A multiplex PCR-based method for the detection and early identification of wood rotting fungi in standing trees

Abstract

Aims: The goal of this research was the development of a PCR‐based assay to identify important decay fungi from wood of hardwood tree species in northern temperate regions. Methods and Results: Eleven taxon‐specific primers were designed for PCR amplification of either nuclear or mitochondrial ribosomal DNA regions of Armillaria spp., Ganoderma spp., Hericium spp., Hypoxylon thouarsianum var. thouarsianum, Inonotus/Phellinus‐group, Laetiporus spp., Perenniporia fraxinea, Pleurotus spp., Schizophyllum spp., Stereum spp. and Trametes spp. Multiplex PCR reactions were developed and optimized to detect fungal DNA and identify each taxon with a sensitivity of at least 1 pg of target DNA in the template. This assay correctly identified the agents of decay in 82% of tested wood samples. Conclusions: The development and optimization of multiplex PCRs allowed for reliable identification of wood rotting fungi directly from wood. Significance and Impact of the Study: Early detection of wood decay fungi is crucial for assessment of tree stability in urban landscapes. Furthermore, this method may prove useful for prediction of the severity and the evolution of decay in standing trees.

Management of plant health risks associated with processing of plant-based wastes: A review

“…Molecular methods for identification of P. ramorum include conventional PCR (Kox et al, 2002;Wagner and Werres, 2003;Lane et al, 2003;Guglielmo et al, 2007) and Real-Time PCR Hayden et al, 2004;Belbahri et al, 2007;Bilodeau et al, 2007). PCR methods are generally more specific for P. ramorum.…”

European aspen (Populus tremula L.) genetic diversity assessed by molecular methods correlation with P. tremulae infection incidence

“…Such decay leads to structural deterioration of the woody tissues, which may lead to the loss of wood mechanical strength (Guglielmo et al, 2007). A common practice to ascertain stem decay affected trees is often restricted to visuall assessment, thus such inspection alows to identify only trees with an advanced stage of decay.…”

Molecular Tools for Detection of Plant Pathogenic Fungi and Fungicide Resistance

“…Although the efficiency of amplification is strongly influenced by amplicon size (shorter amplicons may be amplified preferentially over longer ones), an accurate and careful design of primers and the optimisation of their relative concentrations are required to overcome this drawback and get an equilibrate detection of all target fungi. Multiplex PCR technique has been used for the simultaneous detection and differentiation of Podosphaera xanthii and Golovinomyces cichoracearum in sunflower ; for detecting Phytophthora lateralis in cedar trees and water samples, including detection of an internal control in the same reaction (Winton & Hansen, 2001); for determining the mating type of the pathogens Tapesia yallundae and T. acuformis (Dyer et al, 2001); for differentiating two pathotypes of Verticilliun albo-atrum infecting hop (Radiek et al, 2004) and for distinguishing among eleven taxons of wood decay fungi infecting hardwood trees (Guglielmo et al, 2007). Due to the complexity of the design this technique has recently been replaced by other multiplexing techniques including multiplex real-time PCR (see below).…”