Leaf soluble sugars and starch are important components of nonstructural carbohydrates (NSCs), which are crucial for plant growth, development, and reproduction. Although there is a large body of research focusing on the regulation of plant NSC (soluble sugars and starch) concentrations, the response of foliar NSC concentrations to continuous nitrogen (N) and phosphorus (P) addition is still unclear, especially in tropical forests. Here, we used a long-term manipulative field experiment to investigate the response of leaf NSC concentrations to continuous N and P addition (3-, 5-, and 8-year fertilization) in a tropical forest in southern China. We found significant species-specific variation in leaf NSC concentrations in this tropical forest. Phosphorus addition dramatically decreased both leaf soluble sugar and starch concentrations, while N addition had no significant effects on leaf soluble sugar and starch concentrations. These results suggest that, in plants growing in P-limiting tropical soil, leaf NSC concentrations are regulated by soil P availability rather than N availability. Moreover, the negative relationships between NSC concentrations and leaf mass per area (LMA) revealed that NSCs could supply excess carbon (C) for leaf expansion under P addition. This was further supported by the increased structural P fraction after P fertilization in our previous study at the same site. We conclude that soil P availability strongly regulates leaf starch and soluble sugar concentrations in the tropical tree species included in this study. The response of leaf NSC concentrations to long-term N and P addition can reflect the close relationships between plant C dynamics and soil nutrient availability in tropical forests. Maintaining relatively higher leaf NSC concentrations in tropical plants can be a potential mechanism for adapting to P-deficient conditions.

1. Previous macrophysiological studies suggested that temperature-driven colour lightness and body size variations strongly influence biogeographical patterns in ectotherms, but whether these trait-environment relationships scale to local assemblages and the extent to which they can be modified by dispersal remains largely unexplored. We test whether the predictions of the thermal melanism hypothesis and the Bergmann’s rule hold for local assemblages. We also assess whether these trait-environment relationships are more important for species adapted to less stable (lentic) habitats, due to their greater dispersal propensity compared to those adapted to stable (lotic) habitats. 2. We quantified the colour lightness and body volume of 99 European dragon- and damselflies (Odonata) and combined these trait information with survey data for 518 local assemblages across Europe. Based on this continent-wide yet spatially explicit dataset, we tested for effects temperature and precipitation on the colour lightness and body volume of local assemblages and assessed differences in their relative importance and strength between lentic and lotic assemblages, while accounting for spatial and phylogenetic autocorrelation. 3. The colour lightness of assemblages of odonates increased and body size decreased with increasing temperature. Trait-environment relationships in the average and phylogenetic predicted component were equally important for assemblages of both habitat types but were stronger in lentic assemblages when accounting for phylogenetic autocorrelation. 4. Our results show that the mechanism underlying colour lightness and body size variations scale to local assemblages, indicating their general importance. These mechanisms were of equal evolutionary significance for lentic and lotic species, but higher dispersal ability seems to enable lentic species to cope better with historical climatic changes. The documented differences between lentic and lotic assemblages also highlight the importance of integrating interactions of thermal adaptations with proxies of the dispersal ability of species into trait-based models, for improving our understanding of climate-driven biological responses.

Leaf shape is a defining feature of how we recognise and classify plant species. Although there is extensive variation in leaf shape within many species, few studies have disentangled the underlying genetic architecture. We characterised the genetic architecture of leaf shape variation in Eurasian aspen (Populus tremula L.) by performing a genome wide association studies (GWAS) for physiognomy traits. To ascertain the roles of identified GWAS candidate genes within the leaf development transcriptional program, we performed gene co-expression network analyses from a developmental series, which is publicly available at http://aspleaf.plantgenie.org. We additionally used gene expression measurements across the population to analyse GWAS candidate genes in the context of a population-wide co-expression network and to identify genes that were differentially expressed between groups of individuals with contrasting leaf shapes. These data were integrated with expression GWAS (eQTL) results to define a set of candidate genes associated with leaf shape variation. Our results identified no clear adaptive link to leaf shape variation and indicate that leaf shape traits are genetically complex, likely determined by numerous small-effect variations in gene expression. Genes associated with shape variation were peripheral within the population-wide co-expression network, were not highly connected within the leaf development co-expression network and exhibited signatures of relaxed selection. As such, our results are consistent with the omnigenic model.

Predator-prey interactions are critical to understand how communities function. However, we need to describe intraspecific variation in diet to accurately depict those interactions. Harbor seals (Phoca vitulina, Linnaeus 1758) are an abundant marine predator that prey on species of conservation concern. We estimated intrapopulation feeding diversity of harbor seals in the Salish Sea relative to sex, time, and location with a novel approach that combined molecular techniques, repeated cross-sectional sampling of scat, and a specialization metric (within-individual consistency in diet). Based on 1,083 scat samples collected from five haul-out sites during four non-sequential years, we quantified diet using metabarcoding techniques, and determined the sex of the scat depositor using a molecular assay. Results suggest that intrapopulation feeding diversity was pervasive. Specialization was high over short periods (24 - 48 hours,〖PS〗_i = 0.392, 95% CI = 0.013, R = 100,000) and variable in time and space. Females showed more specialization than males, particularly during summer and fall, and demersal and benthic prey species were correlated with more specialized diets. The latter finding suggests that this type of prey likely require specific foraging strategies and that there are trade-offs between pelagic and benthic foraging styles for harbor seals. This differential feeding on prey species, as well as between sexes of harbor seals, indicate that predator-prey interactions in harbor seals are complex and that each sex may have a different impact on species of conservation concern. As such, describing intraspecific variation in diet may unravel hitherto unknown complex predator-prey interactions in the community.

1) The more demanding requirements of DNA preservation for genomic research can be difficult to meet when field conditions limit the methodological approaches that can be used, or cause samples to be stored in suboptimal conditions. Such limitations may increase rates of DNA degradation, potentially rendering samples unusable for applications such as genome-wide sequencing. Nonetheless, little is known about the impact of suboptimal sampling conditions. 2) We evaluated the performance of two widely used preservation solutions (1. DESS: 20% DMSO, 0.25M EDTA, NaCl saturated solution, and 2. ethanol) under a range of storage conditions over a three-month period (sampling at 1 day, 1 week, 2 weeks, 1 month, and 3 months) to provide practical guidelines for DNA preservation. DNA degradation was quantified as the reduction in average DNA fragment size over time (DNA fragmentation) because the size distribution of DNA segments plays a key role in generating genomic datasets. Tissues were collected from a marine teleost species, the Australasian snapper, Chrysophrys auratus. 3) We found that the storage solution has a dramatic effect on DNA preservation. In DESS, DNA was only moderately degraded after three months of storage while DNA stored in ethanol showed high levels of DNA degradation already within 24 hours, making samples unsuitable for next-generation-sequencing. 4) We recommend DESS as the most promising solution to improve DNA preservation. These results provide practical and economical advice to improve DNA preservation when sampling for genome-wide applications. Keywords: DMSO, DNA preservation, ethanol, fish, next-generation-sequencing, NGS, snapper

Secondary contact between closely related species can lead to the formation of hybrid zones, allowing for interspecific gene flow among taxa. Species replacement can take place if one of the species possesses a competitive advantage over the other, resulting in hybrid zone movement. This displacement may leave a genomic footprint across the landscape in the form of asymmetric introgression of selectively neutral alleles from the displaced to the advancing species. Hybrid zone movement has been suggested for marbled newts in the Iberian Peninsula, supported by the presence of a Triturus marmoratus stronghold surrounded by populations of the supposedly advancing T. pygmaeus in the northwest of the Lisbon Peninsula, i.e., an enclave. Moreover, a newly constructed two-species distribution model suggests that climate conditions following the Last Glacial Maximum may have favoured T. pygmaeus over T. marmoratus along the Atlantic coast. To test for the presence of a T. marmoratus genomic footprint in the area that may have witnessed species displacement, we developed and employed 54 nuclear SNPs and one mitochondrial DNA marker. We found no additional enclaves nor genetic traces of T. marmoratus in T. pygmaeus populations. Therefore, two main hypothesis arise in the absence of a genomic footprint: i) species replacement without hybridisation, either in allopatry or in sympatry under strong reproductive isolation; or ii) displacement with hybridisation where the footprint was eroded due to strong purifying selection. We predict testing for a genomic footprint north of the reported enclave could confirm that species replacement in the marbled newts occurred with hybridisation.

Small mammal abundances are frequently limited by resource availability but predators can exert strong lethal (direct mortality) and non-lethal limitations (e.g. depressed site-level activity). Artificially increasing resource availability for small mammals provides a unique opportunity to examine predator-prey interactions. We monitored the 3-year response of arboreal rodents and their predators at nest platforms (n = 598; 23 young forest sites), using annual inspections and remote cameras (n = 168). One year after adding nest platforms we found a 2.9 to 9.2-fold increase in red tree vole (Arborimus longicaudus) use at the site-level, but little use by potential predators. Predator use of nest platforms began in year two and increased in year three of the study. Most potential nest predators were positively correlated with tree vole presence at nest platforms but effect size and direction varied with temporal grain considered (e.g. hour vs day time-bin widths). Flying squirrels (Glaucomys humboldtensis) were positively correlated with disturbances caused by digging birds. Using a Cormack-Jolly-Seber model and encounter histories produced from visual re-captures of marked tree voles, we estimated apparent annual survival to be 0.099 ± 0.057 (x̄ ± 1 SE) for females and 0.005 ± 0.014 for males. Weasels (Mustela spp.), an active seeking predator, preyed upon tree voles most frequently with 10% of weasel detections resulting in mortality of a tree vole (n = 8) whereas owls, an ambush predator, did not prey upon tree voles at nest platforms even though they were detected at similar frequencies as weasels. Weasels also exerted potential non-lethal effects and we observed a >10-fold reduction in the number of tree vole detections per week after weasel detection. Our evidence indicates that predators exert direct and indirect effects on tree vole populations with active seeking predators being the most important predators at nest sites.

Anthropogenic perturbations such as harvesting often select against a large body size, and are predicted to induce rapid evolution towards smaller body sizes and earlier maturation. However, the evolvability of body size and size-correlated traits remains seldom evaluated in wild populations. Here, we use a laboratory experiment over 6 generations to measure the ability of wild-caught medaka fish (Oryzias latipes) to evolve in response to bidirectional size-dependent selection mimicking opposite harvest regimes. Specifically, we imposed selection against a small body size (Large line), against a large body size (Small line) or random selection (Control line), and measured correlated responses across multiple phenotypic, life-history and endocrine traits. As expected, the Large line evolved faster somatic growth and delayed maturation, but also evolved smaller body sizes at hatch, with no change in average levels of pituitary gene expressions of luteinizing, follicle-stimulating or growth (GH) hormones. In contrast, the Small medaka line was unable to evolve smaller body sizes or earlier maturation, but showed marginally-significant signs of increased reproductive investment, including larger egg sizes and elevated pituitary GH production. Natural selection on medaka body size was too weak to significantly hinder the effect of artificial selection, indicating that the asymmetric body-size response to size-dependent selection reflected an asymmetry in body-size evolvability. Our results show that trait evolvability may be contingent upon the direction of selection, and that a detailed knowledge of trait evolutionary potential is needed to forecast population response to anthropogenic change.

1. Plant leaf stomata are the gatekeepers of the atmosphere-plant interface and are essential building blocks of land surface models as they control transpiration and photosynthesis. Although more stomatal trait data is needed to significantly reduce the error in these model predictions, recording these traits is time-consuming and no standardized protocol is currently available. Some attempts were made to automate stomatal detection from photomicrographs, however, these approaches have the disadvantage of using classic image processing or targeting a narrow taxonomic entity which makes these technologies less robust and generalizable to other plant species. We propose an easy-to-use and adaptable workflow from leaf to label. A methodology for automatic stomata detection was developed using deep neural networks according to the state-of-the-art and its applicability demonstrated across the phylogeny of the angiosperms. 2. We used a patch-based approach for training/tuning three different deep learning architectures. For training, we used 431 micrographs taken from leaf prints made according the nail polish method from herbarium specimens of 19 species. The best performing architecture was tested on 595 images of 16 additional species spread across the angiosperm phylogeny. 3. The nail polish method was successfully applied in 78% of the species sampled here. The VGG19 architecture slightly outperformed the basic shallow and deep architectures, with a confidence threshold equal to 0.7 resulting in an optimal trade-off between precision and recall. Applying this threshold the VGG19 architecture obtained an average F-score of 0.87, 0.89 and 0.67 on the training, validation and unseen test set, respectively. The average accuracy was very high (94%) for computed stomatal counts on unseen images of species used for training. 4. The leaf-to-label pipeline is an easy-to-use workflow for researchers of different areas of expertise interested in detecting stomata more efficiently. The described methodology was based on multiple species and well-established methods so that it can serve as a reference for future work.

Recurrent sea urchin mass mortality has recently affected eastern Atlantic populations of the barren-forming sea urchin Diadema africanum. This new episode of die-off affords the opportunity to determine common meteorological and oceanographic conditions that may promote disease outbreaks. The population dynamics of this sea urchin species are well known—urchin barrens have persisted for many decades along most of the coastlines off the archipelagos of Madeira, Selvages and the Canary Islands, where they limit macroalgae biomass growth. However, this new and explosive mortality event decimated the sea urchin population by 93% on Tenerife and La Palma Islands. Two severe episodes of southwestern rough sea that lead to winter storms, in February 2010 (Xynthia) and February 2018 (Emma), preceded both mass mortality events. The autumn and winter months of those years were anomalous and characterized by swells with an average wave height above 2 m that hit the south and southwest sides of the islands. The amoeba Paramoeba brachiphila was the only pathogen isolated this time from the moribund and dead sea urchins, suggesting that the amoeba was the primary cause of the mortality. This new sea urchin die-off event supports the “killer-storm” hypothesis that has been already described for western Atlantic coasts. These anomalous southwest storms during winters generate pronounced underwater sediment movement and large-scale vertical mixing, detected in local tide gauge, which may promote paramoebiasis. This study presents valuable insights about climate-mediated changes in disease frequency and its impacts on the future of coastal marine ecosystems in the Atlantic.

Fungi are a key component of tropical biodiversity. Due to their inconspicuous and largely subterranean nature, they are however usually neglected in biodiversity inventories. The goal of this study was to identify the key determinants of fungal richness, community composition, and turnover in tropical rainforests. We tested specifically for the effect of soil properties, habitat, and locality in Amazonia. For these analyses, we used high-throughput sequencing data of short and long reads of fungal DNA present in soil and organic litter samples, combining existing and novel genomic data. Habitat type (phytophysiognomies) emerges as the strongest factor in explaining fungal community composition. Naturally open areas – campinas – are the richest habitat overall. Soil properties have different effects depending on the soil layer (litter or mineral soil) and the choice of genetic marker. We suggest that campinas could be a neglected hotspot of fungal diversity. An underlying cause for their rich diversity may be the overall low soil fertility, which increases the reliance on biotic interactions essential for nutrient absorption in these environments, notably ectomycorrhizal fungi–plant associations. Our results highlight the advantages of using both short and long DNA reads produced through high-throughput sequencing to characterize fungal diversity. While short-reads can suffice for diversity and community comparison, long-reads add taxonomic precision and have the potential to reveal population diversity.

Over thirty species of littoral marine Gammaridea occur along the coasts of the North Atlantic. From one to several species can coexist in a single region. There is an evident, inverse relationship between egg incubation time and temperature (from 14 to > 120 days) and consequent trends in the size of the animals on reaching maturity (from 5 mm in warmer waters to 30 mm in the coldest ones) and in lifespan (from < 6 months to > 5 years). Littoral gammarids are a good example of the shrinking size effect of increasing temperatures and size-related species diversity. In large species the annual cohorts of the population (3 to 5 annual size groups) functionally replace the adults of smaller species. The ongoing warming of the European Arctic seas may extend the distribution limits of boreal species so that more Gammarus species may appear on northern coasts hitherto occupied by just one or at most two species.

The Southern Ocean is one of the most isolated marine ecosystems, characterized by high levels of endemism, diversity, and biomass. Ascidians are among the dominant groups in Antarctic benthic assemblages, thus recording the evolutionary patterns of this group is crucial to improve our current understanding of the assembly of this polar ocean. We studied the genetic variation within Cnemidocarpa verrucosa sensu lato, one of the most widely distributed abundant and studied ascidian species in Antarctica. Using a mitochondrial and a nuclear gene (COI and 18S), the phylogeography of fifteen populations distributed along the Antarctic Peninsula and South America (Burdwood Bank/MPA Namuncurá) was characterized, where the bimodal distribution of the genetic distance suggested the existence of two species within the nominal C. verrucosa. When re-evaluating morphological traits to distinguish between genetically defined species, the presence of a basal disc in one of the genotypes could be a morphological trait to differentiate the species. These results are surprising due to the large research that has been carried out with the conspicuous C. verrucosa with no differentiation between species. Furthermore, it provides important tools to distinguish species in the field and laboratory. But also, these results give new insights to patterns of differentiation between closely related species that are distributed in sympatry, where the permeability of species boundaries still needs to be well understood.

Population dynamics models combine density-dependence and environmental effects. Ignoring sampling uncertainty might lead to biased estimation of the strength of density-dependence. This is typically addressed using state-space model approaches, which integrate sampling error and population process estimates. Such models seldom include an explicit link between the sampling procedures and the true abundance, which is common in capture-recapture settings. However, many of the models proposed to estimate abundance in the presence of heterogeneity lead to incomplete likelihood functions and cannot be straightforwardly included in state-space models. We assessed the importance of estimating sampling error explicitly by taking an intermediate approach between ignoring uncertainty in abundance estimates and fully specified state-space models for density-dependence estimation based on autoregressive processes. First, we estimated individual capture probabilities based on a heterogeneity model, using a conditional multinomial likelihood, followed by a Horvitz-Thompson estimate for abundance. Second, we estimated coefficients of autoregressive models for the log abundance. Inference was performed using the methodology of integrated nested Laplace approximation (INLA). We performed an extensive simulation study to compare our approach with estimates disregarding capture history information, and using R-package VGAM, for different parameter specifications. The methods were then applied to a real dataset of gray-sided voles Myodes rufocanus from Northern Norway. We found that density-dependence estimation was improved when explicitly modelling sampling error in scenarios with low innovation variances, in which differences in coverage reached up to 8% in estimating the coefficients of the autoregressive processes. In this case, the bias also increased assuming a Poisson distribution in the observational model. For high innovation variances, the differences between methods were small and it appeared less important to model heterogeneity.

1. The reduction of plant diversity following eutrophication threatens many ecosystems worldwide. Yet, the mechanisms by which species are lost following nutrient enrichment are still not completely understood, nor are the details of when such mechanisms act during the growing season, which hampers understanding and the development of mitigation strategies. 2. Using a common garden competition experiment, we found that early-season differences in growth rates among five perennial grass species measured in monoculture predicted short-term competitive dominance in pairwise combinations and that this effect was stronger under a fertilisation treatment. 3. We also examined the role of early-season growth rate in determining the outcome of competition along an experimental nutrient gradient in an alpine meadow. Early differences in growth rate between species predicted short-term competitive dominance under both ambient and fertilized conditions and competitive exclusion under fertilized conditions. 4. The results of these two studies suggests that plant species growing faster during the early stage of the growing season gain a competitive advantage over species that initially grow more slowly, and that this advantage is magnified under fertilisation. This finding is consistent with the theory of asymmetric competition for light in which fast-growing species can intercept incident light and hence outcompete and exclude slower-growing (and hence shorter) species. We predict that the current chronic nutrient inputs into many terrestrial ecosystems worldwide will reduce plant diversity and maintain low biodiversity state by continuously favouring fast-growing species. Biodiversity management strategies should focus on controlling nutrient inputs and reducing the growth of fast-growing species early in the season.

We use adaptive dynamics models to study how changes in the abiotic environment affect patterns of evolutionary dynamics and diversity in evolving communities of organisms with complex phenotypes. The models are based on the logistic competition model and environmental changes are implemented as a temporal change of the carrying capacity as a function of phenotype. In general we observe that environmental changes cause a reduction in the number of species, in total population size, and in phenotypic diversity. The rate of environmental change is crucial for determining whether a community survives or undergoes extinction. Until some critical rate of environmental changes, species are able to follow evolutionarily the shifting phenotypic optimum of the carrying capacity, and many communities adapt to the changing conditions and converge to new stationary states. When environmental changes stop, such communities gradually restore their initial phenotypic diversity.

The biogeographical distribution of diversity among populations of threatened mammalian species is generally investigated through population genetics. However, intraspecific phenotypic diversity is rarely assessed beyond taxonomy-focused linear measurements or qualitative descriptions. Here, we use a technique widely used in the evolutionary sciences – geometric morphometrics – to characterize shape diversity in the skull of an endangered marsupial, the northern quoll, across its 5,000 km distribution range along the northern Australian coast. Skull shape is a proxy of feeding, behaviour, and phenotypic differentiation, allowing us to ask if populations can be distinguished and if patterns of variation indicate adaptability to changing environmental conditions. We analysed skull shape in 101 individuals across the four mainland populations and several islands. We assessed the contribution of population, size, sex, rainfall, temperature, and latitude/longitude to skull shape variation through Principal Components, Procrustes ANOVA, and variation partitioning analyses. Regardless of land area inhabited, northern quoll populations harbour similar amounts of broadly overlapping skull shape variation. Size predicted skull shape best, coinciding with braincase size variation and differences in the cheekbone shape. Size-adjusted population differences explained less variation with far smaller effect sizes, relating to changes in insertion areas of masticatory muscles, as well as the upper muzzle and incisor region. Climatic and geographic variables contributed little or nothing. Strikingly, the vast majority of shape variation - 76% - remained unexplained. Our results suggest a uniform within-species scope for shape variation, possibly due to phenotypic plasticity or allometric constraints. The lack of local adaptation indicates that cross-breeding between populations will not reduce local morphological skull (and probably general musculoskeletal) adaptation because none exists. However, the potential for heritable morphological variation (e.g. specialization to local diets) seems exceedingly limited. We conclude that 3D geometric morphometrics can provide a comprehensive, statistically rigorous phenomic contribution to genetics-based conservation studies.

The Sanjiang Plain is the biggest freshwater wetland locating within northeastern China. Due to climate change and human activities, that wetland has degraded to a successional gradient from the original flooded wetland to dry shrub vegetation and a forest area with lower ground water level, resulting in changes in soil microbiologic structure and functions. The present study investigated the microbial diversity and community structure in relation to soil properties along this gradient. The soil physic-chemical properties changed significantly with degradation. The Shannon variety of soil fungi as well as bacteria varied significantly with successional stage (both P < 0.05). The community structures of soil bacteria and fungi in the early successional stages (i.e., the wetland) were significantly structured via total phosphorus, available nitrogen and total nitrogen concentrations in soils, while those in the later successional stages (i.e., forests) were significantly structured by soil organic carbon, soil pH and available phosphorus concentrations. Our results indicated that variations in the soil environment affected soil microbial communities along a successional gradient from wetland to forests are mainly. These outcomes indicate that above ground plant composition is a forceful determinant of the structure as well as functions of bacterial and fungal communities, might finally causing substantial alterations in ecosystem activity.