Research Centre for Agrobiology and Pedology (CRA-ABP), Consiglio per la Ricerca e Sperimentazione in Agricoltura, via di Lanciola 12/a, 50125, Florence, Italy.
Exp Appl Acarol. 2014 Feb;62(2):161-70. doi: 10.1007/s10493-013-9733-3. Epub 2013 Sep 13.
Trisetacus juniperinus (Nalepa) sensu Keifer (Acari: Eriophyoidea: Phytoptidae) causes irregular development of buds, shoot deformations and stunted growth of trees, resulting in a serious threat to nurseries and young stands of Cupressus sempervirens L. (Mediterranean cypress). Recently, some cypress clones selected for their resistance to the fungal canker agent Seiridium cardinale (Wag.) have shown high susceptibility to the mite. Considering its tiny body, its hidden lifestyle inside the buds and the probable occurrence of other species (the vagrant Epitrimerus cupressi (Keifer) is common on the Mediterranean cypress in Italy), detection and monitoring of T. juniperinus require taxonomic expertise and are often time-consuming and challenging before serious damage is discernible. In the present study, a rapid, cost-effective PCR-based method was developed and validated to detect T. juniperinus on cypresses. The cytochrome c oxidase subunit I gene was amplified with degenerate and specific primers, but the latter were the only ones able to discriminate between T. juniperinus and E. cupressi. PCR products distinguished the two species both in a pool of individuals in a mixed population of both species and in single individuals, indicating the sensitivity of the detection method. PCR-RFLP (restriction fragment length polymorphism) by means of XmnI and XbaI endonucleases separated the two species. Furthermore, a washing-sieving protocol was used to make mite collection from the tree sample faster and simpler; this procedure did not interfere with the molecular detection of the species. The possibility of the routine use of this assay to monitor quarantine eriophyoids infesting plant material is discussed.
三齿小爪螨（Nalepa）（蜱螨目：叶螨科）会导致芽发育异常、嫩枝变形和树木生长受阻，对柏树苗圃和幼林构成严重威胁。最近，一些柏树苗因对真菌溃疡病原体丝核菌（Seiridium cardinale（Wag.））具有抗性而被选择，但它们对该螨的敏感性较高。考虑到其微小的体型、在芽内隐藏的生活方式以及可能存在其他物种（意大利地中海柏上常见的流浪者柏小爪螨（Epitrimerus cupressi（Keifer））），检测和监测三齿小爪螨需要分类学专业知识，并且在明显损害发生之前，通常既耗时又具有挑战性。在本研究中，开发并验证了一种快速、具有成本效益的基于 PCR 的方法，用于检测柏树上的三齿小爪螨。使用简并和特异性引物扩增细胞色素 c 氧化酶亚基 I 基因，但只有后者能够区分三齿小爪螨和柏小爪螨。PCR 产物在两种物种的个体混合种群和单个个体的混合物中都能区分这两个物种，表明该检测方法具有敏感性。通过 XmnI 和 XbaI 内切酶进行 PCR-RFLP（限制性片段长度多态性）可将两种物种分开。此外，采用洗涤筛选方案使从树木样本中收集螨虫更快、更简单；该程序不干扰物种的分子检测。讨论了常规使用该检测方法监测侵染植物材料的检疫叶螨的可能性。
