Host genotype controls ecological change in the leaf fungal microbiome.

Leaf fungal microbiomes can be fundamental drivers of host plant success, as they contain pathogens that devastate crop plants and taxa that enhance nutrient uptake, discourage herbivory, and antagonize pathogens. We measured leaf fungal diversity with amplicon sequencing across an entire growing season in a diversity panel of switchgrass (Panicum virgatum). We also sampled a replicated subset of genotypes across 3 additional sites to compare the importance of time, space, ecology, and genetics. We found a strong successional pattern in the microbiome shaped both by host genetics and environmental factors. Further, we used genome-wide association (GWA) mapping and RNA sequencing to show that 3 cysteine-rich receptor-like kinases (crRLKs) were linked to a genetic locus associated with microbiome structure. We confirmed GWAS results in an independent set of genotypes for both the internal transcribed spacer (ITS) and large subunit (LSU) ribosomal DNA markers. Fungal pathogens were central to microbial covariance networks, and genotypes susceptible to pathogens differed in their expression of the 3 crRLKs, suggesting that host immune genes are a principal means of controlling the entire leaf microbiome.

Stochastic and deterministic processes shape bioenergy crop microbiomes along a vertical soil niche.

Sustainable biofuel cropping systems aim to address climate change while meeting energy needs. Understanding how soil and plant-associated microbes respond to these different cropping systems is key to promoting agriculture sustainability and evaluating changes in ecosystem functions. Here, we leverage a long-term biofuel cropping system field experiment to dissect soil and root microbiome changes across a soil-depth gradient in poplar, restored prairie and switchgrass to understand their effects on the microbial communities. High throughput amplicon sequencing of the fungal internal transcribed spacer (ITS) and prokaryotic 16S DNA regions showed a common trend of root and soil microbial community richness decreasing and evenness increasing with depth. Ecological niche (root vs. soil) had the strongest effect on community structure, followed by depth, then crop. Stochastic processes dominated the structuring of fungal communities in deeper soil layers while operational taxonomic units (OTUs) in surface soil layers were more likely to co-occur and to be enriched by plant hosts. Prokaryotic communities were dispersal limited at deeper depths. Microbial networks showed a higher density, connectedness, average degree and module size in deeper soils. We observed a decrease in fungal-fungal links and an increase of bacteria-bacteria links with increasing depth in all crops, particularly in the root microbiome.

CONSTAX2: improved taxonomic classification of environmental DNA markers.

SUMMARY: CONSTAX-the CONSensus TAXonomy classifier-was developed for accurate and reproducible taxonomic annotation of fungal rDNA amplicon sequences and is based upon a consensus approach of RDP, SINTAX and UTAX algorithms. CONSTAX2 extends these features to classify prokaryotes as well as eukaryotes and incorporates BLAST-based classifiers to reduce classification errors. Additionally, CONSTAX2 implements a conda-installable command-line tool with improved classification metrics, faster training, multithreading support, capacity to incorporate external taxonomic databases and new isolate matching and high-level taxonomy tools, replete with documentation and example tutorials. AVAILABILITY AND IMPLEMENTATION: CONSTAX2 is available at https://github.com/liberjul/CONSTAXv2, and is packaged for Linux and MacOS from Bioconda with use under the MIT License. A tutorial and documentation are available at https://constax.readthedocs.io/en/latest/. Data and scripts associated with the manuscript are available at https://github.com/liberjul/CONSTAXv2_ms_code. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Fungal, Bacterial, and Archaeal Diversity in Soils Beneath Native and Introduced Plants in Fiji, South Pacific.

The Fiji Islands is an archipelago of more than 330 islands located in the tropics of the South Pacific Ocean. Microbial diversity and biogeography in this region is still not understood. Here, we present the first molecular characterization of fungal, bacterial, and archaeal communities in soils from different habitats within the largest Fijian island, Viti Levu. Soil samples were collected from under native vegetation in maritime-, forest-, stream-, grassland-, and casuarina-dominated habitats, as well as from under the introduced agricultural crops sugarcane, cassava, pine, and mahogany. Soil microbial diversity was analyzed through MiSeq amplicon sequencing of 16S (for prokaryotes), ITS, LSU ribosomal DNA (for fungi). Prokaryotic communities were dominated by Proteobacteria (~ 25%), Acidobacteria (~ 19%), and Actinobacteria (~ 17%), and there were no indicator species associated with particular habitats. ITS and LSU were congruent in ß-diversity patterns of fungi, and fungal communities were dominated by Ascomycota (~ 57-64%), followed by Basidiomycota (~ 20-23%) and Mucoromycota (~ 10%) according to ITS, or Chytridiomycota (~ 9%) according to LSU. Indicator species analysis of fungi found statistical associations of Cenococcum, Wilcoxina, and Rhizopogon to Pinus caribaea. We hypothesize these obligate biotrophic fungi were co-introduced with their host plant. Entoloma was statistically associated with grassland soils, and Fusarium and Lecythophora with soils under cassava. Observed richness varied from 65 (casuarina) to 404 OTUs (cassava) for fungi according to ITS region, and from 1268 (pine) to 2931 OTUs (cassava) for bacteria and archaea. A major finding of this research is that nearly 25% of the fungal OTUs are poorly classified, indicative of novel biodiversity in this region. This preliminary survey provides important baseline data on fungal, bacterial, and archaeal diversity and biogeography in the Fiji Islands.

CONSTAX: a tool for improved taxonomic resolution of environmental fungal ITS sequences.

BACKGROUND: One of the most crucial steps in high-throughput sequence-based microbiome studies is the taxonomic assignment of sequences belonging to operational taxonomic units (OTUs). Without taxonomic classification, functional and biological information of microbial communities cannot be inferred or interpreted. The internal transcribed spacer (ITS) region of the ribosomal DNA is the conventional marker region for fungal community studies. While bioinformatics pipelines that cluster reads into OTUs have received much attention in the literature, less attention has been given to the taxonomic classification of these sequences, upon which biological inference is dependent. RESULTS: Here we compare how three common fungal OTU taxonomic assignment tools (RDP Classifier, UTAX, and SINTAX) handle ITS fungal sequence data. The classification power, defined as the proportion of assigned OTUs at a given taxonomic rank, varied among the classifiers. Classifiers were generally consistent (assignment of the same taxonomy to a given OTU) across datasets and ranks; a small number of OTUs were assigned unique classifications across programs. We developed CONSTAX (CONSensus TAXonomy), a Python tool that compares taxonomic classifications of the three programs and merges them into an improved consensus taxonomy. This tool also produces summary classification outputs that are useful for downstream analyses. CONCLUSIONS: Our results demonstrate that independent taxonomy assignment tools classify unique members of the fungal community, and greater classification power is realized by generating consensus taxonomy of available classifiers with CONSTAX.

Bifiguratus adelaidae, gen. et sp. nov., a new member of Mucoromycotina in endophytic and soil-dwelling habitats.

Illumina amplicon sequencing of soil in a temperate pine forest in the southeastern United States detected an abundant, nitrogen (N)-responsive fungal genotype of unknown phylogenetic affiliation. Two isolates with ribosomal sequences consistent with that genotype were subsequently obtained. Examination of records in GenBank revealed that a genetically similar fungus had been isolated previously as an endophyte of moss in a pine forest in the southwestern United States. The three isolates were characterized using morphological, genomic, and multilocus molecular data (18S, internal transcribed spacer [ITS], and 28S rRNA sequences). Phylogenetic and maximum likelihood phylogenomic reconstructions revealed that the taxon represents a novel lineage in Mucoromycotina, only preceded by Calcarisporiella, the earliest diverging lineage in the subphylum. Sequences for the novel taxon are frequently detected in environmental sequencing studies, and it is currently part of UNITE's dynamic list of most wanted fungi. The fungus is dimorphic, grows best at room temperature, and is associated with a wide variety of bacteria. Here, a new monotypic genus, Bifiguratus, is proposed, typified by Bifiguratus adelaidae.

Mycorrhization of pecan (Carya illinoinensis) with black truffles: Tuber melanosporum and Tuber brumale.

Pecan, Carya illinoinensis, is an economically important nut producing tree that can establish ectomycorrhizal symbiosis with a high diversity of fungi. In the southern USA, truffles (Tuber spp.) sometimes fruit prolifically in cultivated pecan orchards and regularly associate with pecan roots as ectomycorrhizae (ECMs). It has been demonstrated that some valuable European truffle species (e.g., Tuber borchii and Tuber aestivum) can form ECMs with pecan seedlings in nursery conditions. Thus, pecan may represent an attractive alternative host to forest trees for truffle growers given the potential for co-cropping truffles and pecans. To further explore the capacity of pecan to host truffle symbionts, pecan seedlings were inoculated with species of black truffles that are economically important in Europe, T. melanosporum and T. brumale. Ectomycorrhizae were characterized molecularly and their morphology was described in detail. Mycorrhization rates on pecan roots were assessed over a 2-year period. Tuber melanosporum and T. brumale produced well-formed ECMs with a level of root colonization in the first year of 37.3 and 34.5%, respectively. After 24 months, the level of mycorrhization increased for T. brumale (49.4%) and decreased for T. melanosporum (10.5%) inversely to that of non-target ECM greenhouse contaminants (e.g., Sphaerosporella brunnea, Trichophaea woolhopeia, Pulvinula constellatio). To assess whether mating types segregated in T. melanosporum as been reported for other host species, we amplified the mating-type locus from single T. melanosporum ECM belonging to different seedlings over a 2-year period. The two mating idiomorphs were nearly equally represented along the 2-year time span: MAT 1-1-1 decreased from 59.4% in the first year to 48.5% in the second year after inoculation. Data reported in this study add to knowledge on the mycorrhization of pecan trees with commercial truffles and has application to truffle and nut co-cropping systems.

The Truffle Microbiome: Species and Geography Effects on Bacteria Associated with Fruiting Bodies of Hypogeous Pezizales.

Fungi that produce their fruiting bodies underground within the soil profile are known commonly as truffles. Truffle fruiting bodies harbor a diverse but poorly understood microbial community of bacteria, yeasts, and filamentous fungi. In this study, we used next-generation 454 amplicon pyrosequencing of the V1 and V4 region of the bacterial 16S ribosomal DNA (rDNA) in order to characterize and compare effects of truffle species and geographic origin on the truffle microbiome. We compared truffle microbiomes of the glebal tissue for eight truffle species belonging to four distinct genera within the Pezizales: Tuber, Terfezia, Leucangium, and Kalapuya. The bacterial community within truffles was dominated by Proteobacteria, Bacterioides, Actinobacteria, and Firmicutes. Bacterial richness within truffles was quite low overall, with between 2-23 operational taxonomic units (OTUs). Notably, we found a single Bradyrhizobium OTU to be dominant within truffle species belonging to the genus Tuber, irrespective of geographic origin, but not in other truffle genera sampled. This study offers relevant insights into the truffle microbiome and raises questions concerning the recruitment and function of these fungal-associated bacteria consortia.

Characterizing root-associated fungal communities and soils of Douglas-fir (Pseudotsuga menziesii) stands that naturally produce Oregon white truffles (Tuber oregonense and Tuber gibbosum).

Many truffle species in the genus Tuber are endemic to North America. Some of these have commercial value such as Tuber oregonense and Tuber gibbosum, commonly known as Oregon white truffles. Most of what is known about the ecology of these truffles comes from observational data. These truffle species form ectomycorrhizas with Douglas-fir (Pseudotsuga menziesii) and sometimes fruit abundantly in early successional forest regrowth. The goal of this study was to characterize fungal communities and soils associated with truffle-producing Douglas-fir sites. We extracted DNA from roots of five trees at four different truffle-producing Douglas-fir sites (n = 20). We amplified the internal transcribed spacer (ITS) region of the nuclear ribosomal DNA (nrDNA) and sequenced amplicons with 454 pyrosequencing. After quality filtering, we assembled 15,713 sequences into 150 fungal operational taxonomic units (OTUs). Pezizomycetes (Tuber and Pyronemataceae) were the most abundant taxa detected followed by Helotiales. Agaricomycetes represented most by Thelephoraceae, Russulaceae, and Inocybaceae were also abundant. A total of five Tuber species were detected. T. oregonense was the most abundant OTU, followed by T. gibbosum and Wilcoxina mikolae. Fungal root endophytes were also detected and well represented by Chalara and Phialocephala spp. Fungal community structure and soil chemistry differed between sites. This study represents the first characterization of the fungal communities in Douglas-fir stands producing Oregon white truffles. We found that Tuber species can be dominant ectomycorrhizal symbionts of Douglas-fir. Truffle fungi are also important in forest health, food webs, and as a non-timber forest resource that can contribute to rural economies.

Assessment of ectomycorrhizal biodiversity in Tuber macrosporum productive sites.

Tuber macrosporum Vittad. is a truffle with superb organoleptic properties, whose cultivation is still in its infancy. For the first time we have aimed to provide information on ectomycorrhizal communities in natural and cultivated T. macrosporum sites. Ectomycorrhizal morphotypes were identified using ITS nrDNA sequencing and sorted into molecular operational taxonomic unit (MOTU). We detected 16 MOTUs in the T. macrosporum cultivated plantation. Ascomycota were the most abundant (86.4%) with Helvellaceae, Pyronemataceae and Pezizaceae the most common. Twenty-two MOTUs were collected in the natural T. macrosporum site. Basidiomycota morphotypes were plentiful (70.6%) and Thelephoraceae dominated. Each site had different taxa belowground with only T. macrosporum in common, being more abundant in the natural (18.2%) than in the cultivated (14.4%) site. Species richness, Simpson and Shannon diversity indices, taxonomic diversity, distinctness and variation of taxonomic distinctness were lower in the cultivated than in the natural site.

The microbiome structure of decomposing plant leaves in soil depends on plant species, soil pore sizes, and soil moisture content.

Microbial communities are known as the primary decomposers of all the carbon accumulated in the soil. However, how important soil structure and its conventional or organic management, moisture content, and how different plant species impact this process are less understood. To answer these questions, we generated a soil microcosm with decomposing corn and soy leaves, as well as soil adjacent to the leaves, and compared it to control samples. We then used high-throughput amplicon sequencing of the ITS and 16S rDNA regions to characterize these microbiomes. Leaf microbiomes were the least diverse and the most even in terms of OTU richness and abundance compared to near soil and far soil, especially in their bacterial component. Microbial composition was significantly and primarily affected by niche (leaves vs. soil) but also by soil management type and plant species in the fungal microbiome, while moisture content and pore sizes were more important drivers for the bacterial communities. The pore size effect was significantly dependent on moisture content, but only in the organic management type. Overall, our results refine our understanding of the decomposition of carbon residues in the soil and the factors that influence it, which are key for environmental sustainability and for evaluating changes in ecosystem functions.

Interannual dynamics of Tuber melanosporum and fungal communities in productive black truffle orchards amended with truffle nests.

Truffle growers devote great efforts to improve black truffle productivity, developing agronomic practices such as 'truffle nests' (peat amendments that are supplemented with truffle spore inoculum). It has been hypothesized that improved fruiting associated with nests is linked to stimulation of truffle mycelia previously established in soil or to changes generated in soil fungal community. To assess this, we used real-time PCR to quantify black truffle extraradical mycelium during 2 years after nests installation. We also characterized the fungal community via high-throughput amplicon sequencing of the ITS region of rRNA genes. We found that neither the abundance of truffle mycelium in nests nor in the soil-nest interphase was higher than in the bulk soil, which indicates that nests do not improve mycelial growth. The fungal community in nests showed lower richness and Shannon index and was compositionally different from that of soil, which suggests that nests may act as an open niche for fungal colonization that facilitates truffle fruiting. The ectomycorrhizal fungal community showed lower richness in nests. However, no negative relationships between amount of truffle mycelium and reads of other ectomycorrhizal fungi were found, thus countering the hypothesis that ectomycorrhizal competition plays a role in the nest effect.

Regional biogeography versus intra-annual dynamics of the root and soil microbiome.

BACKGROUND: Root and soil microbial communities constitute the below-ground plant microbiome, are drivers of nutrient cycling, and affect plant productivity. However, our understanding of their spatiotemporal patterns is confounded by exogenous factors that covary spatially, such as changes in host plant species, climate, and edaphic factors. These spatiotemporal patterns likely differ across microbiome domains (bacteria and fungi) and niches (root vs. soil). RESULTS: To capture spatial patterns at a regional scale, we sampled the below-ground microbiome of switchgrass monocultures of five sites spanning > 3 degrees of latitude within the Great Lakes region. To capture temporal patterns, we sampled the below-ground microbiome across the growing season within a single site. We compared the strength of spatiotemporal factors to nitrogen addition determining the major drivers in our perennial cropping system. All microbial communities were most strongly structured by sampling site, though collection date also had strong effects; in contrast, nitrogen addition had little to no effect on communities. Though all microbial communities were found to have significant spatiotemporal patterns, sampling site and collection date better explained bacterial than fungal community structure, which appeared more defined by stochastic processes. Root communities, especially bacterial, were more temporally structured than soil communities which were more spatially structured, both across and within sampling sites. Finally, we characterized a core set of taxa in the switchgrass microbiome that persists across space and time. These core taxa represented < 6% of total species richness but > 27% of relative abundance, with potential nitrogen fixing bacteria and fungal mutualists dominating the root community and saprotrophs dominating the soil community. CONCLUSIONS: Our results highlight the dynamic variability of plant microbiome composition and assembly across space and time, even within a single variety of a plant species. Root and soil fungal community compositions appeared spatiotemporally paired, while root and soil bacterial communities showed a temporal lag in compositional similarity suggesting active recruitment of soil bacteria into the root niche throughout the growing season. A better understanding of the drivers of these differential responses to space and time may improve our ability to predict microbial community structure and function under novel conditions.

Co-responses of bacterial and fungal communities to fire management treatments in Mediterranean pyrophytic ecosystems.

Cistus scrublands are pyrophytic ecosystems and occur widely across Mediterranean regions. Management of these scrublands is critical to prevent major disturbances, such as recurring wildfires. This is because management appears to compromise the synergies necessary for forest health and the provision of ecosystem services. Furthermore, it supports high microbial diversity, opening questions of how forest management practices impact belowground associated diversity as research related to this issue is scarce. This study aims to investigate the effects of different fire prevention treatments and site history on bacterial and fungi co-response and co-occurrence patterns over a fire-risky scrubland ecosystem. Two different site histories were studied by applying three different fire prevention treatments and samples were analyzed by amplification and sequencing of ITS2 and 16S rDNA for fungi and bacteria, respectively. The data revealed that site history, especially regarding fire occurrence, strongly influenced the microbial community. Young burnt areas tended to have a more homogeneous and lower microbial diversity, suggesting environmental filtering to a heat-resistant community. In comparison, young clearing history also showed a significant impact on the fungal community but not on the bacteria. Some bacteria genera were efficient predictors of fungal diversity and richness. For instance, Ktedonobacter and Desertibacter were a predictor of the presence of the edible mycorrhizal bolete Boletus edulis. These results demonstrate fungal and bacterial community co-response to fire prevention treatments and provide new tools for forecasting forest management impacts on microbial communities.

Mycorrhization of pecan trees (Carya illinoinensis) with commercial truffle species: Tuber aestivum Vittad. and Tuber borchii Vittad.

Pecan (Carya illinoinensis) is an economically important nut tree native to the Mississippi basin and cultivated worldwide. In North America, species of truffles are regularly found fruiting in productive pecan orchards and the truffle genus Tuber appears to be abundant in pecan ectomycorrhizal (EM) communities. As an initial step to determine the feasibility of co-cropping European truffle species with pecan, we evaluated whether mycorrhizae of highly esteemed European truffle species (Tuber aestivum Vittad. T. borchii and T. macrosporum) could be formed on pecan seedlings. Seedlings were inoculated with truffle spores and were grown in a greenhouse for 10 months. Levels of EM colonization were estimated visually and quantified by counting EM tips. Ectomycorrhizae were identified both morphologically and molecularly with species-specific amplification and by sequencing of the ITS region of the nuclear ribosomal DNA (nrDNA). Both T. borchii and T. aestivum spores produced well-formed ectomycorrhizae on pecan seedlings with average root colonization levels of about 62% and 42%, respectively, whereas no ectomycorrhizae of T. macrosporum were formed. The anatomy and morphology of these truffle ectomycorrhizae on pecan was characterized. The co-cropping of T. aestivum and T. borchii may hold promise as an additional stream of revenue to pecan growers, although, further studies are needed to assess whether this symbiosis is maintained after planting in the field and whether truffle production can be supported by this host species.

Mycorrhization of Quercus robur L., Quercus cerris L. and Corylus avellana L. seedlings with Tuber macrosporum Vittad.

Tuber macrosporum Vittad. is not a common truffle species, but with remarkable organoleptic qualities and much economic interest. After the addition of truffle spore slurry, 30 seedlings of Quercus robur L., Quercus cerris L. and Corylus avellana L. were grown inside a greenhouse for 11 months before evaluation of the mycorrhizal level. Two different potting mixes were used: a natural soil-based potting mix for Q. robur, Q. cerris and C. avellana and a peat-based potting mix for Q. robur. Quercus robur planted in soil potting mix was the most receptive towards the truffle spore inoculum, with a level of formation of T. macrosporum ectomycorrhizas (ECMs) of approximately 14 %, ranging from a minimum of ∼4 % to a maximum of ∼44 % in different seedlings. No T. macrosporum ECMs developed on Q. cerris (soil potting mix) or on Q. robur (peat potting mix), whereas a low percentage of ECMs was detected on only three C. avellana (soil potting mix) seedlings. The fungus Sphaerosporella brunnea (Alb. & Schwein.) Svrcek & Kubicka was also detected as a contaminant on almost half the truffle-inoculated seedlings. A new detailed description of the morphological and anatomical characteristics of T. macrosporum ECMs and their DNA-based verification with species-specific markers were also reported.

Non-target impacts of fungicide disturbance on phyllosphere yeasts in conventional and no-till management.

Fungicides reduce fungal pathogen populations and are essential to food security. Understanding the impacts of fungicides on crop microbiomes is vital to minimizing unintended consequences while maintaining their use for plant protection. However, fungicide disturbance of plant microbiomes has received limited attention, and has not been examined in different agricultural management systems. We used amplicon sequencing of fungi and prokaryotes in maize and soybean microbiomes before and after foliar fungicide application in leaves and roots from plots under long-term no-till and conventional tillage management. We examined fungicide disturbance and resilience, which revealed consistent non-target effects and greater resiliency under no-till management. Fungicides lowered pathogen abundance in maize and soybean and decreased the abundance of Tremellomycetes yeasts, especially Bulleribasidiaceae, including core microbiome members. Fungicide application reduced network complexity in the soybean phyllosphere, which revealed altered co-occurrence patterns between yeast species of Bulleribasidiaceae, and Sphingomonas and Hymenobacter in fungicide treated plots. Results indicate that foliar fungicides lower pathogen and non-target fungal abundance and may impact prokaryotes indirectly. Treatment effects were confined to the phyllosphere and did not impact belowground microbial communities. Overall, these results demonstrate the resilience of no-till management to fungicide disturbance, a potential novel ecosystem service provided by no-till agriculture.

Soil Origin and Plant Genotype Modulate Switchgrass Aboveground Productivity and Root Microbiome Assembly.

Switchgrass (Panicum virgatum) is a model perennial grass for bioenergy production that can be productive in agricultural lands that are not suitable for food production. There is growing interest in whether its associated microbiome may be adaptive in low- or no-input cultivation systems. However, the relative impact of plant genotype and soil factors on plant microbiome and biomass are a challenge to decouple. To address this, a common garden greenhouse experiment was carried out using six common switchgrass genotypes, which were each grown in four different marginal soils collected from long-term bioenergy research sites in Michigan and Wisconsin. We characterized the fungal and bacterial root communities with high-throughput amplicon sequencing of the ITS and 16S rDNA markers, and collected phenological plant traits during plant growth, as well as soil chemical traits. At harvest, we measured the total plant aerial dry biomass. Significant differences in richness and Shannon diversity across soils but not between plant genotypes were found. Generalized linear models showed an interaction between soil and genotype for fungal richness but not for bacterial richness. Community structure was also strongly shaped by soil origin and soil origin × plant genotype interactions. Overall, plant genotype effects were significant but low. Random Forest models indicate that important factors impacting switchgrass biomass included NO3-, Ca2+, PO43-, and microbial biodiversity. We identified 54 fungal and 52 bacterial predictors of plant aerial biomass, which included several operational taxonomic units belonging to Glomeraceae and Rhizobiaceae, fungal and bacterial lineages that are involved in provisioning nutrients to plants. IMPORTANCE Greenhouse gas reduction, carbon sequestration, and environmental remediation are top research themes within the U.S. Department of Energy funded bioenergy research centers. The utilization of unproductive agricultural land for bioenergy crop production is one of the most promising directions to achieve these goals. Switchgrass is a model biofuel system: it is adapted to a wide variety of geographical regions in North America, it is protective of soil and water resources, and it can be productive in low-fertility soils, but its profitability depends greatly on the biomass yield. Beneficial microbes have known roles in modulating plant biomass production but their interaction with soil geography, and switchgrass cultivars were not thoroughly studied. This study aims to fill important knowledge gaps and to serve as a foundation for switchgrass biomass promotion through microbe selection with an ultimate goal of facilitating sustainable bioenergy crop production.

Effect of earthworms on mycorrhization, root morphology and biomass of silver fir seedlings inoculated with black summer truffle (Tuber aestivum Vittad.).

Species of the genus Tuber have gained a lot of attention in recent decades due to their aromatic hypogenous fruitbodies, which can bring high prices on the market. The tendency in truffle production is to infect oak, hazel, beech, etc. in greenhouse conditions. We aimed to show whether silver fir (Abies alba Mill.) can be an appropriate host partner for commercial mycorrhization with truffles, and how earthworms in the inoculation substrate would affect the mycorrhization dynamics. Silver fir seedlings inoculated with Tuber. aestivum were analyzed for root system parameters and mycorrhization, how earthworms affect the bare root system, and if mycorrhization parameters change when earthworms are added to the inoculation substrate. Seedlings were analyzed 6 and 12 months after spore inoculation. Mycorrhization with or without earthworms revealed contrasting effects on fine root biomass and morphology of silver fir seedlings. Only a few of the assessed fine root parameters showed statistically significant response, namely higher fine root biomass and fine root tip density in inoculated seedlings without earthworms 6 months after inoculation, lower fine root tip density when earthworms were added, the specific root tip density increased in inoculated seedlings without earthworms 12 months after inoculation, and general negative effect of earthworm on branching density. Silver fir was confirmed as a suitable host partner for commercial mycorrhization with truffles, with 6% and 35% mycorrhization 6 months after inoculation and between 36% and 55% mycorrhization 12 months after inoculation. The effect of earthworms on mycorrhization of silver fir with Tuber aestivum was positive only after 6 months of mycorrhization, while this effect disappeared and turned insignificantly negative after 12 months due to the secondary effect of grazing on ectomycorrhizal root tips.

Lack of Linkages among Fruiting Depth, Weight, and Maturity in Irrigated Truffle Fungi Marks the Complexity of Relationships among Morphogenetic Stages.

The highly prized black truffle (Tuber melanosporum) has become a model species for ectomycorrhizal fungi biology. However, several questions concerning its reproductive phase remain unanswered. To provide new hypotheses on the fruitbody formation process, we have explored the causal links among development characters of black truffle fruitbodies that are primarily linked to either the mating process, fruitbody growing stage, or maturation. Path analysis was applied to test causal models outlining the relationships among fruitbody development characters such as fruiting depth, weight, shape, and spore maturity. These characters were investigated over a two-season survey and three soil typologies (plus peat-based substrate) under irrigated conditions. We found a clear and generalized relationship between fruitbody weight and shape. Among clusters of fruitbodies we found a positive relationship between the weight of the largest fruitbody and the weight of the remaining fruitbodies. However, no generalized relationships among characters linked to different development stages appeared. Our results were noticeably consistent across soil typologies, both for fruitbodies growing singly and in clusters, indicating that early-developing fruitbody characters did not influence characters linked to subsequent morphogenetic stages. The lack of links among stages opens new perspectives for pre-harvest quality management with stage-specific cultivation practices.