A molecular diagnostic assay for the detection and identification of wood decay fungi of conifers

Ten taxon-specific primers were designed to amplify the Internal Transcribed Spacer of the rRNA operon of several important decay fungi of coniferous wood, including Armillaria spp., Echinodontium spp., Fomitopsis pinicola, Fuscoporia torulosa, Heterobasidion annosum sensu lato (s.l.), Onnia spp., Phaeolus schweinitzii, Phellinus weirii s.l., Pholiota spp. and Porodaedalea spp. Primers designed in this study and in a previous one for the identification of Laetiporus sulphureus and Stereum spp. were combined in two multiplex PCRs, which were tested for efficiency and specificity, and detected at least 1 pg of fungal target DNA. Target DNA at concentrations of 10-1 pg or lower can be detected with this assay applying SYBR® Green Real Time PCR. Validation assays performed on 129 naturally infected wood samples or fruiting bodies confirmed the reliability of the multiplex PCR-based diagnostic method. This method represents a simple and rapid diagnostic tool for the detection of a number of destructive wood decay fungi in conifer wood.