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A qPCR-based method for the detection and quantification of the peach powdery mildew ( Podosphaera pannosa ) in epidemiological studies
European Journal of Plant Pathology ( IF 1.8 ) Pub Date : 2020-10-17 , DOI: 10.1007/s10658-020-02136-0
Neus Marimon , Iban Eduardo , Maela León , Mónica Berbegal , Josep Armengol , Jordi Luque

A qPCR-based method was developed to detect and quantify Podosphaera pannosa, the main causal agent of peach powdery mildew. A primer pair was designed to target part of the ITS region of the fungal ribosomal DNA, which proved to be highly specific and sensitive. A minimum of 2.81 pg µL− 1 of P. pannosa DNA and 6 conidia mL− 1 in artificially-prepared conidia suspensions were found to be the limit of detection. Moreover, a quantification of conidia placed on plastic tapes commonly used in volumetric air samplers was performed. Regression equations on conidia quantification obtained either from aqueous conidia suspensions or conidia placed on plastic tapes were similar. The protocol was further validated in field conditions by estimating the number of P. pannosa conidia obtained with an air sampler, by both microscopic and molecular quantification. Both techniques detected the peaks of conidia production during a 4-month sampling period, and a significant correlation (r = 0.772) was observed between both quantification methods. Additionally, the molecular method was applied to detect latent fungal inoculum in different plant parts of peach trees. The pathogen was detected mainly on the bark of affected twigs, and to a lesser extent, in foliar buds. The method developed here can be applied in the study of P. pannosa epidemiology and can help in improving the management of this pathogen through its early detection and quantification.



中文翻译:

流行病学研究中基于qPCR的桃白粉病(Podosphaera pannosa)检测和定量方法

开发了一种基于qPCR的方法来检测和定量桃白粉病的主要病原体Podosphaera pannosa。设计引物对以靶向真菌核糖体DNA ITS区的一部分为目标,事实证明该酶具有高度特异性和敏感性。至少2.81 pg µL − 1P. pannosa DNA和6个分生孢子mL − 1在人工制备的分生孢子中发现悬浮液是检测的极限。此外,对放置在体积空气采样器中常用的塑料胶带上的分生孢子进行了定量分析。从含水分生孢子悬浮液或置于塑料带上的分生孢子获得的分生孢子定量回归方程相似。通过在现场条件下通过显微镜和分子定量估计用空气采样器获得的P. pannosa分生孢子的数量,进一步验证了该方案。两种技术都在4个月的采样期内检测到了分生孢子的峰值,并且存在显着的相关性(r 两种定量方法之间观察到= 0.772)。另外,该分子方法被用于检测桃树不同植物部位的潜在真菌接种物。病原体主要在受影响的树枝的树皮上检出,而在叶芽中检出程度较小。此处开发的方法可用于研究P. pannosa流行病学,并可以通过早期发现和量化来帮助改善这种病原体的管理。

更新日期:2020-11-12
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