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Divergent biotic and abiotic filtering of root endosphere and rhizosphere soil fungal communities along ecological gradients

Plant roots assemble two distinct microbial compartments: the rhizosphere (microbes in soil surrounding roots) and the endosphere (microbes within roots). Our knowledge of fungal community assembly in these compartments is limited, especially in wetlands. We tested the hypothesis that biotic factors would have direct effects on rhizosphere and endosphere assembly, while abiotic factors would have direct and indirect effects. Using a field study, we examined the influences of salinity, water level and biotic factors on baldcypress (Taxodium distichum) fungal communities. We found that endosphere fungi were correlated with host density and canopy cover as opposed to rhizosphere, suggesting that hosts can impose selective filters on fungi colonizing into their roots. Meanwhile, local abiotic conditions strongly influenced both rhizosphere and endosphere diversity in opposite patterns: e.g. highest endosphere diversity (hump-shaped) while lowest rhizosphere diversity (U-shaped) at intermediate salinity levels. These results indicate that the assembly and structure for the root endosphere and rhizosphere within a host can be shaped by different processes. Our results also highlight the importance of assessing how environmental changes affect plant and plant-associated fungal communities in wetland ecosystems where saltwater intrusion and sea level rise are major threats to both plant and fungal communities.

FEMS Microbiology Ecology

Fungal endophyte effects on invasive Phragmites australis performance in field and growth chamber environments

Manipulating plant microbiomes may provide control of invasive species. Invasive Phragmites australis has spread rapidly in North American wetlands, causing significant declines in native biodiversity. To test microbiome effects on host growth, we inoculated four common fungal endophytes into replicated Phragmites genotypes and monitored their growth in field and growth chamber environments. Inoculations were highly successful in the growth chamber but inoculated plants in the field were rapidly colonized by diverse endophytes from the local environment. There were significant genotype effects and minimal inoculation effects in both experiments with a significant inoculation × genotype interaction on tiller height in the field. Our results demonstrate that endophyte inoculation treatments are feasible, but repeated inoculations may be required to maintain high titer in plants subject to endophyte colonization from the local environment. Future studies should investigate a wider range of fungal endophytes to identify taxa that inhibit Phragmites and other invaders.

Fungal Ecology

Octocoral diseases in a changing ocean

Octocorals (Cnidaria, Octocorallia) constitute a geographically widely distributed and common group of marine invertebrates commonly referred to as “soft-corals,” “sea fans,” “horny corals,” “sea feathers,” and “sea plumes.” They are found from shallow coastal habitats to mesophotic and abyssal depths. Octocorals are important members of most Atlantic-Caribbean, Indo-Pacific, and Mediterranean coastal and mesophotic reef communities; however, information about their susceptibility to diseases, predation, and competition, and their relationship with changing environmental conditions is limited. At least 19 diseases have been observed in at least 42 common octocoral species throughout their range. Twelve of these have been reported in the wider Caribbean (CA), one in Brazil (BR), two in the Mediterranean (ME), one in the Eastern Pacific (EP), and three in the western Pacific (WP). Pathogenic and/or environmental causes have been identified for eight diseases, including viruses, terrestrial fungi, protozoans, bacteria and cyanobacteria, filamentous algae, parasitic copepods, and high temperature. Only a few of the suspected pathogens have been tested with Koch’s postulates. At least eight disease outbreaks have led to extensive octocoral mortalities in the CA, ME, BR, and EP with detrimental ecological consequences. The fungal disease Aspergillosis has produced the highest mortalities in the CA and the EP. Other fungi, protozoans, and the bacterium Vibrio coralliilyticus were identified as potential causes of the death of millions of colonies in two Mediterranean disease outbreaks. Bacterial and fungal agents seemed to be responsible for the mass mortalities in Brazil and the WP. Most outbreaks in all regions were linked to high thermal anomalies associated with climate change, which seems to be the major driver. Other biological stressors such as predation and/or competition produce injuries that may contribute to the spread of infections and mortality. Overfishing of common predators could lead to population explosions of octocoral-feeding species that produced mass mortalities in some Caribbean localities. Our lack of knowledge of causes and pathogenesis of octocoral diseases parallels that of hard corals. New diseases are being described almost every year concomitant with increasing seawater temperatures. The ecological and economic consequences could be significant, with drastic changes in the seascape of shallow coral reefs and other coastal marine habitats and reduction of their ecological services. Given our limited knowledge, our best options for recovery of octocorals and coral reefs in general include sound management of coastal fisheries, development and tourism; reduction of land- and sea-based pollution; and abating effects of climate change.

Book chapter

Ambrosia beetles (Coleoptera: Curculionidae) can directly transmit the fungal pathogens responsible for Rapid ʻŌhiʻa Death

The ecologically and culturally vital tree species, ʻōhiʻa lehua ( Metrosideros polymorpha ), is threatened by the fungal pathogens Ceratocystis lukuohia and Ceratocystis huliohia , the causal agents of the disease complex called Rapid ʻŌhiʻa Death (ROD). Four invasive ambrosia beetle (Coleoptera: Curculionidae: Scolytinae) species in the Xyleborini tribe colonize ROD Ceratocystis -infested ‘ōhiʻa and produce inoculum through their frass; however, the potential for direct transmission of the ROD fungi by these beetles was unknown. We fulfilled Leach's rules to support insect transmission of ROD by documenting the visitation of these ambrosia beetles to healthy ‘ōhiʻa trees, culturing C. lukuohia and C. huliohia from the ROD-associated beetles using three different collection methods at multiple study sites, and challenging healthy ʻōhiʻa seedlings with beetles that were exposed to both C. lukuohia and C. huliohia cultures. We documented all four invasive ROD-associated ambrosia beetle species including Xyleborinus saxesenii , Xyleborus affinis , Xyleborus ferrugineus , and Xyleborus perforans to regularly visit healthy ʻōhiʻa trees on sticky traps. Viable Ceratocystis propagules were isolated from all species, and C. lukuohia was most commonly isolated of the two ROD-causing fungi. Consistently across all collection techniques, ROD Ceratocystis spp. were detected on just under 3% of all assayed beetles, with the highest detection rate from X. affinis . All four beetle species were capable of directly transmitting both pathogens to healthy ʻōhiʻa seedlings with a high rate of transfer. Ceratocystis spp. are highly virulent pathogens in trees, and a single inoculation can result in tree death, therefore any direct transmission is a cause for concern. After meeting the criteria of Leach's rules, we propose that Xi. saxesenii , X. affinis , X. ferrugineus , and X. perforans are vectors of C. lukuohia and C. huliohia , particularly in areas of high ROD pressure and tree stress.

Hawaii

Studies to assess natural resource recovery and evaluate monitoring methods for restored bottomland hardwood forests

The Natural Resource Damage Assessment and Restoration process assesses natural resource injury due to oil or chemical spills and calculates the damages to compensate the public for those injuries. Ecological restoration provides a means for recovering resources injured or lost due to contamination from oil or chemical spills by restoring the injured site after remediation, or acquiring or reconstructing equivalent resources off site to replace those lost due to the spill. In the case of restored forests, once restoration is implemented, monitoring of forest ecology helps keep recovery on track, with the maturation of forest vegetation, recovered soil conditions, and development of microbial, fungal, and faunal communities, necessary for ecologically functioning forests. This series of papers focuses on applying methods for monitoring restoration progress in forest vegetation and soils, and amphibian, avian, and mammalian communities, assessing strengths and weaknesses of different methods, and evaluating levels of effort needed to obtain accurate indications of forest ecological condition.

Integrated Environmental Assessment and Management

Context-dependent variation in persistence of host populations in the face of disease

Research Highlight : Valenzuela-Sánchez, A., Azat, C., Cunningham, A. A., Delgado, S., Bacigalupe, L. D., Beltrand, J., Serrano, J. M., Sentenac, H., Haddow, N., Toledo, V., Schmidt, B. R., & Cayuela, H. (2022). Interpopulation differences in male reproductive effort drive the population dynamics of a host exposed to an emerging fungal pathogen. Journal of Animal Ecology , 00 , 1– 12. https://doi.org/10.1111/1365-2656.13603 . Understanding the nuances of population persistence in the face of a stressor can help predict extinction risk and guide conservation actions. However, the exact mechanisms driving population stability may not always be known. In this paper, Valenzuela-Sánchez et al. (2022) integrate long-term mark–recapture data, focal measurements of reproductive effort, a population matrix model and inferences on life-history variation to reveal differences in demographic response to disease in a susceptible frog species ( Rhinoderma darwinii ). Valenzuela-Sánchez et al. found that demographic compensation via recruitment explained the positive population growth rate in their high disease prevalence population whereas the low disease prevalence population did not compensate and thus had decreasing population growth. Compensatory recruitment was likely due to the high probability of males brooding, and the high number of brooded larvae in the high prevalence population compared to low prevalence and disease-free populations. Valenzuela-Sánchez et al. also document faster generation times in the high prevalence population, which may indicate a faster life history that may be contributing to the population's ability to compensate for reduced survival. Lastly, the authors find a positive relationship between disease prevalence and the proportion of juveniles in a given population that suggest that there may be a threshold for disease prevalence that triggers increased reproductive effort. Altogether, their study provides novel support for increased reproductive effort as the pathway for compensatory recruitment leading to increasing population growth despite strong negative effects of disease on adult survival. Their results also caution the overgeneralization of the effects of stressors (e.g. disease) on population dynamics, where context-dependent responses may differ among host populations of a given species.

Journal of Animal Ecology

Resource availability controls fungal diversity across a plant diversity gradient

Despite decades of research, the ecological determinants of microbial diversity remain poorly understood. Here, we test two alternative hypotheses concerning the factors regulating fungal diversity in soil. The first states that higher levels of plant detritus production increase the supply of limiting resources (i.e. organic substrates) thereby increasing fungal diversity. Alternatively, greater plant diversity increases the range of organic substrates entering soil, thereby increasing the number of niches to be filled by a greater array of heterotrophic fungi. These two hypotheses were simultaneously examined in experimental plant communities consisting of one to 16 species that have been maintained for a decade. We used ribosomal intergenic spacer analysis (RISA), in combination with cloning and sequencing, to quantify fungal community composition and diversity within the experimental plant communities. We used soil microbial biomass as a temporally integrated measure of resource supply. Plant diversity was unrelated to fungal diversity, but fungal diversity was a unimodal function of resource supply. Canonical correspondence analysis (CCA) indicated that plant diversity showed a relationship to fungal community composition, although the occurrence of RISA bands and operational taxonomic units (OTUs) did not differ among the treatments. The relationship between fungal diversity and resource availability parallels similar relationships reported for grasslands, tropical forests, coral reefs, and other biotic communities, strongly suggesting that the same underlying mechanisms determine the diversity of organisms at multiple scales. ?? 2006 Blackwell Publishing Ltd/CNRS.

Ecology Letters

Threats posed by the Fungal Kingdom to humans, wildlife, and agriculture

The Fungal Kingdom includes at least six million eukaryotic species and is remarkable with respect to its profound impact on global health, biodiversity, ecology, agriculture, manufacturing, and biomedical research. Approximately 625 fungal species have been reported to infect vertebrates, 200 of which can be human-associated, either as commensals and members of our microbiome or as pathogens that cause infectious diseases. These organisms pose a growing threat to human health with the global increase in the incidence of invasive fungal infections, prevalence of fungal allergy, and the evolution of fungal pathogens resistant to some or all current classes of antifungals. More broadly, there has been an unprecedented and worldwide emergence of fungal pathogens impacting animal and plant biodiversity. Approximately 8,000 species of fungi and Oomycetes are associated with plant disease. Indeed, across agriculture, such fungal diseases of plants include new devastating epidemics of trees and jeopardize food security worldwide by causing epidemics in staple and commodity crops that feed billions. Further, ingestion of mycotoxins contributes to ill health and causes cancer. Coordinated international research efforts, enhanced technology translation, and greater policy outreach by scientists are needed to more fully understand the biology and drivers that underlie the emergence of fungal diseases and to mitigate against their impacts. Here, we focus on poignant examples of emerging fungal threats in each of three areas: human health, wildlife biodiversity, and food security.

mBio

Possible control of acute outbreaks of a marine fungal pathogen by nominally herbivorous tropical reef fish

Primary producers in terrestrial and marine systems can be affected by fungal pathogens threatening the provision of critical ecosystem services. Crustose coralline algae (CCA) are ecologically important members of tropical reef systems and are impacted by coralline fungal disease (CFD) which manifests as overgrowth of the CCA crust by fungal lesions causing partial to complete mortality of the CCA host. No natural controls for CFD have been identified, but nominally herbivorous fish could play a role by consuming pathogenic fungi. We documented preferential grazing on fungal lesions by adults of six common reef-dwelling species of herbivorous Acanthuridae and Labridae, (surgeonfish and parrotfish) which collectively demonstrated an ~ 80-fold higher grazing rate on fungal lesions relative to their proportionate benthic coverage, and a preference for lesions over other palatable substrata (e.g. live scleractinian coral, CCA, or algae). Furthermore, we recorded a ~ 600% increase in live CFD lesion size over an approximately 2-week period when grazing by herbivorous fish was experimentally excluded suggesting that herbivorous reef fish could control CFD progression by directly reducing biomass of the fungal pathogen. Removal rates may be sufficient to allow CCA to recover from infection and explain historically observed natural waning behaviour after an outbreak. Thus, in addition to their well-known role as determinants of macroalgal overgrowth of reefs, herbivorous fish could thus also be important in control of diseases affecting crustose coralline algae that stabilize the foundation of coral reef substrata.

Palmyra Atoll National Wildlife Refuge

A culture-based survey of fungi in soil from bat hibernacula in the eastern United States and its implications for detection of Geomyces destructans, the causal agent of bat white-nose syndrome

The recent emergence of white-nose syndrome (WNS), a fungal disease causing unprecedented mortality among hibernating bats of eastern North America, has revealed a knowledge gap regarding fungal communities associated with bats and their hibernacula. We used culture-based techniques to investigate the diversity of fungi in soil samples collected from 24 bat hibernacula in the eastern United States. Ribosomal RNA regions (internal transcribed spacer and partial intergenic spacer) were sequenced to preliminarily characterize isolates. Geomyces species were one of the most abundant and diverse groups cultured, representing approximately 33% of all isolates. Geomyces destructans was isolated from soil samples from three hibernacula in states where WNS is known to occur, and many of the other cultured Geomyces isolates likely represent undescribed taxa. Further characterization of the diversity of fungi that occur in hibernacula will both facilitate an improved understanding of the ecology of G. destructans within this complex fungal community and provide an opportunity to identify characteristics that differentiate G. destructans from non-pathogenic relatives.

Mycologia

Functional visual sensitivity to ultraviolet wavelengths in the Pileated Woodpecker (Dryocopus pileatus), and its influence on foraging substrate selection

Most diurnal birds are presumed visually sensitive to near ultraviolet (UV) wavelengths, however, controlled behavioral studies investigating UV sensitivity remain few. Although woodpeckers are important as primary cavity excavators and nuisance animals, published work on their visual systems is limited. We developed a novel foraging-based behavioral assay designed to test UV sensitivity in the Pileated Woodpecker ( Dryocopus pileatus ). We acclimated 21 wild-caught woodpeckers to foraging for frozen mealworms within 1.2 m sections of peeled cedar ( Thuja spp.) poles. We then tested the functional significance of UV cues by placing frozen mealworms behind UV-reflective covers, UV-absorptive covers, or decayed red pine substrates within the same 1.2 m poles in independent experiments. Behavioral responses were greater toward both UV-reflective and UV-absorptive substrates in three experiments. Study subjects therefore reliably differentiated and attended to two distinct UV conditions of a foraging substrate. Cue-naïve subjects showed a preference for UV-absorptive substrates, suggesting that woodpeckers may be pre-disposed to foraging from such substrates. Behavioral responses were greater toward decayed pine substrates (UV-reflective) than sound pine substrates suggesting that decayed pine can be a useful foraging cue. The finding that cue-naïve subjects selected UV-absorbing foraging substrates has implications for ecological interactions of woodpeckers with fungi. Woodpeckers transport fungal spores, and communication methods analogous to those of plant-pollinator mutualisms (i.e. UV-absorbing patterns) may have evolved to support woodpecker-fungus mutualisms.

Physiology & Behavior

Novel genome characteristics contribute to the invasiveness of Phragmites australis (common reed)

The rapid invasion of the non-native Phragmites australis (Poaceae, subfamily Arundinoideae) is a major threat to native wetland ecosystems in North America and elsewhere. We describe the first reference genome for P . australis and compare invasive (ssp. australis ) and native (ssp. americanus ) genotypes collected from replicated populations across the Laurentian Great Lakes to deduce genomic bases driving its invasive success. Here, we report novel genomic features including a Phragmites lineage-specific whole genome duplication, followed by gene loss and preferential retention of genes associated with transcription factors and regulatory functions in the remaining duplicates. Comparative transcriptomic analyses revealed that genes associated with biotic stress and defence responses were expressed at a higher basal level in invasive genotypes, but native genotypes showed a stronger induction of defence responses when challenged by a fungal endophyte. The reference genome and transcriptomes, combined with previous ecological and environmental data, add to our understanding of mechanisms leading to invasiveness and support the development of novel, genomics-assisted management approaches for invasive Phragmites .

Michigan, Ohio

Fish can get diseases too

Infectious diseases are increasingly recognized as an important component of the ecology of fish in the wild. Many of the viral, bacterial, protozoan and fungal pathogens of fish that were initially discovered in captive fish have their origin among wild populations; however, the impact of disease among these free-ranging stocks has been difficult to study. At the WFRC, combinations of field and laboratory investigations, aided by the tools of molecular biology, have begun to provide information on the ecology of infectious diseases among natural populations of fish in both freshwater and marine ecosystems.

Fact Sheet

Fungal life-styles and ecosystem dynamics: Biological aspects of plant pathogens, plant endophytes and saprophytes

This chapter discusses various biochemical, genetic, ecological, and evolutionary aspects of fungi that express either symbiotic or saprophytic life-styles. An enormous pool of potential pathogens exists in both agricultural and natural ecosystems, and virtually all plant species are susceptible to one or more fungal pathogens. Fungal pathogens have the potential to impact on the genetic structure of populations of individual plant species, the composition of plant communities and the process of plant succession. Endophytic fungi exist for at least part of their life cycles within the tissues of a plant host. This group of fungi is distinguished from plant pathogens because they do not elicit significant disease symptoms. However, endophytes do maintain the genetic and biochemical mechanisms required for infection and colonization of plant hosts. Fungi that obtain chemical nutrients from dead organic matter are known as saprophytes and are critical to the dynamics and resilience of ecosystems. There are two modes of saprophytic growth: one in which biomolecules that are amenable to transport across cell walls and membranes are directly absorbed, and another in which fungi must actively convert complex biopolymers into subunit forms amenable to transportation into cells. Regardless of life-style, fungi employ similar biochemical mechanisms for the acquisition and conversion of nutrients into complex biomolecules that are necessary for vegetative growth, production and dissemination of progeny, organismal competition, and survival during periods of nutrient deprivation or environmental inclemency.

Advances in Botanical Research

Manipulation of the Symbiodiniaceae microbiome confers multigenerational impacts on symbioses and reproductive ecology of its Exaiptasia diaphana host

Symbiodiniaceae-associated microbiota strongly influence cnidarian symbioses. We systematically reduced the bacterial and fungal communities associated with Symbiodiniaceae to study potential effects on the cnidarian holobiont Exaiptasia diaphana (Aiptasia). Clonal anemones were inoculated with xenic Breviolum minutum (SSB01) and microbiome-manipulated cultures after antibacterial or antifungal treatment. The asexual reproduction of pedal laceration allowed for three generations of clonal aposymbiotic Aiptasia to be utilised in this study, from the initial adult generation (G0), to the first (G1), and second (G2) generation. We inoculated small and large G1 Aiptasia with SSB01 algae and monitored onset of symbiosis, rate of algal proliferation, and holobiont characteristics. Sequencing the 16S and 18S rRNA gene regions identified significant differences in the bacterial and fungal communities of the G0 and G1 generations, alongside differences between the size classes of small and large G1 anemones. The microbiome of larger G1 individuals was distinct to the smaller G1 anemones, suggesting a microbiome maturation process. Control Breviolum minutum cultures exhibited a significantly greater proliferation rate in large G1 anemones when compared to antibacterial or antifungal treated cultures, whereas the opposite trend was documented in the small G1 anemones. Although no differences were observed between algal photochemical parameters, or the growth and polyp activity of G1 juveniles, we observed a significant influence in the production of G2 clones between treatments. Overall, we provide strong ecological implications of manipulating Symbiodiniaceae microbiome, not for the algae themselves, but for the maturation of the host Aiptasia, as well as for the cnidarian holobiont over multiple generations.

The ISME Journal

Effects of host species and environment on the skin microbiome of Plethodontid salamanders

The amphibian skin microbiome is recognized for its role in defence against pathogens, including the deadly fungal pathogen Batrachochytrium dendrobatidis (Bd). Yet, we have little understanding of evolutionary and ecological processes that structure these communities, especially for salamanders and closely related species. We investigated patterns in the distribution of bacterial communities on Plethodon salamander skin across host species and environments. Quantifying salamander skin microbiome structure contributes to our understanding of how host-associated bacteria are distributed across the landscape, among host species, and their putative relationship with disease. We characterized skin microbiome structure (alpha-diversity, beta-diversity and bacterial operational taxonomic unit [OTU] abundances) using 16S rRNA gene sequencing for co-occurring Plethodon salamander species (35 Plethodon cinereus , 17 Plethodon glutinosus , 10 Plethodon cylindraceus ) at three localities to differentiate the effects of host species from environmental factors on the microbiome. We sampled the microbiome of P. cinereus along an elevational gradient ( n = 50, 700–1,000 m a.s.l.) at one locality to determine whether elevation predicts microbiome structure. Finally, we quantified prevalence and abundance of putatively anti-Bd bacteria to determine if Bd-inhibitory bacteria are dominant microbiome members. Co-occurring salamanders had similar microbiome structure, but among sites salamanders had dissimilar microbiome structure for beta-diversity and abundance of 28 bacterial OTUs. We found that alpha-diversity increased with elevation, beta-diversity and the abundance of 17 bacterial OTUs changed with elevation (16 OTUs decreasing, 1 OTU increasing). We detected 11 putatively anti-Bd bacterial OTUs that were present on 90% of salamanders and made up an average relative abundance of 83% ( SD ± 8.5) per salamander. All salamanders tested negative for Bd. We conclude that environment is more influential in shaping skin microbiome structure than host differences in these congeneric species, and suggest that environmental characteristics that covary with elevation influence microbiome structure. High prevalence and abundance of anti-Bd bacteria may contribute to low Bd levels in these populations of Plethodon salamanders.

Journal of Animal Ecology

Has the time come for big science in wildlife health?

The consequences of wildlife emerging diseases are global and profound with increased burden on the public health system, negative impacts on the global economy, declines and extinctions of wildlife species, and subsequent loss of ecological integrity. Examples of health threats to wildlife include Batrachochytrium dendrobatidis , which causes a cutaneous fungal infection of amphibians and is linked to declines of amphibians globally; and the recently discovered Pseudogymnoascus (Geomyces) destructans , the etiologic agent of white nose syndrome which has caused precipitous declines of North American bat species. Of particular concern are the novel pathogens that have emerged as they are particularly devastating and challenging to manage. A big science approach to wildlife health research is needed if we are to make significant and enduring progress in managing these diseases. The advent of new analytical models and bench assays will provide us with the mathematical and molecular tools to identify and anticipate threats to wildlife, and understand the ecology and epidemiology of these diseases. Specifically, new molecular diagnostic techniques have opened up avenues for pathogen discovery, and the application of spatially referenced databases allows for risk assessments that can assist in targeting surveillance. Long-term, systematic collection of data for wildlife health and integration with other datasets is also essential. Multidisciplinary research programs should be expanded to increase our understanding of the drivers of emerging diseases and allow for the development of better disease prevention and management tools, such as vaccines. Finally, we need to create a National Fish and Wildlife Health Network that provides the operational framework (governance, policies, procedures, etc.) by which entities with a stake in wildlife health cooperate and collaborate to achieve optimal outcomes for human, animal, and ecosystem health.

EcoHealth

Fungi, fire, and feedbacks: Grasses and wildfire interact to alter ectomycorrhizal fungal communities and decrease tree seedling growth

Wildfire and its effects, including changes to soil biota and the introduction of invasive or seeded grasses, can cause long-term shifts in ecological communities. Post-wildfire establishment of long-lived trees and shrubs is a critical bottleneck to recovering native plant communities. Ectomycorrhizal fungi (EMF) can improve plant responses to stressors and influence seedling establishment following wildfire, but little is known about how introduced grasses alter plant-fungal relationships and influence woody plant recovery. We investigated how piñon pine ( Pinus edulis ) EMF colonization and growth responded to soil wildfire history and novel grasses. Piñon seedlings were grown in soils from areas that burned in a stand-replacing fire nearly two decades prior or in soils from unburned piñon-juniper woodlands. Each piñon was grown with an invasive grass ( Bromus tectorum ), a native rhizomatous grass ( Pascopyrum smithii ) or another piñon seedling. Even ∼20 years after fire, EMF community composition in burned areas differed from that of unburned woodlands. Fire history and plant neighbor identity interacted to affect EMF abundance. Piñon seedling biomass was positively associated with EMF abundance in unburned woodland soils, but not in post-burn soils, suggesting that the EMF community in unburned woodlands is more beneficial. Importantly, the presence of either an invasive or native grass had a negative effect on seedling growth and EMF abundance, resulting in an average 61.4 % drop in EMF abundance and altered EMF community composition. Our findings suggest that plant species interactions, long-term effects of fire on soil, and EMF may determine the trajectory of woodland recovery following wildfire.

Colorado