Zhang Yubao, Wang Yajun, Xie Zhongkui, Wang Ruoyu, Guo Zhihong, He Yuhui
Northwest Institute of Eco-Environment and Resources, Chinese Academy of Sciences, Lanzhou 730000, China.
Plant Pathol J. 2020 Apr 1;36(2):170-178. doi: 10.5423/PPJ.OA.04.2019.0096.
The (LMoV) impedes the growth and quality of lily crops in Lanzhou, China. Recently (ArMV) has been detected in LMoV-infected plants in this region, causing plant stunting as well as severe foliar symptoms, and likely posing a threat to lily production. Consequently, there is a need to develop simple, sensitive, and reliable detection methods for these two viruses to prevent them from spreading. Reverse transcription (RT) loop-mediated isothermal amplification (LAMP) assays have been developed to detect LMoV and ArMV using two primer pairs that match six conserved sequences of LMoV and ArMV coat proteins, respectively. RT-LAMP assay results were visually assessed in reaction tubes using green fluorescence and gel electrophoresis. Our assays successfully detected both LMoV and ArMV in lily plants without the occurrence of viral cross-reactivity from other lily viruses. Optimal conditions for LAMP reactions were 65°C and 60°C for 60 min for LMoV and ArMV, respectively. Detection sensitivity for both RT-LAMP assays was a hundredfold greater than that of our comparative RT-polymerase chain reaction assays. We have also found this relatively rapid, target specific and sensitive method can also be used for samples collected in the field and may be especially useful in regions with limited or no laboratory facilities.
百合斑驳病毒（LMoV）阻碍了中国兰州百合作物的生长并影响其品质。最近，在该地区感染LMoV的植物中检测到了建兰花叶病毒（ArMV），它会导致植株矮化以及严重的叶片症状，可能对百合生产构成威胁。因此，需要开发针对这两种病毒的简单、灵敏且可靠的检测方法，以防止它们传播。已开发出逆转录（RT）环介导等温扩增（LAMP）检测方法，使用分别与LMoV和ArMV外壳蛋白的六个保守序列匹配的两对引物来检测LMoV和ArMV。使用绿色荧光和凝胶电泳在反应管中直观评估RT-LAMP检测结果。我们的检测方法成功检测到了百合植株中的LMoV和ArMV，且未出现与其他百合病毒的病毒交叉反应。LAMP反应的最佳条件是，检测LMoV时为65°C 60分钟，检测ArMV时为60°C 60分钟。两种RT-LAMP检测方法的检测灵敏度均比我们的比较RT-聚合酶链反应检测方法高一百倍。我们还发现，这种相对快速、靶向特异性且灵敏的方法也可用于现场采集的样本，在实验室设施有限或没有实验室设施的地区可能特别有用。
