Abstract Verticillium wilt is a major disease of olive (Olea europaea) orchards worldwide. To control the disease, use of a reliable method for pre-planting screening of new planting stocks for infection by V. dahliae is essential. In this study, 20 symptomatic and asymptomatic olive trees, sampled from Zahak region of Sistan in Iran, were analysed with real-time and nested PCR for in planta detection of V. dahliae. We could confirm the presence of V. dahliae in 12 out of 20 (60%) sampled trees using real-time PCR, whereas nested PCR produced amplicons for all tested symptomatic and asymptomatic samples (100% of tested trees), indicating that nested PCR was more efficient method than real-time PCR for in planta detection of V. dahliae. We used the designed nested PCR to determine also the pathotype of V. dahliae using Vdf2/Vdr2 primer set at the second round, which could produce the 297-bp amplicon for all tested samples infected with non-defoliating pathotype of V. dahliae. We showed that although the Verticillium wilt is spread in the olive cultivation areas in Zahak region of Sistan, nested PCR can be deployed for the timely detection and thereby prevention of the further spread of V. dahliae in the olive cultivation areas.

中文翻译：

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使用实时 PCR 和巢式 PCR 鉴定和植物内检测受感染橄榄树中大丽轮枝菌的可靠方法

摘要 黄萎病是世界范围内橄榄（Olea europaea）果园的主要病害。为了控制该病害，必须使用可靠的方法对新种植种群进行种植前筛选以防止大丽花弧菌感染。在这项研究中，对来自伊朗锡斯坦 Zahak 地区的 20 棵有症状和无症状的橄榄树进行了实时和巢式 PCR 分析，用于在植物中检测大丽花。我们可以使用实时 PCR 确认 20 棵采样树中有 12 棵（60%）存在大丽花，而巢式 PCR 为所有测试的有症状和无症状样本（100% 的测试树）产生了扩增子，表明巢式 PCR是比实时 PCR 更有效的方法在大丽花的植物中检测。我们还使用设计的巢式 PCR 来确定 V 的病理类型。大丽花在第二轮使用 Vdf2/Vdr2 引物组，这可以为所有受大丽花非脱叶病型感染的测试样品产生 297-bp 扩增子。我们表明，虽然黄萎病在锡斯坦扎哈克地区的橄榄种植区传播，但可以部署嵌套 PCR 以及时检测，从而防止大丽花在橄榄种植区的进一步传播。