Inter-plant communication through mycorrhizal networks mediates complex adaptive behaviour in plant communities

Adaptive behaviour of plants, including rapid changes in physiology, gene regulation and defence response, can be altered when linked to neighbouring plants by a mycorrhizal network (MN). Mechanisms underlying the behavioural changes include mycorrhizal fungal colonization by the MN or interplant communication via transfer of nutrients, defence signals or allelochemicals. We focus this review on our new findings in ectomycorrhizal ecosystems, and also review recent advances in arbuscular mycorrhizal systems. We have found that the behavioural changes in ectomycorrhizal plants depend on environmental cues, the identity of the plant neighbour and the characteristics of the MN. The hierarchical integration of this phenomenon with other biological networks at broader scales in forest ecosystems, and the consequences we have observed when it is interrupted, indicate that underground ‘tree talk’ is a foundational process in the complex adaptive nature of forest ecosystems

Kin-selected signal transfer through mycorrhizal networks in Douglas-fir

Mycorrhizal networks create pathways for movement of resources and information molecules belowground. A mycorrhizal network is formed when two or more plants are linked by the same mycorrhizal fungus. Experiments have demonstrated movement of carbon and nitrogen between Douglas-fir and neighboring plants in response to source-sink dynamics, seasonality, and differences in age of linked plants. Furthermore, the network appears to act a conduit for information chemicals, where defense chemicals are transferred in response to herbivory or pathogen attack. Because of recent evidence implying the capacity for Douglas-fir to recognize kin, as well as differential colonization of Douglas-fir by ectomycorrhizas based on tree relatedness, this thesis aimed to determine whether Douglas-fir would preferentially transfer carbon and/or nitrogen through mycorrhizal networks to kin over strangers in response to herbivory treatment. Using seedlings with and without access to a mycorrhizal network (restricted or permitted via mesh pore size), stable isotope probing was used to track carbon and nitrogen in the system. One seedling of a pair was designated as the 'donor' and defoliated immediately prior to photosynthesizing with 99%-¹³C-CO₂ as well as pulse-labelling with 99%-¹⁵N ammonium nitrate. Both a greenhouse and field experiments were performed to corroborate results. Transfer was determined by measuring δ¹³C and δ¹³N in tissues (needle, stem, root) of kin and stranger seedlings. Data was analyzed using linear mixed effects models. Significantly more carbon was transferred to kin than strangers, and through the mycorrhizal network than when the mycorrhizal network was blocked. Furthermore, herbivory (in the form of western spruce budworm defoliation as well as manual defoliation) induced transfer of carbon to kin over strangers. Douglas-fir families differed in their tendency to transfer carbon and nitrogen to kin. Molecules potentially involved in defense signaling were identified using liquid chromatography coupled with mass spectroscopy. Ectomycorrhizal fungi that can form mycorrhizal networks were found on all seedlings. We conclude that preferential carbon transfer through mycorrhizal networks occurs between kin in Douglas-fir and is amplified by herbivory stress. Herbivory is not necessary for transfer, as some transfer also occurred in the no-herbivory treatment.Forestry, Faculty ofGraduat

Root Fragment Amendments Increase Nematode Density and Mycobiome Stochasticity in Douglas-Fir Seedlings

Relatively little is known about whole-plant fungal communities (mycobiome) and associated soil nematodes, especially with respect to woody plant seedlings and disturbance caused by forest harvesting. In a growth chamber experiment, we tested simulated clear-cut soil conditions on shoot biomass, total soil nematode density, and the shoot and root mycobiome of Douglas-fir, Pseudotsuga menziesii (Mirb.) Franco, seedlings. Soil treatments included unamended bare soil and soil amended with root segments of kinnikinnick, Arctostaphylos uva-ursi (L.) Spreng., pinegrass, Calamagrostis rubescens Buckley, or P. menziesii seedlings. We used next-generation Illumina sequencing and the PIPITS pipeline to obtain fungal taxa used for mycobiome community richness and Jaccard-based taxonomic normalized stochasticity ratio to assess mycobiome community assembly stochasticity. Total nematode density, measured from Baermann funnel extractions, increased in soils supplemented with A. uva-ursi or C. rubescens root segments. Root mycobiomes were more stochastic in the A. uva-ursi than P. menziesii or the bare conditions, whereas the shoot mycobiome was more stochastic in the C. rubescens treatment than in the P. menziesii treatment. Our results suggest that refugia plants impact the phyto-biome, in this case plant-associated nematodes and the stochasticity of root and shoot mycobiome community assembly, while not showing noticeable impacts on above-ground plant growth.Forestry, Faculty ofNon UBCForest and Conservation Sciences, Department ofReviewedFacultyResearche

Community structure of ericoid mycorrhizas and root-associated fungi of Vaccinium membranaceum across an elevation gradient in the Canadian Rocky Mountains

individuals were examined using culture-dependent sequencing and ARISA (automated ribosomal intergenic spacer analysis). Our results demonstrated that high elevation fungal communities, characterised by Rhizoscyphus ericae, differ from lower elevation communities, where Phialocephala fortinii was the most frequently isolated fungus. Co-occurrence analysis indicated that, overall, fungi tended to occur together more often than would be expected by chance. At the scale of the individual host plant, facilitation may play a more important role than competition in shaping fungal communities in these ecosystems.

Arbuscular mycorrhizal fungi in oat-pea intercropping

Abstract Arbuscular mycorrhizal fungal diversity can be altered by intercropping plant species, as well as N fertilizer applications. This study examined the effects of oat-pea intercropping and N fertilizer addition on the richness and diversity of mycorrhizal species, as well as identified the most common arbuscular mycorrhizal fungi (AMF) genera recruited for oats and peas in two growing seasons (2019 and 2020). The AMF diversity was higher in an intercropped system compared to their respective monocropping system. Under drier conditions in 2019, arbuscular mycorrhizal richness decreased with N fertilizer addition in sole peas and increased with N fertilizer addition in sole oats, but no significant change in richness was observed in oat-pea intercropping. During the wetter growing season 2020, arbuscular mycorrhizal diversity increased when oat and pea were intercropped, compared to either sole oat or sole pea. Diversispora in sole pea was a significant indicator differentiating the root associated AMF community from sole oat. Claroideoglomus richness increased in peas in 2020, thus this genus could be moisture dependent. Paraglomus richness in oat-pea intercropping was similar to sole oat in 2019, and similar to sole pea in 2020. This can suggest that Paraglomus is an indicator of plant stress under intercropping, as based on the premise that stressed plants release more exudates, and the subsequent mycorrhizal associations favor these plants with higher exudation. Future investigations can further reveal the functions and benefits of these mycorrhizal genera in annual monocrop and intercropping systems

Host and habitat filtering in seedling root-associated fungal communities: taxonomic and functional diversity are altered in ‘novel’ soils

Climatic and land use changes have significant consequences for the distribution of tree species, both through natural dispersal processes and following management prescriptions. Responses to these changes will be expressed most strongly in seedlings near current species range boundaries. In northern temperate forest ecosystems, where changes are already being observed, ectomycorrhizal fungi contribute significantly to successful tree establishment. We hypothesised that communities of fungal symbionts might therefore play a role in facilitating, or limiting, host seedling range expansion. To test this hypothesis, ectomycorrhizal communities of interior Douglas-fir and interior lodgepole pine seedlings were analysed in a common greenhouse environment following growth in five soils collected along an ecosystem gradient. Currently, Douglas-fir’s natural distribution encompasses three of the five soils, whereas lodgepole pine’s extends much further north. Host filtering was evident amongst the 29 fungal species encountered: 7 were shared, 9 exclusive to Douglas-fir and 13 exclusive to lodgepole pine. Seedlings of both host species formed symbioses with each soil fungal community, thus Douglas-fir did so even where those soils came from outside its current distribution. However, these latter communities displayed significant taxonomic and functional differences to those found within the host distribution, indicative of habitat filtering. In contrast, lodgepole pine fungal communities displayed high functional similarity across the soil gradient. Taxonomic and/or functional shifts in Douglas-fir fungal communities may prove ecologically significant during the predicted northward migration of this species; especially in combination with changes in climate and management operations, such as seed transfer across geographical regions for forestry purposes

Methods for Improving Human Gut Microbiome Data by Reducing Variability through Sample Processing and Storage of Stool.

Gut microbiome community analysis is used to understand many diseases like inflammatory bowel disease, obesity, and diabetes. Sampling methods are an important consideration for human microbiome research, yet are not emphasized in many studies. In this study, we demonstrate that the preparation, handling, and storage of human faeces are critical processes that alter the outcomes of downstream DNA-based bacterial community analyses via qPCR. We found that stool subsampling resulted in large variability of gut microbiome data due to different microenvironments harbouring various taxa within an individual stool. However, we reduced intra-sample variability by homogenizing the entire stool sample in liquid nitrogen and subsampling from the resulting crushed powder prior to DNA extraction. We experimentally determined that the bacterial taxa varied with room temperature storage beyond 15 minutes and beyond three days storage in a domestic frost-free freezer. While freeze thawing only had an effect on bacterial taxa abundance beyond four cycles, the use of samples stored in RNAlater should be avoided as overall DNA yields were reduced as well as the detection of bacterial taxa. Overall we provide solutions for processing and storing human stool samples that reduce variability of microbiome data. We recommend that stool is frozen within 15 minutes of being defecated, stored in a domestic frost-free freezer for less than three days, and homogenized prior to DNA extraction. Adoption of these simple protocols will have a significant and positive impact on future human microbiome research

Stool storage in a domestic frost-free freezer affects the abundance of bacterial taxa.

<p>A homogenized stool sample was stored in a domestic freezer for 0, 3,7,14, and 30 days, DNA was extracted and used for qPCR to compare bacterial taxa abundance. All bacterial taxa tested showed some change in abundance by day 30. *, p < 0.05.</p

RNAlater reduces DNA yields from stool samples.

<p>DNA extracted from samples not stored in RNAlater was significantly greater (p<0.0001) than samples stored in RNAlater.</p

Stool stored in nucleic acid stabilizer prior to processing did not protect against bacterial taxa changes.

<p>Stool was either stored with or without RNAlater (Qiagen) prior to freezing and then processing stool samples. Detection of Firmicutes, <i>Lactobacillus</i> spp. and <i>Bifidobacteria</i> spp. was reduced after storage in RNAlater. *, p = 0.05.</p