Plantago asiatica mosaic virus (PlAMV), which infects lilies worldwide, constitutes a potential risk factor for lily production. Development of effective and sensitive methods for the detection and specific quantification of PlAMV is required to prevent the spread of PlAMV in lilies. In this study, we established a sensitive and accurate quantification method for PlAMV infection in lily samples using a reverse transcription droplet digital polymerase chain reaction (RT-ddPCR) assay. The specificity and sensitivity of the proposed RT-ddPCR assay was evaluated for precise detection and quantification of PlAMV. The assay exhibited high specificity for PlAMV and showed no cross-reactivity with other lily viruses. It was about 10-fold more sensitive than reverse-transcription followed by quantitative polymerase chain reaction (RT-qPCR). RT-ddPCR (R² = 0.9985) and RT-qPCR (R² = 0.9869) showed high degrees of linearity. In addition, field validation tests on 92 lily samples confirmed that RT-ddPCR (45/92) has higher sensitivity than RT-qPCR (8/92). In summary, RT-ddPCR outperforms RT-qPCR in the analysis of low-viral-load samples and can be utilized for PlAMV infection diagnosis, especially in plant quarantine inspection and PlAMV-free certification programs.

车前草花叶病毒 (PlAMV) 感染全世界的百合，是百合生产的潜在风险因素。需要开发有效和灵敏的方法来检测和特定量化 PlAMV，以防止 PlAMV 在百合中传播。在这项研究中，我们使用逆转录液滴数字聚合酶链反应建立了一种灵敏准确的百合样品 PlAMV 感染定量方法(RT-ddPCR) 测定。评估了所提出的 RT-ddPCR 测定的特异性和灵敏度，以精确检测和定量 PlAMV。该测定对 PlAMV 表现出高度特异性，并且与其他百合病毒没有交叉反应。它比逆转录后进行定量聚合酶链反应 (RT-qPCR) 的灵敏度高约 10 倍。RT-ddPCR ( R ²  = 0.9985) 和 RT-qPCR ( R ² = 0.9869) 表现出高度的线性。此外，对 92 个百合样品的现场验证测试证实，RT-ddPCR (45/92) 比 RT-qPCR (8/92) 具有更高的灵敏度。总之，RT-ddPCR 在低病毒载量样本分析方面优于 RT-qPCR，可用于 PlAMV 感染诊断，特别是在植物检疫检验和无 PlAMV 认证项目中。