The north American Pine mushroom tricholoma magnivelare (peck) redhead: in vitro mycelial culture, ectomycorrhizal synthesis trials and preliminary shiro analysis
Creators: name: Fogarty, Fidel
Publisher: name: University of British Columbia
Year: 2009

description: Tissue cultures of Tricholoma magnivelare (Peck) Redhead were isolated from basidiocarps and maintained on MMN agar media. The mycelial mass of resulting colonies was expanded by transfers to fresh media. Isolates were confirmed as T. magnivelare by employing polymerase chain reaction - restriction fragment length polymorphism (PCR-RFLP) analysis to compare DNA from the vegetative mycelium to DNA from sporocarp tissue. Isolates of T. magnivelare were tested in vitro for cellulose, lignin and pectin decomposing abilities. As with many known ectomycorrhizal fungi, none of the isolates were able to degrade any of these substrates, indicating only limited, if any, cellulase, lignase and pectinase production. In vitro ectomycorrhizal synthesis trials were performed by combining T. magnivelare isolates with lodgepole pine, Douglas fir and western hemlock seedlings. Mycelia from five isolates were inoculated onto root systems of 8-month-old seedlings grown aseptically in Erlenmeyer flasks. No ectomycorrhizae were formed. However, some of the lodgepole pine root systems developed slightly swollen, darkened root-tips, similar in appearance to the initial stages of rootlet colonization by a related ectomycorrhizal species, Tricholoma matsutake (Ito et Imai) Singer. The affected rootlets resembled what Japanese researchers described as a 'transient mycorrhization', and appeared similar in some respects to rootlets in the initial stages of a host-parasite relationship. Longer term studies are required to determine i f ectomycorrhizae can be synthesized from these isolates under laboratory conditions. Preliminary field investigations of basidiocarp production and the shiro (underground colony) of T. magnivelare were undertaken on three sampling plots located in a Pine Mushroom Study Area established in the Nass Valley, near Terrace, British Columbia. The numbers and grades of mushrooms in plots were determined and mapped during three sampling periods in autumn, 1994. Clusters of mushrooms were inferred from sporocarp maps however, several more years of intensive monitoring are required to assess spatial and temporal distributions. Soil samples were collected both directly underneath and away from T. magnivelare basidiocarps in order to examine the types and abundance of root-tip morphotypes. Initial macro and micromorphological descriptions were done for the seven root-tip morphotypes found. Preliminary results indicated that a charcoal-grey to black, slightly swollen, elongated and coarsely fissured morphotype was more common directly beneath T. magnivelare basidiocarps than further away from them. This morphotype probably represents one of the latter stages of T. magnivelare rootlet colonization. Another of the morphotypes (white mycelium covering light cinnamon-brown rootlet) may represent an earlier stage of T. magnivelare colonization. Molecular analysis of fungal DNA from excised root-tips is required to confirm the identity of these root-tip morphotypes. I f these were in fact T. magnivelare colonized rootlets then the presence of a mantle and Hartig net supports the theory that the T. magnivelare is ectomycorrhizal in nature. However, rootlet characteristics, similar in some respects to those of a host-parasite relationship (i.e. darkening, necrosis and eventual sloughing of cortical cells) were observed. As suspected, T. magnivelare has some trophic and morphological characteristics typical of an ectomycorrhizal fungus