Phylogenetics, population genetics, molecular epidemiology, and pathogenicity of the Douglas-fir Swiss needle cast pathogen Phaeocryptopus gaeumannii Public Deposited

http://ir.library.oregonstate.edu/concern/graduate_thesis_or_dissertations/fb494b597

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  • A hierarchical series of studies, based mainly on molecular data, was conducted to elucidate the life history of the Douglas-fir Swiss needle cast pathogen Phaeocryptopus gaeumannii at macro- and micro-evolutionary scales. This information was then utilized to design and evaluate molecular diagnostic tools for use in studies on the epidemiology of a Swiss needle cast outbreak near Tillamook, Oregon. Phylogenetic analyses of partial nuclear ribosomal gene sequences indicated that P. gaeumannii, currently classified in the Venturiaceae, is closely related to neither Phaeocryptopus nudus, type of the genus, nor Venturia inaequalis, type of the Venturiaceae. Instead, it is closely related to members of the "sooty molds" (Capnodiales), particularly the common and morphologically similar Douglas-fir epiphyte Rasutoria pseudotsugae (Euantennariaceae). Single-strand conformation polymorphisms, revealing DNA sequence variation in five loci, were used to investigate population biology of P. gaeumannii from a worldwide collection of isolates. In western Oregon, P. gaeumannii population structure suggests a predominantly selfing reproductive mode within two reproductively isolated sympatric lineages. One lineage was widely distributed both locally and abroad. The second lineage was restricted to western Oregon and suggested a correlation with symptoms of Swiss needle cast. A novel application of real-time PCR allowed species-specific detection and quantification of P. gaeumannii and proved a good measure of its biomass in Douglas-fir needles. Compared to other techniques (ergosterol and a DNA probe), real-time PCR correlated best with visual estimates of needle colonization and additionally proved useful early in the first year of the colonization process before visible development of fruiting structures. While all four methods provided evidence that sites expressing a range of disease severity differed in the degree of fungal colonization, only real-time PCR consistently separated both moderately and severely diseased sites from relatively healthy sites. Seedling inoculation experiments, fulfilling Koch's postulates, demonstrated that P. gaeumannii is the causal agent of Swiss needle cast, as observed in the Tillamook epidemic. Furthermore, the incorporation of virulence tests provided independent, non-molecular evidence that Oregon's pathogen population is not homogeneous. One strain, isolated from a severely diseased site, caused significantly greater symptom severity than strains derived from less damaged sites.
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