both assays were successfully validated using field-collected samples of large cardamom that included asymptomatic and mixed infections.