In vitro culture and growth modeling of Tuber spp. and inoculation of hardwoods with T. melanosporum ascospores

Graduation date: 1983In vitro growth of twelve isolates of Tither spp., the true truffles, was quantified by an agar-melt procedure. All isolates grew\ud poorly on media commonly used for the culture of mycorrhizal fungi, but responded markedly to the addition of nitrate, as well as other\ud inorganic ions, to malt and potato-dextrose basal media. The pH of all media containing nitrate rose during incubation, and maximum rates of growth occurred at these higher pH values. Growth response to eight polycarboxylic acid buffers was investigated; one in particular, B-ntethylcarballylic acid is both nontoxic and has a buffering range\ud from pH 2.5 - 8.2 and is suggested for use in subsequent in vitro Tuber studies. Phosphate buffer was also found to be non-inhibitory at moderate concentrations. Of the six isolates studied for temperature response, all had optima around 20 C and declined around 26.5 C. The possible response of other hard-to-culture ectomycorrhizal fungi to nitrate is discussed. A few simple models of fungal growth in shallow Petri dishes were\ud proposed, and the growth data obtained as mycelial dry weight for twelve Tuber isolates were tested for fit. Growth was hypothesied to be uniform and proportional to total biomass, or to occur only at the colony periphery and proportional to the square root of the biomass. Each of these models was altered to include a time-dependent variation: growth was hypothesized to also depend upon the concentration of a metabolite produced by the fungus. Metabolite-dependent\ud models gave better fit than metabolite-independent versions, but differences between peripheral and uniform growth models were less clear. In inoculation studies of oaks and filberts with T.\ud melanosporum ascospores, viability of spores was maintained either by freezing or 5 C storage of ascocarps. Liming improves mycorrhization, although not necessarily seedling vigor. The effect of soil organic\ud matter is minimal on growth and mycorrhizal development of Corylus avellana. Soil inoculation with spores is an effective method of establishing the symbiosis, but a germling drench is less so. A method which nondestructively measures seedling root volume was proposed in order to study mycorrhizal colonization on the same seedling over time