Thladiantha dubia mosaic virus (ThDMV) from the wild Manchurian tuber gourd (Thladiantha dubia Bunge, Cucurbitaceae), is a novel tentative specie of the genus Potyvirus within the family Potyviridae. It may pose a potential risk to both cultivated crops and plants in the natural ecosystem. In the present study, we developed an ELISA procedure for detection of ThDMV. Coat protein (CP) of ThDMV was successfully expressed in Escherichia coli, and used to immunize rabbit for antiserum after being purified by using Ni-NTA affinity column. Results of indirect ELISA showed that the optimum dilutions of the coating antigen and antiserum were 1:160 (1 μg per well), and 1:2000 respectively. Western blot indicated that antibodies in the antiserum could specifically recognize and bind CP of ThDMV. An ELISA method was developed with the antiserum to detect ThDMV in plant extracts. Samples collected from potato fields in Northeast China and Northwest China were analyzed by ELISA and RT-PCR and both gave consistent results. The ELISA assay developed here has potential to be used as a sensitive, reliable and cost-effective tool for ThDMV detection in epidemiological studies.

来自野生东北瓜（Thladiantha dubia Bunge，葫芦科）的Thladiantha dubia花叶病毒（ThDMV）是Potyviridae家族中Potyvirus属的新型暂定物种。它可能对自然生态系统中的农作物和植物都构成潜在风险。在本研究中，我们开发了用于检测ThDMV的ELISA程序。ThDMV的外壳蛋白（CP）在大肠杆菌中成功表达Ni-NTA亲和柱纯化后用于免疫兔抗血清。间接ELISA的结果表明，包被抗原和抗血清的最佳稀释度分别为1：160（每孔1μg）和1：2000。Western印迹表明抗血清中的抗体可以特异性识别并结合ThDMV的CP。开发了一种使用抗血清的ELISA方法来检测植物提取物中的ThDMV。通过ELISA和RT-PCR对东北和西北马铃薯田的样品进行了分析，结果均一致。此处开发的ELISA测定法有可能被用作流行病学研究中ThDMV检测的灵敏，可靠且经济高效的工具。