Biology of the ectomycorrhizal fungal genus, Rhizopogon

The morphology and anatomy of ectomycorrhizas of Rhizopogon arctostaphyli , R. ellenae , R. flavofibrillosus , R. occidentalis , R. rubescens , R. smithii , R. subcaerulescens and R. truncatus synthesized on Ponderosa pine ( Pinus ponderosa ) in glasshouse conditions using spore slurries, are described and compared. All species produced a well-developed Hartig net, and a well-developed fungal mantle. The mantles of R. arctostaphyli , R. smithii and R. subcaerulescens ectomycorrhizas were two-layered with outer mantle hyphae of wider diameter than inner mantle hyphae. The mantle of R. subcaerulescens ectomycorrhizas also had distinctive peg-like structures (cystidia) along peripheral hyphae. Rhizopogon truncatus ectomycorrhizas were tuberculate in morphology and had a rind-like mantle enclosing adjacent roots. In addition, several species exhibited crystal inclusions in the outer mantle, presumably at the interface between mantle and soil.     

Ultrastructure of developing basidiospores in <Emphasis Type="Italic">Rhizopogon roseolus</Emphasis> (= <Emphasis Type="Italic">R. rubescens</Emphasis>)

The ultrastructure of developing basidiospores in Rhizopogon roseolus is described. When viewed in the fruiting body chamber using scanning electron microscopy, basidiospores appear narrowly ellipsoid and have smooth walls. Eight basidiospores are usually produced on the apex of each sterigma on the basidium. Transmission electron micrographs showed that basidiospores formed by movement of cytoplasm (including the nuclei) via the sterigmata, and then each basidiospore eventually became separated from its sterigma by an electron-lucent septum. The sterigma and basidium subsequently collapsed, resulting in spore release. Freshly released spores retained the sterigmal appendage connected to the collapsed basidium. After spore release, the major ultrastructural changes in the spore concerned the lipid bodies and the spore wall. During maturation, lipid bodies formed and then expanded. Before release, the spore wall was homogeneous and electronlucent, but after release the spore wall comprised two distinct layers with electron-dense depositions at the inner wall, and the dense depositions formed an electron-dense third layer. The mature spore wall complex comprised at least four distinct layers: the outer electron-lucent thin double layers, the mottled electron-dense third layer, and the electron-lucent fourth layer in which electron-lucent granular substances were dispersed.     

Bipolar incompatibility system of an ectomycorrhizal basidiomycete, Rhizopogon rubescens

The mating systems of most ectomycorrhizal fungi have not been elucidated because of two reasons. One is the difficulty of obtaining homokaryotic isolates for mating tests caused by the low germination rate of basidiospores, and another is the difficulty of checking dikaryotization caused by the absence or inconsistent production of clamp connections on heterokaryotic mycelia under laboratory conditions. Basidiospore germination of a few ectomycorrhizal fungi has been induced by living roots of their host plants. Based on this information, we examined methods to obtain homokaryotic isolates of Rhizopogon rubescens using its host plant, Pinus thunbergii. The basidiospores of R. rubescens appeared to germinate well on an agar plate, on which axenic pine seedlings were grown in advance to induce germination, even when the seedlings were removed from the plate at the time of spore inoculation. To enhance the production rate of clamp connections on the heterokaryotic mycelia of R. rubescens, the culture medium composition was modified. The pH of the medium was critical for the production of clamp connections, and the optimal pH was higher for the production of clamp connections than for mycelial growth. These findings made it possible to conduct mating tests, and we found that the mating system of R. rubescens is bipolar with a multiallelic mating type factor.     

Priority effects determine the outcome of ectomycorrhizal competition between two Rhizopogon species colonizing Pinus muricata seedlings

Competition is often considered a fundamental process influencing assemblage structure, yet little is known about competition among ectomycorrhizal (EM) fungi. Here, we examine competitive interactions between Rhizopogon occidentalis and Rhizopogon salebrosus in a 6-month microcosm study. Pinus muricata seedlings were grown in three EM treatments: R. occidentalis, R. salebrosus, and R. occidentalis and R. salebrosus. At 2, 4, and 6 months, five seedlings per treatment were harvested and the EM root tip biomass of each species was determined. Root tips in the two-species treatment were identified using molecular techniques. R. occidentalis had similar EM root tip biomass when grown alone or in the presence of R. salebrosus. By contrast, R. salebrosus had significantly lower EM root tip biomass when grown with R. occidentalis than when grown alone, indicating it was a competitive inferior under the conditions tested. Competition was driven by differences in timing of colonization resulting in a strong priority effect for R. occidentalis. Our results, together with two earlier studies, indicate competition may play a more important role in EM interactions than previously recognized.     

On the origins of extreme mycorrhizal specificity in the Monotropoideae (Ericaceae): performance trade-offs during seed germination and seedling development

Fungal-induced seed germination is a phenomenon characteristic of mycorrhizal plants that produce dust-like seeds with only minimal nutritional reserves. In such systems, fungi trigger germination and/or subsidize development. We studied mycorrhizal germination in relation to mycorrhizal specificity in the Monotropoideae, a lineage of dust-seeded non-photosynthetic plants that are dependent upon ectomycorrhizal fungi of forest trees. A total of 1695 seed packets, each containing two to five compartments with seeds from different sources, were buried for up to 2 years near known ectomycorrhizal fungi in six different native forest locations. Upon harvest, seedlings were analysed by cultivation-independent molecular methods to identify their mycorrhizal fungi. We report that (i) germination is only induced by the same fungus that associates with mature plants or by closely related congeners; (ii) seedlings associated with the latter fungi develop less than those associated with maternal fungal species in most settings; and (iii) exceptions to this pattern occur in allopatric settings, where novel plant-fungal associations can result in the greatest seedling development. We interpret these results as evidence of performance trade-offs between breadth of host range and rate of development. We propose that in conjunction with host-derived germination cues, performance trade-offs can explain the extreme mycorrhizal specificity observed at maturity. The allopatric exceptions support the idea that performance trade-offs may be based on a coevolutionary arms race and that host range can be broadened most readily when naive fungal hosts are encountered in novel settings.     

High root concentration and uneven ectomycorrhizal diversity near Sarcodes sanguinea (Ericaceae): a cheater that stimulates its victims?

Sarcodes sanguinea is a nonphotosynthetic mycoheterotrophic plant that obtains all of its fixed carbon from neighboring trees through a shared ectomycorrhizal fungus. We studied the spatial structuring of this tripartite symbiosis in a forest where Sarcodes is abundant, and its only fungal and photosynthetic plant associates are Rhizopogon ellenae and Abies magnifica, respectively. We found disproportionately high concentrations of Abies roots adjacent to Sarcodes roots compared to the surrounding soil. Rhizopogon ellenae colonizes the vast majority of those Abies roots (86-98%), and its abundance tends to decrease with increasing distance from Sarcodes plants. At 500 cm from Sarcodes plants we did not detect R. ellenae, and the ectomycorrhizal community instead was dominated by members of the Russulaceae and Thelephoraceae, which are commonly dominant in other California pinaceous forests. The highly clumped distribution of Abies-R. ellenae ectomycorrhizas indicates that Sarcodes plants either establish within pre-existing clumps, or they stimulate clump formation. Several lines of evidence favor the latter interpretation, suggesting an unexpected mutualistic aspect to the symbiosis. However, the mechanism involved remains unknown.