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This information correspond to our article: "Drivers of contrasting boreal understory vegetation in coniferous and broadleaf deciduous alternative states", published in Ecological Monographs (2023). Please find further methodological details in the article. Data in "Understory_veg_JR.csv" contain abundances of plant understory vegetation. The script for the analyses of this data correspond to "Github_AlternativeStates_2023.R" Data in "Env_Chem_JR.csv" contain environmental and physico-chemical properties in the study plots. The script for the analyses of this data correspond to "Github_Environmental_Data_Analyses.R", Article reference: Rodríguez-Rodríguez, J.C., Fenton, N., Kembel, SW., Mestre, E., Jean, M., and Y. Bergeron. 2023. Drivers of shifts in boreal understory vegetation between coniferous and broadleaf deciduous alternative state. Ecological monographs (accepted - in press). PhD thesis (complete) available at: https://depositum.uqat.ca/id/eprint/1426/

The soil microbiome plays major roles in the below-ground processes and productivity of forest ecosystems. Atmospheric nitrogen (N) deposition is predicted to increase globally and might create disturbances in soil microbial communities, essentially by modifying soil chemistry. However, the impacts of higher N deposition on the soil microbiome in N-limited northern forests are still unclear. For 16 years, we simulated N deposition by adding ammonium nitrate at rates of 3 and 10 times the ambient N deposition directly into soils located in three bioclimatic domains of the eastern Canadian forest (i.e., sugar maple–yellow birch, balsam fir–white birch, and black spruce–feather moss). We identified changes in the microbial communities by isolating the DNA of the L, F, and H soil horizons, as well as by sequencing amplicons of the bacterial 16S rRNA gene and the fungal ITS region. We found that long-term increased N deposition had no effect on soil microbial diversity, but had moderate impacts on the composition of the bacterial and fungal communities. The most noticeable change was the increase in ectomycorrhizal fungi ASV abundance, potentially due to increased tree root growth on fertilized plots. Our work suggests that, in N-limited northern forests, extra N is rapidly mobilized by vegetation, thus minimizing impacts on the soil microbiome.

Neonicotinoids, a class of systemic insecticides, have been widely used for decades against various insect pests. Previous studies have reported non-target effects of neonicotinoids on some beneficial macro- and micro-organisms. Considering the crucial role the soil microbiota plays in sustaining soil fertility, it is critical to understand how neonicotinoid exposure affects the microbial taxonomic composition and gene expression. However, most studies to date have evaluated soil microbial taxonomic compositions or assessed microbial functions based on soil biochemical analysis. In this study, we have applied a metatranscriptomic approach to quantify the variability in soil microbial gene expression in a 2 year soybean/corn crop rotation in Quebec, Canada. We identified weak and temporally inconsistent effects of neonicotinoid application on soil microbial gene expression, as well as a strong temporal variation in soil microbial gene expression among months and years. Neonicotinoid seed treatment altered the expression of a small number of microbial genes, including genes associated with heat shock proteins, regulatory functions, metabolic processes and DNA repair. These changes in gene expression varied during the growing season and between years. Overall, the composition of soil microbial expressed genes seems to be more resilient and less affected by neonicotinoid application than soil microbial taxonomic composition. Our study is among the first to document the effects of neonicotinoid seed treatment on microbial gene expression and highlights the strong temporal variability of soil microbial gene expression and its responses to neonicotinoid seed treatments.

The discovery of major axes of correlated functional variation among species and habitats has revealed the fundamental trade-offs structuring both functional and taxonomic diversity in eukaryotes such as plants. Whether such functional axes exist in the bacterial realm and whether they could explain bacterial taxonomic turnover among ecosystems remains unknown. Here we use a data-driven approach to leverage global genomic and metagenomic datasets to reveal the existence of major axes of functional variation explaining both evolutionary differentiation within Bacteria and their ecological sorting across diverse habitats. We show that metagenomic variation among bacterial communities from various ecosystems is structured along a few axes of correlated functional pathways. Similar clusters of traits explained phylogenetic trait variation among >16,000 bacterial genomes, suggesting that functional turnover among bacterial communities from distinct habitats does not only result from the differential filtering of similar functions among communities, but also from phylogenetic correlations among these functions. Concordantly, functional pathways associated with trait clusters that were most important for defining functional turnover among bacterial communities were also those that had the highest phylogenetic signal in the bacterial genomic phylogeny. This study overall underlines the important role of evolutionary history in shaping contemporary distributions of bacteria across ecosystems.Originality-Significance StatementIn this article, we use a trait screening approach based on genomic and metagenomic data to identify the key functional strategies of bacteria across ecosystems but also across the bacterial tree of life. This novel approach allows us to quantify the role of evolutionary processes in structuring microbial ecological differences among ecosystems. By reducing the high-dimensionality of trait variation observed among microorganisms around a small number of fundamental axes of trait covariation, we make a significant step towards generalization of the drivers of biological diversity in microbes but also across study systems. This research provide a major advance in our understanding of the origin and maintenance of bacterial biological diversity, expanding on related findings for plants and animals.

Phyllosphere microbial communities have received considerable attention given their important influence on their plant hosts and on ecosystem functioning. In a context where climate change threatens the sustainability of ecosystems, it is important to understand how phyllosphere microbes will respond to changes in their environment. We used 16S rRNA gene amplicon sequencing to quantify phyllosphere bacterial communities of black spruce (Picea mariana (Mill.) B.S.P.) exposed to nitrogen canopy enrichment and soil warming in the boreal forest of Quebec, Canada. The treatments were applied from April to September 2015 and the sampling was done in September. Neither treatment influenced the overall community structure and diversity of black spruce phyllosphere bacterial communities. However, some bacterial taxa and inferred microbial functions did differ among treatments, revealing in particular a stronger response of some bacteria to soil warming rather than nitrogen enrichment. Our results suggest that soil warming could potentially induce more changes in phyllosphere bacterial taxa abundances and functions than could nitrogen addition, with potential consequences for microbial diversity and boreal forest ecosystem function under likely climate change scenarios. Our study suggests avenues for further research to integrate a more mechanistic understanding of the importance of phyllosphere microbes for black spruce and boreal forest ecosystems.

Purpose: Natural and anthropogenic causes have produced changes in tree dominance from coniferous to broadleaf deciduous forests, generating shifts in litter inputs and plant understory composition. The impact of changes in canopy-associated factors on belowground microbial communities remain poorly understood. The objective of this study was to better understand how abiotic and biotic factors in black spruce and trembling aspen forests shape soil microbial community structure. Methods: With high throughput sequencing, we first analyzed differences in microbial communities between microhabitats (tree phyllosphere vs. soil microbiome) and forest types (black spruce vs. trembling aspen). Second, we analyzed how shifts in factors related to each forest type (litter deposition and understory vegetation) affected soil microbial community composition. Results: We found a high microhabitat specificity of bacterial communities interacting with forest type. Shifts in litter deposition and understory vegetation between forest types did not influence microbial community composition, but the legacy effects of each forest type defined soil bacterial and fungal communities. Fungal community composition was more strongly influenced by forest type compared with bacterial communities, and both were correlated with several soil physicochemical properties that differed among forest types. Conclusion: This study expands our knowledge of the microbial composition of tree phyllosphere and soil microbial communities in black spruce and trembling aspen forests and their correlation with abiotic and biotic factors in each forest type. Our study demonstrates the resistance of microorganisms to variation in canopy-related factors and the importance of legacy effects of forest type in defining soil microbial community composition.

Methylobacterium is a group of methylotrophic microbes associated with soil, fresh water, and particularly the phyllosphere, the aerial part of plants that has been well-studied in terms of physiology but whose evolutionary history and taxonomy are unclear. Recent work has suggested that Methylobacterium is much more diverse than thought previously, questioning its status as an ecologically and phylogenetically coherent taxonomic genus. However, taxonomic and evolutionary studies of Methylobacterium have mostly been restricted to model species, often isolated from habitats other than the phyllosphere, and have yet to utilize comprehensive phylogenomic methods to examine gene trees, gene content, or synteny. By analyzing 189 Methylobacterium genomes from a wide range of habitats, including the phyllosphere, we inferred a robust phylogenetic tree while explicitly accounting for the impact of horizontal gene transfers. We showed that Methylobacterium contains four evolutionary distinct groups of bacteria (namely A, B, C, D), characterized by different genome size, GC content, gene content and genome architecture, revealing the dynamic nature of Methylobacterium genomes. In addition of recovering 59 described species, we identified 45 candidate species, mostly phyllosphere-associated, stressing the significance of plants as a reservoir of Methylobacterium diversity. We inferred an ancient transition from a free-living lifestyle to association with plant roots in Methylobacteriaceae ancestor, followed by phyllosphere association of three of the major groups (A, B, D), which early branching in Methylobacterium history was heavily obscured by HGT. Together, our work lays the foundations for a thorough redefinition of Methylobacterium taxonomy, beginning with the abandon of Methylorubrum.

Plant-associated microbes are essential for promoting plant well-being, maintaining biodiversity, and supporting ecosystem function. However, little is known about the geographic distribution of plant-microbe symbioses and how they are formed and change along latitudinal gradients. Here we identified leaf bacteria for 328 plant species sampled from 10 forests along a tropical to temperate gradient in China. We analyzed the diversity and composition of plant leaf-associated bacteria and quantified the contributions of hosts, habitats, and neighborhood plants to the plant-bacterial symbiosis. We found a strong latitudinal gradient in leaf bacterial diversity and composition. Bacterial assemblages on leaves were most strongly selected by host plants, and the selection pressure increased with latitude. In contrast, at low latitudes and at large geographical scales multiple factors were found to jointly regulate bacterial community composition. Our result also showed that plant-bacteria symbiotic networks were structured by network hub bacteria taxa with high co-occurrence network centrality, and the abundance of temperate hub taxa was more influenced by host plants than that in tropical forests. For the first time, we documented a previously unrecognized latitudinal gradient in plant-bacterial symbioses that was regulated by a joint effect of multiple factors at low latitudes but mostly by host selection at high latitudes, implying that leaf microbiomes are likely to respond differently to global change along the latitudinal gradient.

Climate change is prompting plants to migrate and establish novel interactions in new habitats. Because of the pivotal roles that microbes have on plant health and function, it is important to understand the ecological consequences of these shifts in host-microbe interactions with range expansion. Here we examine how the diversity of plant-associated microbes varies along the host's current range and extended range according with climate change predictions, and assess the relative influence of host genotype (seed provenance) and environment in structuring the host microbiome. We collected sugar maple seeds from across the species current range, then planted them in temperate and mixedwood/transitional forests (current range) and in the boreal region (beyond range but predicted future range in response to climate change). We used amplicon sequencing to quantify bacterial, fungal, and mycorrhizal communities from seedling leaves and roots. Variation among sites and regions were the main drivers of the differences in host microbial communities, whereas seed provenance did not play a large role. No unifying pattern was observed for microbial community richness, diversity, or specialization, demonstrating the complexity of responses of different taxa on above- and belowground plant compartments. Along the latitudinal gradient, we (1) observed reductions in mycorrhizal diversity that can negatively impact maple establishment; (2) and revealed reductions in fungal leaf pathogens that can have opposite effects. Our results highlight the need for an integrated approach including the examination of various microbial taxa on different plant compartments to improve our understanding of plant range shifts and plant-microbe interactions.

Neonicotinoids, a class of systemic insecticides, have been widely used for decades against various insect pests. Past studies have reported non-target effects of neonicotinoids on some beneficial macro- and micro-organisms. Given the crucial role that the soil microbiota plays in sustaining soil fertility, it is critical to understand how microbial taxonomic composition and gene expression respond to neonicotinoid exposure. To date, few studies have focused on this question, and these studies have evaluated the shifts in soil microbial taxonomic composition or used soil biochemical analyses to assess the changes in microbial functions. In this study, we have applied a metatranscriptomic approach to quantify the variability in soil microbial gene expression in a two-year soybean/corn crop rotation in Quebec, Canada. We identified weak and temporally inconsistent effects of neonicotinoid application on soil microbial gene expression, as well as a strong temporal variation in soil microbial gene expression among months and years. Neonicotinoid seed treatment altered the expression of a small number of microbial genes, including genes associated with heat shock proteins, regulatory functions, metabolic processes and DNA repair. These changes in gene expression varied during the growing season and between years. Overall, the composition of soil microbial expressed genes seems to be more resilient and less affected by neonicotinoid application than soil microbial taxonomic composition. Our study is among the first to document the effects of neonicotinoid seed treatment on microbial gene expression and highlights the strong temporal variability of soil microbial gene expression and its responses to neonicotinoid seed treatments.IMPORTANCEThis work provides the first example of the impacts of neonicotinoid seed treatment on community-wide soil microbial gene expression in an experimental design representing real farming conditions. Neonicotinoid pesticides have attracted a great deal of attention in recent years due to their potential non-target impacts on ecological communities and their functions. Our paper represents the first use of metatranscriptomic sequencing to offer real-time and in-depth insights into the non-target effects of this pesticide on soil microbial gene expression and on potentially beneficial soil microorganisms.

Methylobacterium is a prevalent bacterial genus of the phyllosphere. Despite its ubiquity, little is known about the extent to which its diversity reflects neutral processes like migration and drift, versus environmental filtering of life history strategies and adaptations. In two temperate forests, we investigated how phylogenetic diversity within Methylobacterium was structured by biogeography, seasonality, and growth strategies. Using deep, culture-independent barcoded marker gene sequencing coupled with culture-based approaches, we uncovered a considerable diversity of Methylobacterium in the phyllosphere. We cultured different subsets of Methylobacterium lineages depending upon the temperature of isolation and growth (20 °C or 30 °C), suggesting long-term adaptation to temperature. To a lesser extent than temperature adaptation, Methylobacterium diversity was also structured across large (>100km; between forests) and small geographical scales (Methylobacterium is phylogenetically structured into lineages with distinct growth strategies, which helps explain their differential abundance across regions, host tree species, and time. This works paves the way for further investigation of adaptive strategies and traits within a ubiquitous phyllosphere genus.ImportanceMethylobacterium is a bacterial group tied to plants. Despite its ubiquity and importance to their hosts, little is known about the processes driving Methylobacterium community dynamics. By combining traditional culture-dependent and independent (metabarcoding) approaches, we monitored Methylobacterium diversity in two temperate forests over a growing season. On the surface of tree leaves, we discovered remarkably diverse and dynamic Methylobacterium communities over short temporal (from June to October) and spatial scales (within 1.2 km). Because we cultured different subsets of Methylobacterium diversity depending on the temperature of incubation, we suspected that these dynamics partly reflected climatic adaptation. By culturing strains in lab conditions mimicking seasonal variations, we found that diversity and environmental variations were indeed good predictors of Methylobacterium growth performances. Our findings suggest that Methylobacterium community dynamics at the surface of tree leaves results from the succession of strains with contrasted growth strategies in response to environmental variations.

Litter decomposition in forests is affected by tree species composition which produces litters with specific characteristics and alters soil surface conditions and nutrient availability. The impact of such changes in soil microsite conditions on litter decomposition is however poorly elucidated. We studied the decomposition of sugar maple leaves in 22 plots varying from pure hardwoods to mixedwoods to conifer-dominated stands across four sites in the sugar maple bioclimatic domain in Quebec. Mass loss and nutrient release were assessed over a 2-year experiment. Proxies of moisture availability measured under the canopy were more valuable for explaining the variability in leaf litter decomposition rates than soil temperature, litter quality and general site measurements (e.g. air temperature, rainwater). Throughfall amounts and soil volumetric water content (VWC) explained 56% of the variability in mass loss constants. Sugar maple leaf litter under hardwoods also decomposed more rapidly than under conifer-dominated stands. Slower litter decomposition under conifer-dominated stands was again more largely controlled by the lower soil VWC than the more acidic forest floor created by the needles. This study highlights that proxies of moisture availability measured at the microsite level can be key variables to predict litter dynamics in forests.

Functional diversity holds the promise of understanding ecosystems in ways unattainable by taxonomic diversity studies. Underlying this promise is the intuition that investigating the diversity of what organisms actually do (i.e., their functional traits) within ecosystems will generate more reliable insights into the ways these ecosystems behave, compared to considering only species diversity. But this promise also rests on several conceptual and methodological (i.e., epistemic) assumptions that cut across various theories and domains of ecology. These assumptions should be clearly addressed, notably for the sake of an effective comparison and integration across domains, and for assessing whether or not to use functional diversity approaches for developing ecological management strategies. The objective of this contribution is to identify and critically analyze the most salient of these assumptions. To this aim, we provide an epistemic roadmap that pinpoints these assumptions along a set of historical, conceptual, empirical, theoretical, and normative dimensions.

Bacterial endophytes may be fairly host‐specific; nonetheless, an important subset of taxa may be shared among numerous host species forming a community‐wide core microbiome. Moreover, other key factors, particularly the supply of limiting macronutrients and disturbances, may supersede the importance of host identity. We tested the following four non‐mutually exclusive hypotheses: (a) The Host Identity Hypothesis: endophytes vary substantially among different host‐plant species. (b) The Core Microbiome Hypothesis: a subset of microbial taxa will be shared among all host‐plant species. (c). The Soil Resource Supply Hypothesis: endophytes vary substantially among habitats with experimentally elevated levels of macronutrients. (d) The Disturbance–Disruption Hypothesis: disturbances created by the periodic application of antibiotics structure bacterial endophyte communities. We tested these hypotheses by characterizing endophytes using high‐throughput sequencing among seedlings of five phylogenetically diverse tree species nested within a long‐term, full factorial nitrogen, phosphorus and potassium soil fertilization experiment. We artificially disturbed one of our focal species by applying antibiotics every 10–14 days for 29 months within the soil (N, P, K) fertilization experiment. While we detected a significant effect of host identity and soil nutrient additions, together they explained little variation in endophyte community composition (

Based on the bilateral relationship between the gut microbiota and formation/function of the enteric nervous system (ENS), we sought to determine whether antibiotics-induced dysbiosis might impact the expressivity of genetically-induced ENS abnormalities. To address this, we took advantage of the TashT mouse model of Hirschsprung disease, in which colonic aganglionosis and hypoganglionosis are both much more severe in males. These defects result into two male-biased colon motility phenotypes: either megacolon that is lethal around weaning age or chronic constipation in adults, the latter being also associated with an increased proportion of nitrergic neurons in the distal ENS. Induction of dysbiosis using a cocktail of broad-spectrum antibiotics specifically impacted the colonic ENS of TashTTg/Tg mice in a stage-dependent manner. It further decreased the neuronal density at post-weaning age and differentially modulated the otherwise increased proportion of nitrergic neurons, which appeared normalized around weaning age and further increased at post-weaning age. These changes delayed the development of megacolon around weaning age but led to premature onset of severe constipation later on. Finally, local inhibition of nitric oxide signaling improved motility and prevented death by megacolon. We thus conclude that exposure to antibiotics can negatively influence the expressivity of a genetically-induced enteric neuropathy.

Trophic networks are composed of many organisms hosting microbiota that interact with their hosts and with each other. Yet, our knowledge of the factors driving variation in microbiota and their interactions in wild communities is limited. To investigate the relation among host microbiota across a trophic network, we studied the bacterial microbiota of two species of primary producers (downy and holm oaks), a primary consumer (caterpillars), and a secondary consumer (blue tits) at nine sites in Corsica. To quantify bacterial microbiota, we amplified 16S rRNA gene sequences in blue tit feces, caterpillars, and leaf samples. Our results showed that hosts from adjacent trophic levels had a more similar bacterial microbiota than hosts separated by two trophic levels. Our results also revealed a difference between bacterial microbiota present on the two oak species, and among leaves from different sites. The main drivers of bacterial microbiota variation within each trophic level differed across spatial scales, and sharing the same tree or nest box increased similarity in bacterial microbiota for caterpillars and blue tits. This study quantifies host microbiota interactions across a three-level trophic network and illustrates how the factors shaping bacterial microbiota composition vary among different hosts.

Cover cropping is a soil conservation practice that may reduce the impacts of the economically important pathogen Pseudomonas syringae on crops including squash (Cucurbita pepo). To date, no studies have directly quantified the effect of rye cover crops on P. syringae populations, nor on the bacterial community of squash leaves. In this work, we tested the hypothesis that the protective effects of cover cropping on squash may be mediated by cover cropping effects on the plant’s microbiota that in turn protects against P. syringae. Using combined 16S sequencing and culture-based approaches, we showed that rye cover cropping protects squash against P. syringae, by decreasing pathogen population size on squash leaves and increasing fruit health and marketability at harvest. We also found evidence of a strong effect of rye cover crops on bacterial communities of the squash phyllosphere. Those findings were more striking early in the growing season. Finally, we identified numerous phyllosphere bacteria belonging to the genera Sphingomonas, Methylobacterium and Pseudomonas that were promoted by rye cover crops. Overall, our findings suggest cover cropping is effective for the sustainable management of P. syringae on squash and may provide a reservoir of potential microbial biocontrol agents colonizing the phyllosphere.

Phyllosphere bacterial diversity is shaped through interactions between hosts and microbes. Most studies having focused on pairwise associations between host taxa and their symbionts, little is yet understood about the influence of the host community as a whole in shaping these interactions. Envisioning phyllosphere bacterial communities as a spatially structured network of communities linked by dispersal (i.e., metacommunities) can help us better understand the relative importance of species sorting among host populations and species versus dispersal from the neighboring host community for bacterial community assembly in forest ecosystems. Here we investigate drivers of metacommunity structure of epiphytic bacteria of the phyllosphere among 33 tree host species distributed across a large‐scale transition from deciduous to boreal forest. We expect the identity and traits of hosts to play an important role in determining phyllosphere bacterial composition. We further hypothesize that bacterial dispersal from neighboring host species will modulate the match between a focal host species and its microbiota, and shape opportunities for host specialization of phyllosphere bacteria at local and regional scales. We defined specialization as the level of phylogenetic similarity among hosts that a bacterial symbiont associates with. We found that host taxonomic identity and traits were important drivers of bacterial community turnover and variation in host specialization across the landscape. Dispersal from neighboring communities further played a role in homogenizing bacterial communities. The microbiota of focal hosts such as sugar maple was thus increasingly similar to that of neighboring host species along the transition from deciduous to boreal forest. Specialization of bacterial taxa on sugar maple was further positively correlated with the relative abundance of this host in the landscape, revealing a role for the host community context in shaping evolutionary relationships between phyllosphere bacteria and their tree hosts. These results overall suggest that the dispersal of phyllosphere bacteria from the dominant tree community members may be constraining the match between tree species and their symbionts, particularly at their range limits. We also demonstrate that considering host‐associated microbial communities as part of metacommunities within the host landscape is a promising tool for improving our understanding of host‐symbiont matching.

BackgroundThe phyllosphere and soil are dynamic habitats for microbial communities. Non-pathogenic microbiota, including leaf and soil beneficial bacteria, plays a crucial role in plant growth and health, as well as in soil fertility and organic matter production. In sustainable agriculture, it is important to understand the composition of these bacterial communities, their changes in response to disturbances, and their resilience to agricultural practices. Widespread pesticide application may have had non-target impacts on these beneficial microorganisms. Neonicotinoids are a family of systemic insecticides being vastly used to control soil and foliar pests in recent decades. A few studies have demonstrated the long-term and non-target effects of neonicotinoids on agroecosystem microbiota, but the generality of these findings remains unclear. In this study, we used 16S rRNA gene amplicon sequencing to characterize the effects of neonicotinoid seed treatment on soil and phyllosphere bacterial community diversity, composition and temporal dynamics in a three-year soybean/corn rotation in Quebec, Canada. Results We found that habitat, host species and time are stronger drivers of variation in bacterial composition than neonicotinoid application. They respectively explained 37.3%, 3.2% and 2.9% of the community variation. However, neonicotinoids did have an impact on bacterial community structure, especially on the taxonomic composition of soil communities (2.6%) and over time (2.4%). They also caused a decrease in soil alpha diversity in the middle of the growing season. While the neonicotinoid treatment favored some bacterial genera known as neonicotinoid biodegraders, there was a decline in the relative abundance of some potentially beneficial soil bacteria, such as the plant growth-promoting rhizobacteria and the bacteria involved in the nitrogen cycle, in response to the pesticide application.ConclusionsOur results indicate that neonicotinoids have non-target effects on phyllosphere and soil bacterial communities in a soybean-corn agroecosystem, especially potentially beneficial bacteria that are vital for plant growth and improve soil fertility. Exploring the interactions among bacteria and other organisms, as well as the bacterial functional responses to the pesticide treatment, may enhance our understanding of these non-target effects and help us adapt agricultural practices to control these impacts.

Metacommunity theory predicts that strongly connected individuals will harbour similar gut microbiomes (GMs) and affiliating with more individuals should increase GM diversity. Additionally, cross-species bacterial transmission may play a role in how interspecific interactions affect host community dynamics. We tracked sympatric mice (Peromyscus maniculatus) and voles (Myodes gapperi) and constructed social networks for each species and both species together. We tested whether: 1) similarity in GM composition between individuals correlates with their social proximity within and across species; 2) GM diversity correlates with a host’s number of conspecific or heterospecific neighbours. We could not differentiate associations between GM composition and mouse social proximity or habitat. In voles, social proximity explained part of the GM composition. GM composition associated with interspecific social proximity, and mouse GM diversity correlated with number of vole neighbours. Contributions of host-host bacterial transmission to the GM partly follow metacommunity theory but depend on host species.

Background The phyllosphere is an important microbial habitat, but our understanding of how plant hosts drive the composition of their associated leaf microbial communities and whether taxonomic associations between plants and phyllosphere microbes represent adaptive matching remains limited. In this study, we quantify bacterial functional diversity in the phyllosphere of 17 tree species in a diverse neotropical forest using metagenomic shotgun sequencing. We ask how hosts drive the functional composition of phyllosphere communities and their turnover across tree species, using host functional traits and phylogeny. Results Neotropical tree phyllosphere communities are dominated by functions related to the metabolism of carbohydrates, amino acids, and energy acquisition, along with environmental signalling pathways involved in membrane transport. While most functional variation was observed within communities, there is non-random assembly of microbial functions across host species possessing different leaf traits. Metabolic functions related to biosynthesis and degradation of secondary compounds, along with signal transduction and cell–cell adhesion, were particularly important in driving the match between microbial functions and host traits. These microbial functions were also evolutionarily conserved across the host phylogeny. Conclusions Functional profiling based on metagenomic shotgun sequencing offers evidence for the presence of a core functional microbiota across phyllosphere communities of neotropical trees. While functional turnover across phyllosphere communities is relatively small, the association between microbial functions and leaf trait gradients among host species supports a significant role for plant hosts as selective filters on phyllosphere community assembly. This interpretation is supported by the presence of phylogenetic signal for the microbial traits driving inter-community variation across the host phylogeny. Taken together, our results suggest that there is adaptive matching between phyllosphere microbes and their plant hosts.

Background Aims Hirschsprung disease (HSCR) is a life-threatening birth defect in which the distal colon is devoid of enteric neural ganglia. HSCR is treated by surgical removal of aganglionic bowel, but many children continue to have severe problems after surgery. We studied whether administration of glial cell derived neurotrophic factor (GDNF) induces enteric nervous system regeneration in mouse models of HSCR. Methods We performed studies with four mouse models of HSCR: Holstein (HolTg/Tg, a model for trisomy 21-associated HSCR), TashT (TashTTg/Tg, a model for male-biased HSCR), Piebald-lethal (Ednrbs-l//s-l, a model for EDNRB mutation-associated HSCR), and Ret9/- (with aganglionosis induced by mycophenolate). Mice were given rectal enemas containing GDNF or saline (control) from postnatal days 4 through 8. We measured survival times of mice, and colon tissues were analyzed by histology, immunofluorescence, and immunoblots. Neural ganglia regeneration and structure, bowel motility, epithelial permeability, muscle thickness, and neutrophil infiltration were studied in colon tissues and in mice. Stool samples were collected, and microbiomes were analyzed by 16S rRNA gene sequencing. Time-lapse imaging and genetic cell-lineage tracing were used to identify a source of GDNF-targeted neural progenitors. Human aganglionic colon explants from children with HSCR were cultured with GDNF and evaluated for neurogenesis. Results GDNF significantly prolonged mean survival times of HolTg/Tg mice, Ednrbs-l//s-l mice, and male TashTTg/Tg mice, compared with control mice, but not Ret9/- mice (which had mycophenolate toxicity). Mice given GDNF developed neurons and glia in distal bowel tissues that were aganglionic in control mice, had a significant increase in colon motility, and had significant decreases in epithelial permeability, muscle thickness, and neutrophil density. We observed dysbiosis in fecal samples from HolTg/Tg mice compared with feces from wild-type mice; fecal microbiomes of mice given GDNF were similar to those of wild-type mice except for Bacteroides. Exogenous luminal GDNF penetrated aganglionic colon epithelium of HolTg/Tg mice, inducing production of endogenous GDNF, and new enteric neurons and glia appeared to arise from Schwann cells within extrinsic nerves. GDNF application to cultured explants of human aganglionic bowel induced proliferation of Schwann cells and formation of new neurons. Conclusions GDNF prolonged survival, induced enteric neurogenesis, and improved colon structure and function in 3 mouse models of HSCR. Application of GDNF to cultured explants of aganglionic bowel from children with HSCR induced proliferation of Schwann cells and formation of new neurons. GDNF might be developed for treatment of HSCR.

Climate change is causing rapid shifts in species’ range limits, leading to poleward expansions and range losses toward the equator. However, ‘climate debt’, the gap between required and realized range shifts under changing climates, can accumulate when species are unable to track shifting conditions sufficiently rapidly to keep pace with climate changes. Currently, we do not know the rate at which species will keep pace via dispersal to track their climate envelopes, yet understanding potential differences in climate debt is central to estimating how climate change will influence extinction risk. Here, we use historical observations of 155 butterfly species found in Canada to construct climate‐based environmental niche models for each species and then compare projections with observed modern distributions to quantify climate debts. This approach suggests that high levels of climate debt are accumulating within the vast majority of these species. Such failure to track changing climates may arise from some combination of interspecific interactions such as particular food availability for specialists, abiotic barriers such as mountain ranges, or species’ intrinsic dispersal capacities. Our linear models relating climate debt to a variety of biological predictors suggest that the debts we documented are accumulating independently of dispersal ability, diet breadth, and phylogeny. A proxy for range size is the only significant predictor of climate debt, with species with narrower ranges accumulating more debt: this suggests that species with narrow ranges may be at risk from both a reduction of suitable habitat in their current range and the failure to colonize newly available habitat. Identifying the factors, whether intrinsic or imposed by local environmental conditions, that limit species’ capacities to colonize areas beyond their historical limits is vital to conservation planning.

BackgroundThe phyllosphere is an important microbial habitat but our understanding of how plant hosts drive the composition of their associated leaf microbial communities and whether taxonomic associations between plants and phyllosphere microbes represent adaptive matching remains limited. In this study we quantify bacterial functional diversity in the phyllosphere of 17 tree species in a diverse neotropical forest using metagenomic shotgun sequencing. We ask how hosts drive the functional composition of phyllosphere communities and their turnover across tree species, using host functional traits and phylogeny. We compare functional predictions inferred from 16S gene sequencing with functions estimated from metagenomic shotgun sequencing.ResultsNeotropical tree phyllosphere communities are dominated by functions related to the metabolism of carbohydrates, amino acids and energy acquisition, along with environmental signalling pathways involved in membrane transport. While most functional variation was observed within communities, there is non-random assembly of microbial functions across host species possessing different leaf traits. Metabolic functions related to biosynthesis and degradation of secondary compounds, along with signal transduction and cell-cell adhesion were particularly important in driving the match between microbial functions and host traits. These microbial functions were also evolutionarily conserved across the host phylogeny. Functional predictions inferred from 16S gene sequences were weakly correlated with functional annotations from the same samples through metagenomic shotgun sequencing, especially for finer-scale functional annotations.ConclusionsFunctional profiling based on metagenomic shotgun sequencing offers evidence for the presence of a core functional microbiome across phyllosphere communities of neotropical trees. While functional turnover across phyllosphere communities is relatively small, the association between microbial functions and leaf trait gradients among host species supports a significant role for plant hosts as selective filters on phyllosphere community assembly. This interpretation is supported by the presence of phylogenetic signal for the microbial traits driving inter-community variation across the host phylogeny. Our comparison of functional annotations derived from 16S genes versus metagenomic shotgun sequencing suggests caution in using functions inferred from 16S genes for studying ecological dynamics in phyllosphere communities. Taken together, our results suggest that there is adaptive matching between phyllosphere microbes and their plant hosts.

Growing pressures linked to global warming are prompting governments to put policies in place to find alternatives to fossil fuels. In this study, we compared the impact of tree-length harvesting to more intensive full-tree harvesting on the composition of fungi residing in residual stumps 5 years after harvest. In the tree-length treatment, a larger amount of residual material was left around the residual stumps in contrast to the full-tree treatment where a large amount of woody debris was removed. We collected sawdust from five randomly selected residual stumps in five blocks in each of the tree-length and full-tree treatments, yielding a total of 50 samples (25 in each treatment). We characterized the fungal operational taxonomic units (OTUs) present in each stump using high-throughput DNA sequencing of the fungal ITS region. We observed no differences in Shannon diversity between tree-length and full-tree harvesting. Likewise, we observed few differences in the composition of fungal OTUs among tree-length and full-tree samples using non-metric multidimensional scaling. Using the differential abundance analysis implemented with DESeq2, we did, however, detect several associations between specific fungal taxa and the intensity of residual biomass harvest. For example, Peniophorella pallida (Bres.) KH Larss. and Tephromela sp. were found mainly in the full-tree treatment, while Phlebia livida (Pers.) Bres. and Cladophialophora chaetospira (Grove) Crous & Arzanlou were found mainly in the tree-length treatment. While none of the 20 most abundant species in our study were identified as pathogens we did identify one conifer pathogen species Serpula himantioides (Fr.) P. Karst found mainly in the full-tree treatment.

There is an increasing interest in applying trait-based approaches to microbial ecology, but the question of how and why to do it is still lagging behind. By anchoring our discussion of these questions in a framework derived from epistemology, we broaden the scope of trait-based approaches to microbial ecology from one oriented mostly around explanation towards one inclusive of the predictive and integrative potential of these approaches. We use case studies from macro-organismal ecology to concretely show how these goals for knowledge development can be fulfilled and propose clear directions, adapted to the biological reality of microbes, to make the most of recent advancements in the measurement of microbial phenotypes and traits.

Human activities can alter aquatic ecosystems through the input of nutrients and carbon, but there is increasing evidence that these pressures induce nonlinear ecological responses. Nonlinear relationships can contain breakpoints where there is an unexpected change in an ecological response to an environmental driver, which may result in ecological regime shifts. We investigated the occurrence of nonlinearity and breakpoints in relationships between total dissolved nitrogen (TDN), total dissolved phosphorus (TDP), and total dissolved carbon (DOC) concentrations and ecological responses in streams with varying land uses. We calculated breakpoints using piecewise regression, two dimensional Kolmogorov-Smirnov (2DKS), and significant zero crossings (SiZer) methods. We found nonlinearity was common, occurring in half of all analyses, with some evidence of multiple breakpoints. Linearity, by contrast, occurred in less than 14% of cases, on average. Breakpoints were related to land use gradients, with 34–43% agricultural cover associated with DOC and TDN breakpoints, and 15% wetland and 9.5% urban land associated with DOC and nutrient breakpoints, respectively. While these breakpoints are likely specific to our study area, our study contributes to the growing literature of the prevalence and location of ecological breakpoints in streams, providing watershed managers potential criteria for catchment land use thresholds.

Research on biodiversity and ecosystem functioning has demonstrated links between plant diversity and ecosystem functions such as productivity. At other trophic levels, the plant microbiome has been shown to influence host plant fitness and function, and host-associated microbes have been proposed to influence ecosystem function through their role in defining the extended phenotype of host organisms However, the importance of the plant microbiome for ecosystem function has not been quantified in the context of the known importance of plant diversity and traits. Here, using a tree biodiversity-ecosystem functioning experiment, we provide strong support for the hypothesis that leaf bacterial diversity is positively linked to ecosystem productivity, even after accounting for the role of plant diversity. Our results also show that host species identity, functional identity and functional diversity are the main determinants of leaf bacterial community structure and diversity. Our study provides evidence of a positive correlation between plant-associated microbial diversity and terrestrial ecosystem productivity, and a new mechanism by which models of biodiversity-ecosystem functioning relationships can be improved.

The factors that govern assembly of the gut microbiota are insufficiently understood. Here, we test the hypothesis that inter-individual microbiota variation can arise solely from differences in the order and timing by which the gut is colonized early in life. Experiments in which mice were inoculated in sequence either with two complex seed communities or a cocktail of four bacterial strains and a seed community revealed that colonization order influenced both the outcome of community assembly and the ecological success of individual colonizers. Historical contingency and priority effects also occurred in Rag1-/- mice, suggesting that the adaptive immune system is not a major contributor to these processes. In conclusion, this study established a measurable effect of colonization history on gut microbiota assembly in a model in which host and environmental factors were strictly controlled, illuminating a potential cause for the high levels of unexplained individuality in host-associated microbial communities.

15 pages, 6 figures, 3 tables, supplementary data https://doi.org/10.1093/femsec/fiy115, Freshwater bacterioplankton communities are influenced by the inputs of material and bacteria from the surrounding landscape, yet few studies have investigated how different terrestrial inputs affect bacterioplankton. We examined whether the addition of soils collected under various tree species combinations differentially influences lake bacterial communities. Lake water was incubated for 6 days following addition of five different soils. We assessed the taxonomic composition (16S rRNA gene sequencing) and metabolic activity (Biolog Ecoplates) of lake bacteria with and without soil addition, and compared these to initial soil communities. Soil bacterial assemblages showed a strong influence of tree composition, but such community differences were not reflected in the structure of lake communities that developed during the experiment. Bacterial taxa showing the largest abundance increases during incubation were initially present in both lake water and across most soils, and were related to Cytophagales, Burkholderiales and Rhizobiales. No clear metabolic profiles based on inoculum source were found, yet soil-amended communities used 60% more substrate than non-inoculated communities. Overall, we show that terrestrial inputs influence aquatic communities by stimulating the growth and activity of certain ubiquitous taxa distributed across the terrestrial-aquatic continuum, yet different forest soils did not cause predictable changes in lake bacterioplankton assemblages, This work was supported by the Natural Science and Engineering Research Council of Canada (NSERC) and Hydro-Quebec, as well as by the Fons de Reserche Nature et Technologies (FRQNT) and the Canada Research Chairs program

Microbial communities have been shown to play an important role for host health in mammals, especially humans. It is thought that microbes could play an equally important role in other animal hosts such as insects. A growing body of evidence seems to support this, however most of the research effort in understanding host-microbe interactions in insects has been focused on a few well-studied groups such as bees, cockroaches and termites. We studied the effects of the gut-associated microbial community on the growth and survival of the eastern spruce budworm Choristoneura fumiferana, an economically important lepidopteran forest pest in eastern Canada and the northeastern United States. Contrary to our expectations, the gut microbial community of spruce budworm larvae does not appear to influence host growth or survival. Our results agree with the hypothesis that lepidopteran larvae lack resident microbial communities and are not nutritionally dependent on bacterial symbionts.