QuantifyingVerticillium dahliae in soils collected from potato fields using a competitive PCR assay

Abstract

A quantitative PCR assay based on the competitive PCR technique was compared to the classical soil dilution (SD) method for its ability to estimateV. dahliae propagules directly in soils collected from fields under potato production. A strong correlation (r = 0.97) was observed betweenV. dahliae propagules estimated using the quantitative PCR assay and those using the SD method. Coamplification ofV. dahliae DNA with competitor DNA provided accurate quantification in the range of 102 to 107 spores and 1 to 100 microsclerotia/g of soil. The number ofV. dahliae propagules detected in PEI soils ranged from 4.9 to 15.6 and 0.06 to 0.5 microsclerotia/g of soil for PCR assay and SD method, respectively. The strong correlation between PCR assay and SD method and the non significant differences between replications of PCR estimates ofV. dahliae propagules in soils (P< 0.05) show that the PCR assay is reliable and reproducible, and comparable to the SD method. This method is fast, does not depend on the subjectiveness of the traditional plating method, and offers an improvement in speed and precision over currently used methods. In addition, it can be extended to estimateV. dahliae propagules in other pathosystems and finds immediate and practical use in epidemiological studies to determine the effects of various crop management strategies on the dynamics and level of fungal propagules in the soil in order to establish threshold levels for assessing disease risks and develop disease prediction systems.