Atmospheric pollution, soil nutrients and climate effects on Mucoromycota arbuscular mycorrhizal fungi.

Fine root endophyte mycorrhizal fungi in the Endogonales (Mucoromycota arbuscular mycorrhizal fungi, M-AMF) are now recognized as at least as important globally as Glomeromycota AMF (G-AMF), yet little is known about the environmental factors which influence M-AMF diversity and colonization, partly because they typically only co-colonize plants with G-AMF. Wild populations of Lycopodiella inundata predominantly form mycorrhizas with M-AMF and therefore allow focussed study of M-AMF environmental drivers. Using microscopic examination and DNA sequencing we measured M-AMF colonization and diversity over three consecutive seasons and modelled interactions between these response variables and environmental data. Significant relationships were found between M-AMF colonization and soil S, P, C:N ratio, electrical conductivity, and the previously overlooked micronutrient Mn. Estimated N deposition was negatively related to M-AMF colonization. Thirty-nine Endogonales Operational Taxonomic Units (OTUs) were identified in L. inundata roots, a greater diversity than previously recognized in this plant. Endogonales OTU richness correlated negatively with soil C:N while community composition was mostly influenced by soil P. This study provides first evidence that M-AMF have distinct ecological preferences in response to edaphic variables also related to air pollution. Future studies require site-level atmospheric pollution monitoring to guide critical load policy for mycorrhizal fungi in heathlands and grasslands.

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.

Fine-level mycorrhizal specificity in the Monotropoideae (Ericaceae): specificity for fungal species groups.

The Monotropoideae (Ericaceae) are non-photosynthetic angiosperms that obtain fixed carbon from basidiomycete ectomycorrhizal fungi. In previous work, we showed that each plant species is associated with a single genus or a set of closely related genera of ectomycorrhizal fungi. Here we show that the level of specificity is much higher. We used a molecular phylogenetic approach to contrast specificity patterns among eight plant lineages and three fungal genera. We relied on fungal nuclear internal transcribed spacer (nrITS) sequence data obtained from 161 basidiocarps and 85 monotropoid roots representing 286 sampled plants screened using restriction length polymorphisms. From the phylogenetic placement of fungal symbionts in fungal phylograms, we found that three basal (Sarcodes, Pterospora, Pleuricospora) and one derived lineage (Allotropa) of plants target narrow clades of closely related species groups of fungi, and four derived lineages (Monotropa hypopithys species group, Pityopus) target more distant species groups. Within most plant lineages, geography and photobiont association constrain specificity. Specificity extended further in Pterospora andromedea, in which sequence haplotypes at the plastid trn L-F region of 73 plants were significantly associated with different fungal species groups even in sympatry. These results indicate that both the macro- and microevolution of the Monotropoideae are tightly coupled to their mycorrhizal symbionts.

Extreme specificity in epiparasitic Monotropoideae (Ericaceae): widespread phylogenetic and geographical structure.

The Monotropoideae (Ericaceae) are nonphotosynthetic plants that obtain fixed carbon from their fungal mycorrhizal associates. To infer the evolutionary history of this symbiosis we identified both the plant and fungal lineages involved using a molecular phylogenetic approach to screen 331 plants, representing 10 of the 12 described species. For five species no prior molecular data were available; for three species we confirmed prior studies which used limited samples; for five species all previous reports are in conflict with our results, which are supported by sequence analysis of multiple samples and are consistent with the phylogenetic patterns of host plants. The phylogenetic patterns observed indicate that: (i) each of the 13 plant phylogenetic lineages identified is specialized to a different genus or species group within five families of ectomycorrhizal Basidiomycetes; (ii) mycorrhizal specificity is correlated with phylogeny; (iii) in sympatry, there is no overlap in mature plant fungal symbionts even if the fungi and the plants are closely related; and (iv) there are geographical patterns to specificity.

Regional specialization of Sarcodes sanguinea (Ericaceae) on a single fungal symbiont from the Rhizopogon ellenae (Rhizopogonaceae) species complex.

We have sampled the mycorrhizal roots of 76 snow plants (Sarcodes sanguinea, Monotropoideae, Ericaceae) in two areas of the Sierra Nevada of California that are ∼180 km apart. To identify the fungal symbionts associated with these plants, we first analyzed restriction fragment length polymorphisms (RFLPs) of the internal transcribed spacer region (ITS) of the fungal nuclear ribosomal repeat. Fungal ITS-RFLPs were successfully produced from 57 of the 76 plants sampled, and all symbionts shared the same DNA fragment pattern. The morphology of S. sanguinea mycorrhizae was consistent with that expected from a Rhizopogon species in section Amylopogon. To confirm and refine this identification, a total of six fungal ITS sequences were determined from S. sanguinea mycorrhizae. These sequences were analyzed together with eight existing and eight newly determined ITS sequences from Rhizopogon section Amylopogon. The newly determined sequences include an ITS sequence from the fungal symbiont of pine drops (Pterospora andromedea, Monotropoideae, Ericaceae), a plant that was previously reported to be exclusively associated with the Rhizopogon subcaerulescens group. When these sequences were analyzed together, the Sarcodes symbionts grouped tightly with several collections of R. ellenae including the holotype, one collection of R. idahoensis, and one collection of R. semireticulatus. A different lineage comprised collections of R. subgelatinosus, R. subcaerulescens, another collection of R. semireticulatus, and the Pterospora symbiont. We conclude that S. sanguinea associates exclusively with a single species in the R. ellenae species complex throughout our sampling range. These results indicate a much higher level of specificity in S. sanguinea than was previously reported and confirm the emerging pattern that nonphotosynthetic, monotropoid plants generally associate very specifically with a narrow range of ectomycorrhizal fungi.

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.