IngentaConnect Quantitative assessment of phytopathogenic fungi in various subst...

Authors: Lievens, Bart; Brouwer, Margreet¹; Vanachter, Alfons C. R. C.²; Lévesque, C. André³; Cammue, Bruno P. A.¹; Thomma, Bart P. H. J.⁴

Source: Environmental Microbiology, Volume 7, Number 11, November 2005 , pp. 1698-1710(13)

Publisher: Blackwell Publishing

Abstract:

Summary

Detection, identification and quantification of plant pathogens are the cornerstones of preventive plant disease management. To detect multiple pathogens in a single assay, DNA array technology currently is the most suitable technique. However, for sensitive detection, polymerase chain reaction (PCR) amplification before array hybridization is required. To evaluate whether DNA array technology can be used to simultaneously detect and quantify multiple pathogens, a DNA macroarray was designed and optimized for accurate quantification over at least three orders of magnitude of the economically important vascular wilt pathogens Verticillium albo-atrum and Verticillium dahliae. A strong correlation was observed between hybridization signals and pathogen concentrations for standard DNA added to DNA from different origins and for infested samples. While accounting for specific criteria like amount of immobilized detector oligonucleotide and controls for PCR kinetics, accurate quantification of pathogens was achieved in concentration ranges typically encountered in horticultural practice. Subsequently, quantitative assessment of other tomato pathogens (Fusarium oxysporum, Fusarium solani, Pythium ultimum and Rhizoctonia solani) in environmental samples was performed using DNA array technology and correlated to measurements obtained using real-time PCR. As both methods of quantification showed a very high degree of correlation, the reliability and robustness of the DNA array technology is shown.

Document Type: Research article

DOI: 10.1111/j.1462-2920.2005.00816.x

Affiliations: 1: Centre of Microbial and Plant Genetics (CMPG), Katholieke Universiteit Leuven, Kasteelpark Arenberg 20, B-3001 Heverlee-Leuven, Belgium. 2: Scientia Terrae Research Institute, Fortsesteenweg 30A, B-2860 Sint-Katelijne-Waver, Belgium. 3: Agriculture and Agri-Food Canada, Eastern Cereal and Oilseed Research Centre, 960 Carling Avenue, Ottawa, ON K1A 0C6, Canada. 4: Laboratory of Phytopathology, Wageningen University, Binnenhaven 5, 6709 PD Wageningen, the Netherlands.

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Quantitative assessment of phytopathogenic fungi in various substrates using a DNA macroarray