Molecular and morphological investigations of Pezizalean mycorrhizae

A phylogeny of the Pezizales was reconstructed using 18S rDNA sequence data. The data set consisted of 117 taxa. Several approaches were used to generate the phylogenetic trees, including Neighbor Joining consensus (ML algorithm), Neighbor Joining with bootstrap analysis (ML algorithm) and Maximum likelihood using quartet puzzling. The clades resulting from this analysis roughly corresponded to both the traditionally recognized families within the order Pezizales and to the results reported from recent phylogenetic work on this group. The clades delineated by this study included the Helvellaceae, Pezizaceae, Morchellaceae, Discinaceae, Sarcosomataceae, Otideaceae and Sarcoscyphaceae. Ofthese, only the Sarcoscyphaceae had no suspected or proven associates. The Otideaceae and Tuberaceae contained the highest number of proven ectomycorrhizal and biotrophic fungi. The Morchellaceae and Helvellaceae had the second highest number of proven associates. Pezizalean fruitbodies and nearby root tips were collected for analysis with particular attention paid to members of the Otideaceae, Tuberaceae, Morchellaceae and Helvellaceae. The root tips were morphotyped and the the fruitbodies were identified. DNA was extracted from the fruitbodies and the root tips that were potentially ascomycetous. A portion of the nuclear ribosomal DNA was amplified spanning the two internally transcribed spacer regions (ITS 1 and ITS2) in addition to a portion of the 28S gene. This was then digested using the restriction enzymes Alu I, Hinf I and Rsa I. The resulting restriction patterns were analysed, and a match between fruitbody and root tip indicated an association (it being ectomycorrhizal depended upon the presence of a Hartig net in the root tip). No matches were found. Potentially ascomycetous root tip DNA and restriction patterns, in addition to herbarium samples of other Pezizalean fruitbodies were obtained from other sources in an attempt to find matches. Matches were found between Helvella leucomelaena and He/vella latispora fruitbody RFLP patterns and root tips morphotyped as MRA, E-strain and 'lightly colonized' under Abies lasiocarpa, Picea glauca X engelmanni and Pinus contorta var. latifolia. In addition, a match was found between a Trichophaea hemisphaerioides fruitbody RFLP pattern and an E-strain morphotyped tip under Abies lasiocarpa.