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Application of a real-time PCR method for the detection of pine wood nematode, Bursaphelenchus xylophilus, in wood samples from lodgepole pine

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For more content, see Nematologica.

A real-time PCR polymerase chain reaction (real-time PCR) method was developed to detect and differentiate Bursaphelenchus xylophilus (pine wood nematode, PWN) from other wood-inhabiting nematode species. A primer set and a specific Taqman® fluorescent probe were designed to amplify target B. xylophilus heat shock protein 70 sequences. After optimization, this real-time PCR assay was shown to be highly specific and sensitive, detecting at least 0.005 ng of B. xylophilus genomic DNA, as well as DNA extracted from single nematodes. The practical application of this real-time PCR diagnostic method for the detection of B. xylophilus from actual wood samples of lodgepole pine (Pinus contorta, Dougl. var. latifolia) trees containing a heterogeneous population of nematodes, rather than pure cultures or individual nematodes, is demonstrated. This method works well in the presence of potential inhibitors associated with wood after Baermann extraction and thus eliminates the need to produce pure nematode samples through further culturing and/or isolation of nematodes with a high-power microscope.

Affiliations: 1: Natural Resources Canada, Canadian Forest Service, Victoria, British Columbia, Canada, V8Z 1M5; 2: Centre for Plant Health, Canadian Food and Inspection Agency, Sidney, British Columbia, Canada, V8L 1H3


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