Detection of Pythium aphanidermatum in tomato using loop-mediated isothermal amplification (LAMP) with species-specific primers
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A loop-mediated isothermal amplification (LAMP) reaction with a primer set designed from the rDNA ITS sequence of P. aphanidermatum was developed. Results of a specificity test using 57 strains of Pythium spp. indicated that the LAMP assay gave no cross reactions in other 39 Pythium species, 11 strains of Phytophthora spp. and eight other soil borne pathogens. The detection limit was 10 fg of genomic DNA, which was ten times the sensitivity of the polymerase chain reaction. The LAMP assay was applied to hydroponic solution samples from tomato fields, and the results were compared to those of the conventional plating method. LAMP was observed to be effective for the specific detection of P. aphanidermatum. Furthermore, P. aphanidermatum was detected directly in tomato roots infected with P. aphanidermatum without DNA extraction. The LAMP method established in this study is a simple, sensitive and rapid tool for the detection of P. aphanidermatum.
KeywordsPythium aphanidermatum Detection Loop-mediated isothermal amplification (LAMP) Tomato
We are grateful to M. Suzuki, S. Uematsu, Y. Kajitani, Y. Chikuo and M. D. Coffey for providing Pythium spp. isolates.
This work was supported by Research and development projects for application in promoting new policy of agriculture, forestry and fisheries, Ministry of Agriculture, Forestry and Fisheries, Japan.
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