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migratory endoparasitic nematode Pratylenchus thornei is an important pathogen of wheat in Europe, Africa, North America, Asia, the Middle East and Australia. In the Mediterranean basin, it causes also severe yield decline of crops such as chickpea, faba bean (Greco et al., 1984; Glazer and Orion, 1983) and pulse crops (Di Vito et al., 1992). Correct identification and quantification of this nematode are necessary for providing advice to farmers, but are not easily obtained with the traditional way of microscopic observation. We developed a qPCR assay to detect and quantify P. thornei in a short but accurate manner. A qPCR primer set, including two primers and a TaqMan probe, was designed based on the sequence of the β‐1,4‐endoglucanase gene. The assay was optimized by using the primers in a qPCR assay with SYBR green I dye and setting the qPCR program to different annealing temperatures ranging from 62 to 69 °C. Based on the Ct‐values, we retained the program with an annealing temperature of 69 °C. The assay with the probe was very sensitive as it was able to detect a single individual of P. thornei, even when mixed with up to 80 individuals of P.penetrans. The specificity of the qPCR assay including the probe was confirmed by the lack of amplification of DNA from 47 populations of 15 other Pratylenchus species and 9 different isolates from P. thornei. A dilution series from DNA of P. thornei resulted in a standard curve showing a highly significant linearity between the Ct‐values and the dilution rates (R² = 0.98; slope = ‐3.38; E= 97.6 %). The qPCR assay developed in this study proved to be specific and sensitive, thus providing a fast and accurate tool for detection and quantification of this pathogen during research, as well as for
diagnostic labs
Originele taal-2Engels
TitelABSTRACTS 66th International Symposium on Crop Protection
Publicatiedatum20-mei-2014
StatusGepubliceerd - 20-mei-2014

ID: 2806795