Ecosphere

Field ecologists often rely on observational data to understand the impact of environmental stressors and management interventions on natural systems. Natural and anthropogenic events (e.g. wildfires, protected areas, nutrient deposition) do not occur randomly in space, however, which can introduce bias into observational studies--which we refer to as causal selection bias. Field study designs that ignore the non-random occurrence of stressors may yield biased estimates of stressor effects on ecosystems. Matching methods commonly used in economics, political science and epidemiology offer a powerful framework for controlling for causal selection bias by identifying more comparable treatment and control sites. Although these methods are increasingly used in conservation, they are rarely used in ecological field-based studies. Here we review how Propensity Score Matching (PSM) can improve field sampling designs in ecology and strengthen causal identification of stressor effects. Then we apply this approach to a case study examining wildfire effects on forest recovery in California. We conclude with practical recommendations for implementing PSM to improve causal identification of ecological change, which is particularly important for developing effective management interventions.

Ecologists remotely track movement steps of animals (e.g., via global positioning systems) and use step selection functions to study the effect of environmental factors upon their movement decisions. Constructing such functions requires pairing each observed step with some number of unobserved but feasible comparison steps. Larger numbers of comparison steps generally yield better estimates but also incur potentially challenging computational demands. Thus, it is important to determine an appropriate number of comparison steps. No established guidance exists for this decision. Here, we use simulated tracks to assess how many comparison steps are needed, fitting each set of steps to a conditional logistic regression model. We monitor errors in estimated effects for several classes of tracks, identifying the number of comparison steps for which mean relative absolute error in estimated effects is consistently low. By this criterion, 32 comparison steps per observed step are needed for our primary class of simulated tracks. Tracks in more homogeneous landscapes, tracks with shorter mean step lengths, or shorter tracks generally require more comparison steps (ranging from 64 to 128 per observed step) to achieve the same level of accuracy. Longer tracks generally require fewer comparison steps (16 per observed step). These results clearly demonstrate that the number of comparison steps influences how well step selection functions estimate covariate effects and provides initial direction in a research area that currently lacks quantitative guidance. Movement ecologists should take care when selecting the number of comparison steps paired with each observed step because those decisions matter.

O_LISpawning salmon transport marine-derived nutrients (MDN) into riparian forests, influencing soil, plant, and animal communities, yet their effects on fungal communities remain poorly understood. C_LIO_LIWe used DNA metabarcoding to examine fungal responses to three spatial patterns of salmon-derived nitrogen (N) in southwest Alaska: (i) patchy inputs from wildlife-deposited carcasses, (ii) a 21-year carcass relocation experiment, and (iii) natural N gradients with distance from streams. C_LIO_LIDecomposing carcasses increased saprotrophic fungal diversity, identifying taxa responsible for salmon carcass decomposition. Long-term carcass relocation reduced diversity of medium-distance fringe ectomycorrhizal fungi (EMF), whereas recent, patchy carcass inputs increased diversity of both medium-distance fringe and long-distance EMF--guilds often associated with low-nutrient environments. Along natural stream N gradients, EMF responses varied markedly within functional guilds and genera, revealing unexpected variation in N sensitivity among closely related taxa. C_LIO_LIPulsed, spatially heterogeneous nutrient inputs enhanced diversity of typically nitrophobic EMF, likely reflecting their capacity to maintain extensive mycelial networks, exploit nutrient hotspots, and mobilize organic N and phosphorus. The diversity of responses along natural N gradients suggests that mechanisms linking EMF traits to nutrient acquisition and tolerance remain unresolved. Our findings emphasize the importance of linking fungal community composition with functional attributes and nutrient dynamics. C_LI

O_LIWhen eradication is unfeasible, invasive predator control should evaluate how removal affects ecological responses by native species. Assessments often use total invasive predator abundance to evaluate prey responses, yet intraspecific variation in diet and space use means that some subgroups cause disproportionate impacts. Identifying these problem individuals, and the spatial scales over which their impacts operate, can enable targeted spatially explicit removal to maximise impact reduction. However, despite individual-level information is often already collected during trapping operations it is seldom included when analysing predator impacts, potentially biasing the conservation outcomes expected under blanket removal. C_LIO_LIWe use a novel framework and two decades of invasive predator control data to estimate how individual variation in residency status influences the distance-dependent impacts of invasive American mink Neogale vison on water vole Arvicola amphibius occupancy across two prey surveys. We also develop a sub-model to predict mink residency status for individuals with missing age data. C_LIO_LIThe probability of capturing adult mink decreased with elevation and years of control, indicating that long-term control altered the resident population and demographic composition of mink around water vole sites. C_LIO_LIDistance-dependent negative impacts of mink varied by residency status, becoming negligible at approximately 20 km from water vole sites for resident mink and 2 km for transient. The spatial scale of mink impacts was largest during the first vole survey when resident mink were more abundant, and declined rapidly for the second survey, when mink were less abundant and spatially clustered. Our results suggest that water voles have benefited mostly from reducing resident mink rather than the total population, especially in early control phases. C_LIO_LIManagers can use our framework to develop spatially explicit and impact-based strategies, not restricted to invasive species control, to construct empirically informed management buffers around populations of conservation concern. Long-term efforts will change the landscape and invasive predator contexts, and thus we recommend iteratively updating and re-evaluating management outcome evaluations. We argue that incorporating individual heterogeneity improves our understanding of ecological mechanisms influencing management success but that the suitability of targeted strategies should be evaluated for target socioecological contexts. C_LI

Understanding the relationships between soil macrofauna and decomposition is crucial for predicting how land-use change impacts ecosystem function in fragmented systems. This is because soil macrofauna affect decomposition and also respond to the changes in abiotic conditions across habitat gradients. This study investigates edge effects on the macrofauna contributions to decomposition across forest-field ecotones. We used bait lamina assay to quantify aboveground and belowground feeding activity of soil macrofauna in Autumn 2025 in three deciduous forest-old field ecotones and one coniferous forest-old field ecotone, in Southwestern Ontario, Canada. Vegetation diversity and composition, LAI and soil characteristics (i.e., soil organic matter, pH, temperature and moisture) were measured at each plot along the ecotone. Pitfall trap data collected in Summer 2025 at the same sites were used to characterize macrofauna communities. We used generalized linear mixed effects models to estimate the effect of distance to edge, site, and depth into the soil on bait lamina consumption and soil macrofauna, with transect nested within site as random effects. Consumption activity increased with distance into the forest from the field, with the edge representing an intermediate; and, decreased with increasing depth into the soil. In contrast, soil macrofauna abundance, especially isopods, decrease with distance into the forest from the field. These trends varied significantly across sites, so that consumption activity and abundance sometimes remained constant across the ecotone (i.e., site x distance interaction). The results demonstrate that macrofaunal contributions to bait consumption varied along the ecotone, shaped by interacting environmental gradients and shifts in community composition unique to each site.

Identifying optimal grazing intensities for sustainable population growth is crucial for informing management strategies. Community-level studies frequently find that biodiversity peaks at intermediate grazing intensities, known as the Intermediate Disturbance Hypothesis. However, whether this hypothesis applies to population-level performance remains untested. Our stochastic integral projection models, parameterized with five-year demographic data of two co-occurring species, Morina chinensis and Deyeuxia flavens, on the Tibetan Plateau grasslands., indeed show a hump-shaped response in stochastic population growth rate ({lambda}S) to grazing intensities, providing empiral support for the Intermediate Disturbance Hypothesis at population level. Furthermore, populations with demographic compensation among vital rates are better able to buffer temporal variation in annual population growth rate and exhibit a much smaller decline in {lambda}S under heavy grazing. Our study provides mechanistic insights into demographic processes driving population dynamics across grazing levels, thereby better informing grazing management strategies.

Devils gardens are among the most striking ant-plant mutualisms in Amazonian forests. In this system, the tree Duroia hirsuta is associated with the ant Myrmelachista schumanni, which actively removes neighboring vegetation and maintains nearly monodominant patches of the host plant. Despite the apparent efficiency of this system, repeated field observations at the Tiputini Biodiversity Station, Yasuni Biosphere Reserve, revealed that individuals of Palicourea alba recurrently occur within active devils gardens. Palicourea alba closely resembles dead plant material, exhibiting leaf morphology and coloration that strongly mimic the surrounding litter layer, and appears to be uncommon outside these gardens. To our knowledge, this "dead-leaf" masquerade has not been previously documented in this intensively studied system, making it a particularly striking and unexpected observation. To evaluate whether this masquerade facilitates persistence within devils gardens, we surveyed 35 gardens and recorded P. alba in 19 (52.8%). When present, P. alba covered on average 27% of plot area, while mean herbivory across sampled leaves remained low (8.6%). Generalized linear mixed models showed that P. alba cover decreased significantly with increasing herbivory (F = 8.09, p = 0.0159), whereas herbivory increased with leaf-litter cover (F = 8.73, p = 0.0120). Field observations further revealed that many individuals are nearly indistinguishable from dry leaf litter, suggesting a role for visual crypsis or masquerade. Together, these results indicate initially, that the persistence of P. alba within devils gardens is mediated by a multi-layered ecological filtering process. First, masquerade likely reduces detection by M. schumanni, allowing seedlings to escape ant-mediated removal. Second, low herbivory suggests either enemy avoidance or reduced apparency to herbivores within the simplified understory. Third, spatial heterogeneity in leaf-litter cover may create microhabitats where both ant activity and herbivore pressure are modulated, reinforcing establishment success. This system thus represents a previously undocumented mechanism in which plant-litter resemblance enables persistence within a highly structured, biotically filtered habitat, highlighting how subtle trait-mediated interactions may modulate outcomes in otherwise strongly deterministic mutualisms.

A central question in biodiversity conservation is whether species can sustain viable population under current and future atmospheric N deposition. Assessing species viability under N deposition requires demographic studies integrating species vital rates responses to long-term N deposition across different levels. However, studies of this nature are rare. Our integral projection models (IPMs), parameterized with demographic data, revealed differing responses of two functionally similar coexisting species, Stipa bungeana and Leymus secalinus, to 12 years of N deposition at low N addition levels (1.15 and 2.30 g N m-2 yr-1) and high N addition levels (4.60, 9.20, and 13.80 g N m-2 yr-1) on the Loess Plateau grasslands. We found that the reduced survival across N addition levels was partially compensated by increased contributions from growth, shrinkage, and fecundity, alleviating the population decline of S. bungeana (with a longer lifespan and generation time) under different N additions. Contrasting, more positive correlations among vital rate enabled the population of L. secalinus (with a shorter lifespan and generation time) to track N additions, with population growth under low N additions and population decline under high N additions. Our results illustrate that the demographic response to N deposition may vary considerably between functionally similar coexisting species, and species with demographic compensation can buffer populations against N deposition while with demographic lability enable populations to track N deposition. Furthermore, our study demonstrates the potential of using life-history traits to predict species viability under N deposition, thereby informing biodiversity conservation under global change.

Native ash trees are destined for functional extinction in North America due to the spread of the non-native emerald ash borer. Yet, the consequences of ash loss for soil fungi are unclear. To address this, we employed a factorial study of forest soil fungi in two hydropedological soil types beneath four canopy tree species -- including white ash (Fraxinus americana). Sporocarp surveys and community DNA metabarcoding from soil samples revealed patterns in fungal communities related to canopy tree species but not soil type. Ash trees supported a particularly rich soil fungal community that was distinguishable from communities beneath beech, birch, and maple. We identified over 100 fungal taxa (OTUs) that are at risk of decline or loss from the studied forest, due to their association with ash. Our results indicate that canopy tree species influence soil fungi much more broadly than just the species with which they have mycorrhizal associations.

Across species, changing environmental conditions are altering major phenological milestones. In seasonal environments, this can involve shifts in the timing of activity onset, growth, reproduction, and senescence; however, predicting these responses can be challenging and often requires species-specific information. In this study, we investigated phenological patterns of bumble bee colony development using observations from wild nests of two common species, Bombus griseocollis and B. impatiens. Using observations of nest traffic, we documented dates of nest-searching, peak worker activity, first gyne (new queen) production, and colony senescence over three years of field data collection. We also tested whether colony growth was density dependent, because longstanding life history models have shown that colony growth patterns determine the optimal timing of reproduction in constant environments. Here, we present a novel extension of these models, using them to infer that density-independent colony growth should be associated with extended phenology in warmer years, while density-dependent should have consistent phenology. In total, we found 79 wild nests, including 34 reproductive colonies (19 B. griseocolis and 15 B. impatiens). Bombus impatiens colonies were larger, had a longer activity period, and showed density-independent growth and later reproduction in warmer years, while B. griseocollis colonies were smaller, shorter-lived, and showed density-dependent growth and similar dates of reproduction across years. Both species had higher gyne production in warmer years. These results highlight the role of life history theory for predicting interspecific variation in phenological changes, as well as a research framework for exploring possible evolutionary mismatches of existing life histories in changing environments.

Mortality-inducing disturbances are important, ubiquitous drivers of community composition and function. Importantly, human activities and climate change are increasingly altering disturbance regimes. Most disturbance studies focus on the effects of current disturbance regimes, rarely considering those of historical regimes. However, recent theoretical work predicts that historical regimes can leave persistent legacies, modulating the communitys response to novel disturbances and invasive species. Here, we complement this theoretical approach using a model bacterial system that experienced disturbance regimes for [~]120 generations, followed by novel regimes and invasions for another [~]120 generations. Our results show persistent effects of historical legacies on disturbance-diversity relationships. Furthermore, some combinations of past and novel regimes promote invasion with increasing resident diversity, while others prevent it; legacies may explain conflicting diversity-invasibility relationships. These findings demonstrate the importance of historical legacies in disturbance-prone ecosystems, and underscore the challenges in predicting future community responses to disturbance regime changes.

Many Sockeye salmon (Oncorhynchus nerka) populations have declined over recent decades, and climate change is likely to exacerbate these declines through direct and indirect ecological effects. The response to the associated environmental changes is likely to vary among life stages, populations, and regions. Quantitative estimates of climate change driven impacts that account for this variability could fill a critical gap and provide forward-looking insights into how sockeye are expected to respond to future climate-driven change across their lifecycle. To address this need we developed a hierarchical population dynamics model parameterized with juvenile, adult return and spawner abundance data from 13 sockeye salmon populations from Washington State to northern British Columbia. We used a formal causal inference framework that paired salmon abundance data with a suite of environmental covariates hypothesized to represent ecological conditions across the lifecycle. We used the model to estimate population-specific responses to each environmental driver, then combined parameter estimates with projections from down-scaled climate change models to estimate productivity responses to anticipated environmental change. We found that historical sockeye productivity was strongly associated with environmental covariates, which explained more interannual variability in return abundance than spawner abundance in most populations. However, the life stages and specific environmental covariates with the largest impacts differed among populations and regions, often displaying a latitudinal gradient. Increases in coastal ocean temperatures and mixed layer depth generally had negative effects though they varied among regions. Increased freshwater summer rearing and return migration temperatures had weaker but consistently negative effects. Under future climate conditions, projected changes in these environmental covariates are expected to result in substantial declines in productivity across most populations. Sockeye salmon display varying degrees of sensitivity to climate change across life stages, populations, and regions. Effective future management will require explicitly accounting for these life stage and population-specific responses.

1Chronic wasting disease (CWD), a fatal, transmissible disease in white-tailed deer (Odocoileus virginianus) and related species, is spreading across North America but has not yet been detected in Vermont, United States (U.S.). The Vermont Department of Fish and Wildlife, along with partner agencies, wants to develop a proactive prevention and response plan in anticipation of the eventual detection of the disease. Between September 2023 and September 2025, staff from the U.S. Geological Survey and the University of Vermont facilitated a structured decision-making (SDM) process with seven State and Federal agencies that have jurisdiction over some aspect of CWD management in Vermont. The aim of this process was to generate and evaluate alternative response plans against a range of long-term objectives important to the agencies. To aid in the evaluation of the alternatives, we developed a linked set of models for white-tailed deer population and disease dynamics, hunter participation and health, forest health, economic consequences, and agricultural opportunities related to the actions being contemplated as part of the response plan. We evaluated over 256 different permutations of management actions and used multi-criteria decision analysis, a branch of decision analysis designed to help decision makers navigate tradeoffs among competing objectives, to summarize the performance of those alternative strategies against the desired outcomes. Proactive actions--those designed to slow the arrival of CWD to Vermont--were moderately effective, but the most important proactive action was surveillance to detect the disease early after arrival, which triggered response actions after detection. With the insights generated by the SDM process and the results of the analyses, the participating agencies were able to identify a preferred strategy and outline the elements of a proactive response plan. This report describes the SDM process, the technical details of the modeling work, and the results of the analyses. It is intended to serve as the technical basis for Vermonts response plan. 2 Executive SummaryChronic wasting disease (CWD) is a fatal disease of deer, moose, and other cervids that is spreading across the United States (U.S.). As of December 2025, the nearest infected wild deer population to Vermont was in Pennsylvania, about 225 miles away; however, the disease has been detected closer to the state in New York and Quebec, Canada, but those detections are assumed to have been isolated and removed without additional spread in wild populations or captive facilities. The primary pathways of arrival to Vermont are natural spread in wild deer, import of infected live animals to captive cervid facilities, or disposal of imported infected deer parts on the Vermont landscape. Once CWD arrives, significant changes to many ecological, social, and economic resources are expected. Many State and Federal agencies could have a role in managing CWD, before and after its arrival. O_LIVermont Department of Fish and Wildlife (VDFW) has statutory responsibility for managing the wild deer and moose populations in the State, including through hunting; C_LIO_LIVermont Agency of Agriculture, Food and Markets (VAAFM) is responsible for oversight of the nine captive cervid facilities in the State; C_LIO_LIVermont Department of Forests, Parks and Recreation (VFPR) manages State Forests and Parks, which provide habitat for deer and recreational access for hunters; C_LIO_LIVermont Department of Health (VDH) provides information and guidance to the public that enables them to make informed decisions that can affect their health, including whether and how CWD poses any risk to humans; C_LIO_LIU.S. Fish and Wildlife Service (USFWS) manages two National Wildlife Refuges in the State, which provide public access, including to hunters; C_LIO_LIU.S. Department of Agriculture Wildlife Services can provide contractual support for wildlife management, including targeted reduction of deer populations; and C_LIO_LIU.S. Forest Service manages extensive forest resources in the State, which provide habitat for deer and access for hunters. C_LI These agencies have expressed interest in co-development of a long-term collaborative plan for management of CWD. From September 2023 through September 2025, they engaged in a series of meetings, facilitated using the principles of structured decision making to explore potential proactive and reactive management strategies, and how well those strategies might perform against the fundamental objectives being sought by the agencies. As part of this work, the U.S. Geological Survey and the University of Vermont developed quantitative predictive models to allow evaluation of alternative strategies against the fundamental objectives. The agencies identified seven fundamental objectives for a long-term (20-year) CWD management plan that arise from their enabling legislation and missions: O_LIMaximize herd health of native cervid populations in the state of Vermont; C_LIO_LIMaximize the public benefit of cervids (consumptive and non-consumptive); C_LIO_LIMinimize harm to other species, unique ecosystems, and forests; C_LIO_LIMinimize costs, staff time, and workload associated with CWD management; C_LIO_LIMinimize the effect (or potential effect) that CWD has on public health; C_LIO_LIMinimize the effect of CWD and its management on local economies; and C_LIO_LIMaintain opportunities for private landowners to engage in agricultural pursuits. C_LI In initial work to explore the effects of actions on the objectives, six broadly different alternative strategies were developed, where the strategies differed in philosophy, investment, timing, and proactive versus reactive emphasis. O_LIUnder the Status Quo strategy, existing protocols would remain in place until CWD is detected in the State, at which point an increase in surveillance and harvest opportunity would be implemented. C_LIO_LIThe Heavy Outreach strategy would rely primarily on outreach and communication with the public, notably hunters, to delay arrival, slow transmission, and mitigate impacts. C_LIO_LIThe Reduce Arrival strategy would invest primarily in proactive measures including both outreach and regulatory changes to slow arrival of CWD to the State. C_LIO_LIThe Prepare and React strategy would invest primarily in reactive measures after CWD arrives, with preparatory measures to detect it as early as possible and have in place the management tools to deploy upon arrival. C_LIO_LIThe Prevent and React strategy would make a moderate investment in both proactive and reactive measures. C_LIO_LIUnder the Kitchen Sink strategy, extensive investment would be made in both proactive and reactive measures. C_LI The estimated mean time to arrival of CWD in the State is 7.7 years under the Status Quo strategy. Concerted efforts to delay arrival could increase this time to 10.6 years, by significantly reducing two of the three arrival pathways. It will, however, take an average of 15-30 years after arrival to detect CWD in the State assuming effort is spread evenly throughout the State, owing to its slow growth in the deer population. Investment in surveillance leads to earlier detection and hence, an earlier opportunity to implement reactive measures. There are notable trade-offs among the alternative strategies in their ability to achieve outcomes of interest to the State, based on the analysis described in this report. O_LIThe arrival and spread of CWD can be slowed quite significantly, particularly with a combination of proactive and reactive measures, with the Kitchen Sink showing a 6-fold reduction in prevalence over 50 years compared to the Status Quo strategy. C_LIO_LIThe number of CWD-positive deer consumed by hunters, their families, and their friends largely tracks the prevalence of CWD in the population, although increased outreach efforts also contribute to reduced consumption. C_LIO_LIDeer harvest, and the economic benefits associated with it, is expected to increase under aggressive CWD management strategies, because the strategies encourage higher harvest to manage the disease, and lower prevalence of CWD keeps more hunters participating. C_LIO_LIThe positive outcomes associated with more aggressive proactive and reactive measures are counterbalanced by increases in costs to agencies, for surveillance, sharpshooting, outreach, and captive cervid management. C_LI Captive cervid farm operations in Vermont were estimated to be a low-risk introduction pathway because of the stringent regulations, agency oversight of importation, and disease testing that is already occurring. However, actions by VDFW could reduce the risk of spread from wild to captive deer. The more aggressive CWD management strategies reduce CWD prevalence and increase the long-term opportunity for captive cervid operations. The one exception to this is the Kitchen Sink strategy, which seeks to reduce the risk of arrival of CWD by proactively closing captive cervid facilities. The size of the statewide deer population is expected to be about the same in 20 years across all strategies, with all alternatives resulting in a decrease of about 5% compared to current population size estimates. In some strategies, the deer population decreases because of the effects of CWD; in others, the deer population decreases because of increased hunting that slows the transmission of CWD. After considering the results from the analysis of the six initial strategies, the participating agencies were interested in more carefully exploring the tradeoffs among three of the objectives: long-term prevalence of CWD in the State; the cost of management of CWD; and the opportunity for private farms to participate in captive cervid farming. An additional 256 strategies, which were permutations of the constituent actions under consideration, were created and analyzed. These results showed that the most effective way to keep CWD prevalence low over the long term was extensive surveillance to allow early detection, followed by sharpshooting or other targeted removal at a local level, to continually eliminate or reduce new incursions of the disease. This approach, however, is quite costly, so the balance between effort and effect was important to the agencies. Regarding captive cervid operations, proactive closing of captive cervid facilities does slightly lower the long-term prevalence of CWD and substantially diminishes the long-term costs of closing infected captive cervid farms in the future, but comes at the cost of farming opportunity. In September 2025, the participating agencies expressed a collective interest in a strategy that preserves captive cervid farming opportunities, invests heavily in surveillance for early detection of the disease, and prepares for targeted removal of deer when detections occur. Such a strategy, while potentially costly, achieves many of the objectives sought by the participating agencies on behalf of the people of Vermont. The agencies, without yet making any commitments, intend to undertake work to develop the details of this plan, discuss its aspects with stakeholders, and build support for its long-term implementation.

Space-for-time Cormack-Jolly-Seber (CJS) models have been developed to estimate survival of migrating animals that are imperfectly detected at spatially discrete sampling stations. However, space-for-time CJS models typically ignore survival over time and age-specific probability of movement, missing the diversity of life history strategies. To address these limitations, we extended space-for-time Cormack-Jolly-Seber models to explicitly incorporate time, age, and individual-level covariates to account for diverse life history strategies. We incorporate a sub-model that includes uncertainty in individuals age using a finite-mixture model. The detection and transition probabilities were parameterized using generalized linear models, facilitating flexible model specification and inclusion of spatiotemporal and individual covariates. We apply the model to detections of juvenile steelhead (Oncorhynchus mykiss) from two populations in the Snake River Basin in Idaho, USA, to estimate trends in survival with respect to population, age, time, and the length of individuals. Additionally, we generalized the model so that it can be applied to other systems and populations with different life history strategies and monitoring infrastructure.

The reintroduction of extinct or endangered species to restore ecosystem function is an essential aspect of rewilding. The Wilder Blean Project at West Blean and Thornden Woods in Canterbury, UK, is committed to rewilding natural processes and enhancing biodiversity in one of England's oldest and largest areas of ancient woodland. The introduction of European bison (Bison bonasus) is an important part of the project. However, how the reintroduction of large herbivores influences local biodiversity and ecosystem functions during the early stages of rewilding remains poorly understood. Soil samples were collected from the same sampling sites before and two years after bison were reintroduced and profiled by metagenomic sequencing using Oxford Nanopore Technologies sequencing platforms. The results showed that the alpha diversity of soil organisms did not change significantly before and after the introduction of European bison, while beta diversity showed modest shifts in community composition. The relative abundance of some nitrogen-fixing and photosynthetic microbial genera showed declines in the 2024 Bison Area, while the mycorrhizal fungus genus Rhizophagus was significantly less abundant than in the 2024 Control Area. Despite relatively stable taxonomic diversity, functional composition differed significantly between the 2022 and 2024 Bison areas and among the 2024 rewilding treatments, revealing a decoupling between taxonomic diversity and functional composition. Amino acid synthesis pathways and carbon metabolism pathways were significantly enriched. These findings highlight the potential of long-read Oxford Nanopore metagenomics to reveal functional shifts that may not be apparent from taxonomic diversity alone. Although these early-stage responses cannot yet predict long-term rewilding trajectories, continued longitudinal monitoring integrating microbial, soil physicochemical, and ecosystem-level measurements will be essential to determine the persistence and ecological significance of these functional shifts.

Replication is central to most experimental and sampling designs, increasing inferential power and capturing fine-scale data heterogeneity. However, its importance remains poorly evaluated in some ecological and evolutionary settings. This is the case of metabarcoding studies using DNA recovered from sedimentary archives, in which biological signals may integrate ecological information through depositional and burial processes, and are often inferred from a single sediment core per site. Here, we evaluated the effect of different types of replication using sedimentary DNA (sedaDNA) metabarcoding data from two genetic markers (mitochondrial COI and nuclear 18S), under a nested sampling design. The design included three intertidal sites, three spatially separated sediment cores per site (biological replicates), two sediment depth horizons per core, and eight PCR (technical) replicates per sediment sample. Variance partitioning showed that site identity and sediment age group together explained >70% of the variation in beta diversity, indicating that among-site spatial variation and stratigraphic variation were the dominant drivers of community composition. In contrast, variation among different cores within sites was small and non-significant (<5%). Among PCR replicates from the same sediment sample, richness varied substantially, whereas Shannon diversity was more consistent. Despite this variability, differences in community composition among technical replicates remained smaller than among biological replicates and site identity, indicating limited influence on broader ecological patterns. Community composition was highly similar among replicate cores within sites, consistent with stratigraphic coherence. These results indicate limited within-site heterogeneity and suggest that, under stratigraphically coherent conditions, increasing biological replication may yield limited additional information, whereas enhancing technical replication and stratigraphic resolution can improve ecological inference from sedaDNA metabarcoding datasets.

Root-associated fungal (RAF) communities can influence tree nutrient acquisition and performance, yet their structuring factors and relationship with tree growth remain poorly understood, particularly near environmental treelines. We characterized root-associated fungal (RAF) communities on fine roots of white spruce (Picea glauca [Moench] Voss) in paired forest and treeline plots across two elevational and one moisture-limited treeline ecotones in Alaska. Using ITS2 DNA metabarcoding of fine root tips, we characterized individual RAF community alpha and beta diversity and tree growth based on basal area increment (BAI) over 5-30 years. As a result, sampling site was the strongest predictor of RAF composition, explaining 19.6% of variation, while soil pH explained an additional 11.7%. Treeline effects on RAF communities were weaker and context-dependent, with treeline trees showing distinct community composition in the Alaska Range, lower alpha diversity across all Hill numbers in Interior Alaska, and higher ECM relative abundance in the Brooks Range. RAF composition did not differ between fast- and slow-growing trees within sites. In contrast, alpha diversity was negatively associated with tree growth: OTU richness, Shannon and Inverted Simpson indices significantly predicted BAI over the previous 5, 10 and 15 years, with fast-growing trees supporting less diverse RAF and ECM communities. This relationship was strongest for recent growth and weakened with longer BAI averaging periods. Our results suggest that high growth of mature P. glauca is not necessarily associated with greater RAF or ECM diversity or specific taxa but may instead rely on fewer dominant or functionally effective fungal partners.

1. Ruby-throated hummingbirds (Archilochus colubris) exhibit flower preferences and readily visit human subsidies like feeders. However, foraging behavior and plant-animal interactions may vary across spatial, temporal, and landscape gradients. Despite the popularity and ubiquity of Ruby-throated Hummingbirds in the eastern U.S., there has never been a quantitative assessment of their flower preferences or feeder use across broad scales. 2. We investigated how feeder visitation, flower visitation, and flower trait preferences vary by latitude, season, and land use in the northeastern United States using over 6 million occurrence records of Ruby-throated Hummingbirds and flowering plants, >2,100 annotated hummingbird-flower interactions, and >2,700 feeder visit occurrences. 3. We found that hummingbird feeder use declined over the year, increased with latitude, and was higher in developed landscapes. Flower visitation increased over the year across all latitudes, with higher visitation in developed landscapes. Finally, we found that native plant use diverged between landscapes, such that the probability of visiting a native flower increased over time in non-developed land uses but declined over time in developed ones, demonstrating that hummingbirds track the advancement of native floral phenology and use non-native, cultivated flowers as a human subsidy due to either availability or preferences. 4. Our preference and network models revealed that while hummingbird-plant network structure was similar across landscapes, the composition of important taxa shifted from native, wild species like Monarda and Impatiens to non-native, cultivated species like Salvia. 5. Using trait-based models of flower visitation, we found that hummingbirds preferred native, tubular, and red/orange flowers fitting the hummingbird pollination syndrome despite visiting >260 different plant species. Red and orange flowers were preferred across all seasons, suggesting color may be a reliable signal of nectar availability across species and contexts. Native and tubular flowers were strongly preferred during the breeding season; however, preferences relaxed during spring and fall migration. 6. These findings reveal the consistent preferences of Ruby-throated Hummingbirds for native, tubular, and red/orange flowers, and underscore how spatial and temporal factors reshape foraging behavior and trait preferences. Our results also highlight the value of community-collected data in characterizing plant-pollinator interactions across broad spatial and temporal scales.

AimTo test whether dispersal ability determines the relative importance of environmental filtering and distinct dispersal pathways in structuring stream macroinvertebrate metacommunities. LocationNzoia River catchment, Lake Victoria basin, western Kenya. TaxonBenthic macroinvertebrates (family level), grouped by dispersal ability into strong fliers, weak fliers, passive aerial dispersers, and obligate aquatics. MethodsMacroinvertebrates were sampled seasonally (2018 to 2021) at 55 sites. Using distance-based redundancy analysis and variation partitioning, we compared the influence of physicochemical water quality on community structure, representing environmental filtering, with the influence of three sets of spatial factors representing distinct dispersal pathways: Euclidean distance (overland dispersal pathway), land-use resistance (land-use mediated dispersal pathway), and Asymmetric Eigenvector Maps built on river-network connectivity (along-watercourse dispersal pathway). In addition, distance-decay relationships and turnover-nestedness partitioning were examined for the full assemblage and each dispersal group. ResultsCommunity assembly shifted predictably along the macroinvertebrate dispersal-ability gradient. Environmental filtering dominated in strong fliers and retained an independent influence in weak fliers and obligate aquatics, but contributed no independent influence in passive aerial dispersers. The overland dispersal pathway structured the three aerial groups. The land use mediated dispersal pathway constrained weak fliers and passive aerial dispersers, and the along-watercourse dispersal pathway structured passive aerial dispersers and obligate aquatics. Passive aerial dispersers showed the strongest dispersal signal, with the overland and along-watercourse pathways contributing equally, and in this group beta diversity reflected richness gradients (nestedness) rather than species replacement. Main conclusionsDispersal ability determined whether environmental filtering or a specific dispersal pathway governed community composition, and no single pathway captured assembly across the full range of dispersal abilities. Dispersal traits and connectivity must therefore be considered jointly when studying and managing aquatic biodiversity in human-modified Afrotropical catchments.

The charismatic migratory monarch butterfly population has declined dramatically, likely precipitated by loss of its breeding host plants (milkweed). Whether restoring milkweed would allow monarch recovery depends on whether additional factors currently limit the population. We investigated road mortality as one such factor. Monarchs cross thousands of roads during fall migration, and traffic volume has increased sharply while the population has plummeted. Using estimates of pre-migration distribution, flight patterns, and road traffic, we estimate that at least 61% (range 61% to 99.99%) of migrating monarchs are road-killed each fall. Although there is high uncertainty in our estimate, its magnitude suggests that roadkill could inhibit recovery of the population. Recovery planning should not only consider increasing the monarchs host plants, but must also address the reality of roadkill.