Transplantation Experiments Research Articles

Range‐wide intraspecific variation reflects past adaptation to climate in a gypsophile Mediterranean shrub

Abstract Phenotypic differences among populations stem from the interaction between neutral and adaptive processes, and phenotypic plasticity. Although clinal trait variation along climatic gradients often evolves in widely distributed species, it is unknown whether substrate specialization, such as that of Mediterranean gypsum plants, has constrained adaptation to climate. Using a common garden experiment with two contrasting watering treatments, we quantified phenotypic plasticity, assessed evidence for footprints of selection using FST – QST comparisons, and evaluated the ecological factors driving genetically based phenotypic differentiation of 11 populations encompassing the full environmental range of the gypsum shrub Lepidium subulatum. We found evidence for genetic differentiation among populations related to climatic differences, with populations from warmer and drier sites showing lower specific leaf area and leaf N, earlier phenology, greater water use efficiency and greater fitness. Multiple lines of evidence suggest that this differentiation was driven by past divergent selection rather than neutral processes. All populations showed high phenotypic plasticity, indicating that plasticity has not been selected against, even in populations from sites with harsher climatic conditions. Synthesis. Our results indicate that despite strong substrate specialization, adaptive differentiation related to climatic gradients occurs in this species. However, we also found that populations from mesic sites may be particularly vulnerable to future climate change given their relatively lower fitness under both wet and dry conditions.

Soil microbial identity explains home-field advantage for litter decomposition.

Unraveling the mechanisms of home-field advantage (HFA) is essential to gain a complete understanding of litter decomposition processes. However, knowledge of the relationships between HFA effects and microbial communities is lacking. To examine HFA effects on litter decomposition, we identified the microbial communities and conducted a reciprocal transplant experiment, including all possible combinations of soil and litter, between sites at two elevations in cool-temperate forests. Soil origin, rather than HFA, was an important factor in controlling litter decomposition processes. Microbiome-wide association analyses identified litter fungi and bacteria specific to the source soil, which completely differed at a low taxonomic level between litter types. The relative abundance of these microbes specific to source soil was positively correlated with litter mass loss. The results indicated that the unique relationships between plant litter and soil microbes through plant-soil linkages drive litter decomposition processes. In the short term, soil disturbances resulting from land-use changes have the potential to disrupt the effect of soil origin and hinder the advancement of litter decomposition. These findings contribute to an understanding of HFA mechanisms and the impacts of land-use change on decomposition processes in forest ecosystems.

Transgenerational salinity tolerance plasticity in rice: Unraveling nongenetic phenotypic modifications and environmental influences.

Transgenerational plasticity in plants enables rapid adaptation to environmental changes, allowing organisms and their offspring to adapt to the environment without altering their underlying DNA. In this study, we investigated the plasticity of transgenerational salinity tolerance of rice plants using a reciprocal transplant experimental strategy. Our aim was to assess whether nongenetic environment-induced phenotypic modifications and transgenerational salinity affect the salinity tolerance of progeny while excluding nuclear genomic factors for two generations. Using salt-tolerant and salt-sensitive rice genotypes, we observed that the parentally salt-stressed salt-sensitive genotype displayed greater growth performance, photosynthetic activity, yield performance, and transcriptional responses than did the parentally nonstressed salt-sensitive plants under salt stress conditions. Surprisingly, salt stress-exposed salt-tolerant progeny did not exhibit as much salinity tolerance as salt stress-exposed salt-sensitive progeny under salt stress. Our findings indicate that the phenotypes of offspring plants differed based on the environment experienced by their ancestors, resulting in heritable transgenerational phenotypic modifications in salt-sensitive genotypes via maternal effects. These results elucidated the mechanisms underlying transgenerational plasticity in salinity tolerance, providing valuable insights into how plants respond to changing environmental conditions.

Knock down of APE1 suppressed gastric cancer metastasis via improving immune disorders caused by myeloid-derived suppressor cells

ABSTRACT Gastric cancer is a highly immunogenic malignancy. Immune tolerance facilitated by myeloid-derived suppressor cells (MDSCs) has been implicated in gastric cancer resistance mechanisms. The potential role of APE1 in regulating gastric cancer metastasis by targeting MDSCs remains uncertain. In this study, the plasmid Plxpsp-mGM-CSF was used to induce high expression of granulocyte-macrophage colony-stimulating factor (GM-CSF) in GES-1 cells. For tumor transplantation experiments, AGS, AGS+GM-CSF and AGS+GM-CSF-siAPE1 cell lines were established by transfection, followed by subcutaneous implantation of tumor cells. MDSCs, Treg cells, IgG, CD3 and CD8 levels were assessed. Transfection with siAPE1 significantly inhibited tumor growth compared to the AGS+GM-CSF group. APE1 gene knockdown modulated the immune system in gastric cancer mice, characterized by a decrease in MDSCs and an increase in Treg cells, IgG, CD3 and CD8. In addition, APE1 gene knockdown resulted in decreased levels of pro-MDSC cytokines (HGF, CCL5, IL-6, CCL12). Furthermore, APE1 gene knockdown inhibited proliferation, migration and invasion of AGS and MKN45 cells. AGS-GM-CSF cell transplantation increased MDSC levels and accelerated tumor growth, whereas APE1 knockdown reduced MDSC levels, inhibited tumor growth and attenuated inflammatory infiltration in gastric cancer tissues. Strategies targeting the APE1/MDSC axis offer a promising approach to the prevention and treatment of gastric cancer, providing new insights into its management.

Microbiome processing of organic nitrogen input supports growth and cyanotoxin production of Microcystis aeruginosa cultures.

Nutrient-induced blooms of the globally abundant freshwater toxic cyanobacterium Microcystis cause worldwide public and ecosystem health concerns. The response of Microcystis growth and toxin production to new and recycled nitrogen (N) inputs, and the impact of heterotrophic bacteria in the Microcystis phycosphere on these processes are not well understood. Here, using microbiome transplant experiments, cyanotoxin analysis, and nanometer-scale stable isotope probing to measure N incorporation and exchange at single cell resolution, we monitored the growth, cyanotoxin production, and microbiome community structure of several Microcystis strains grown on amino acids or proteins as the sole N source. We demonstrate that the type of organic N available shaped the microbial community associated with Microcystis, and external organic N input led to decreased bacterial colonization of Microcystis colonies. Our data also suggest that certain Microcystis strains could directly uptake amino acids, but with lower rates than heterotrophic bacteria. Toxin analysis showed that biomass-specific microcystin production was not impacted by N source (i.e., nitrate, amino acids or protein) but rather by total N availability. Single-cell isotope incorporation revealed that some bacterial communities competed with Microcystis for organic N, but other communities promoted increased N uptake by Microcystis, likely through ammonification or organic N modification. Our laboratory culture data suggest that organic N input could support Microcystis blooms and toxin production in nature, and Microcystis-associated microbial communities likely play critical roles in this process by influencing cyanobacterial succession through either decreasing (via competition) or increasing (via biotransformation) N availability, especially under inorganic N scarcity.

Cu exposure induces liver inflammation via regulating gut microbiota/LPS/liver TLR4 signaling axis

Copper (Cu) serves as an essential cofactor in all organisms, yet excessive Cu exposure is widely recognized for its role in inducing liver inflammation. However, the precise mechanism by which Cu triggers liver inflammation in ducks, particularly in relation to the interplay in gut microbiota regulation, has remained elusive. In this investigation, we sought to elucidate the impact of Cu exposure on liver inflammation through gut-liver axis in ducks. Our findings revealed that Cu exposure markedly elevated liver AST and ALT levels and induced liver inflammation through upregulating pro-inflammatory cytokines (IL-1β, IL-6 and TNF-α) and triggering the LPS/TLR4/NF-κB signaling pathway. Simultaneously, Cu exposure induced alterations in the composition of intestinal flora communities, notably increasing the relative abundance of Sphingobacterium, Campylobacter, Acinetobacter and reducing the relative abundance of Lactobacillus. Cu exposure significantly decreased the protein expression related to intestinal barrier (Occludin, Claudin-1 and ZO-1) and promoted the secretion of intestinal pro-inflammatory cytokines. Furthermore, correlation analysis was observed that intestinal microbiome and gut barrier induced by Cu were closely related to liver inflammation. Fecal microbiota transplantation (FMT) experiments further demonstrated the microbiota-depleted ducks transplanting fecal samples from Cu-exposed ducks disturbed the intestinal dysfunction, which lead to impaire liver function and activate the liver inflammation. Our study provided insights into the mechanism by which Cu exposure induced liver inflammation in ducks through the regulation of gut-liver axis. These results enhanced our comprehension of the potential mechanisms driving Cu-induced hepatotoxicity in avian species.

Precocity of fruiting in Pinus yunnanensis revealed by common garden experiment and association analysis with molecular markers

Shortening the growth cycle and maintaining high yields are the goals pursued by all breeders. Under the distinct ecological conditions of tropical and rainy and dry seasons, many plants experience the phenomenon of early flowering and fruiting under adverse conditions. However, there is still no clear conclusion as to whether early flowering and fruiting will affect the vegetative growth of plants. In this study, the vegetative and reproductive growth traits of 270 families of Pinus yunnanensis of 6 years old in common garden were analyzed. There were significant differences in phenotypic traits of different populations, indicating that the trait differences were genetically stable. The variation of vegetative growth traits mainly comes from within populations, and the variation of reproductive growth traits mainly comes among populations. The average coefficient of variation of reproductive growth traits was significantly higher than that of vegetative growth traits. There was a positive correlation between vegetative growth traits and precocity of fruiting. Ecological environment factors have impact on phenotypic variation, among which the temperature of the wettest season has the greatest impact. According to cluster analysis, the nine populations could be divided into three categories. These cluster results were not due to geographic distances, indicating the discontinuity of phenotypic variation. By association analysis, 9 and 3 SSR loci were found to be significantly associated with the trait of precocity of fruiting, respectively. Among them, PK840 locus had the highest interpretation rate for the trait of precocity of fruiting. There were 10 and 7 unique haplotypes in precocity and non-precocity of fruiting individuals, with 179 bp and 183 bp bands, respectively. According to the breeding goal of high productivity and precocity of fruiting, 54 superior families and 200 superior individuals were selected. This study explored the adaptation strategies of tree growth traits to environmental changes.

Variation and plasticity in life-history traits and fitness of wild Arabidopsis thaliana populations are not related to their genotypic and ecological diversity

BackgroundDespite its implications for population dynamics and evolution, the relationship between genetic and phenotypic variation in wild populations remains unclear. Here, we estimated variation and plasticity in life-history traits and fitness of the annual plant Arabidopsis thaliana in two common garden experiments that differed in environmental conditions. We used up to 306 maternal inbred lines from six Iberian populations characterized by low and high genotypic (based on whole-genome sequences) and ecological (vegetation type) diversity.ResultsLow and high genotypic and ecological diversity was found in edge and core Iberian environments, respectively. Given that selection is expected to be stronger in edge environments and that ecological diversity may enhance both phenotypic variation and plasticity, we expected genotypic diversity to be positively associated with phenotypic variation and plasticity. However, maternal lines, irrespective of the genotypic and ecological diversity of their population of origin, exhibited a substantial amount of phenotypic variation and plasticity for all traits. Furthermore, all populations harbored maternal lines with canalization (robustness) or sensitivity in response to harsher environmental conditions in one of the two experiments.ConclusionsOverall, we conclude that the environmental attributes of each population probably determine their genotypic diversity, but all populations maintain substantial phenotypic variation and plasticity for all traits, which represents an asset to endure in changing environments.

Is competitive ability the key adaptation to benign environments? Revisiting experiments on closely related species of tidal plants.

A central goal in biology is to understand which traits underlie adaptation to different environments. Yet, few studies have examined the relative contribution of competitive ability towards adaptive divergence among species occupying distinct environments. Here, we test the relative importance of competitive ability as an adaptation to relatively benign versus challenging environments, using previously published studies of closely related species pairs of primarily tidal plants subjected to reciprocal removal with transplant experiments in nature. Subordinate species typically occupy more challenging environments and showed consistent evidence for adaptation to challenging conditions, with no significant competitive effect on non-local, dominant species. In contrast, dominant species typically occupy relatively benign environments and performed significantly better than non-local, subordinate species that faced competition from the dominant species. Surprisingly, when the two species were not allowed to compete, the subordinate species performed as well as the dominant species in the benign environments where the subordinate species do not occur. These results suggest that competitive ability is the most important adaptation distinguishing the species that occupy relatively benign environments. The limited scope and number of suitable experimental studies encourage future work to test if these results are generalizable across taxa and environments.

Are novel or locally adapted pathogens more devastating and why? Resolving opposing hypotheses.

There is a rich literature highlighting that pathogens are generally better adapted to infect local than novel hosts, and a separate seemingly contradictory literature indicating that novel pathogens pose the greatest threat to biodiversity and public health. Here, using Batrachochytrium dendrobatidis, the fungus associated with worldwide amphibian declines, we test the hypothesis that there is enough variance in "novel" (quantified by geographic and phylogenetic distance) host-pathogen outcomes to pose substantial risk of pathogen introductions despite local adaptation being common. Our continental-scale common garden experiment and global-scale meta-analysis demonstrate that local amphibian-fungal interactions result in higher pathogen prevalence, pathogen growth, and host mortality, but novel interactions led to variable consequences with especially virulent host-pathogen combinations still occurring. Thus, while most pathogen introductions are benign, enough variance exists in novel host-pathogen outcomes that moving organisms around the planet greatly increases the chance of pathogen introductions causing profound harm.

Genomic, climatic, and cultural diversity of maize landraces from the Himalayan Kingdom of Bhutan

Societal Impact StatementBhutan is an ancient kingdom in the Himalayan range and one of the most rugged, geodiverse, and mountainous agricultural countries in the world. Historically secluded and geographically isolated, Bhutan is a hotspot for Himalayan agrobiodiversity where small‐scale agriculture supports the livelihoods of a large share of the resident population. Here, Bhutanese maize agrobiodiversity is explored to unlock its adaptation potential using genomics and participatory variety selection in combination with climate research. We show that Bhutanese traditional farmers maintain a wealth of diversity that may support the sustainable intensification of maize cropping in the Himalayas and beyond.Summary Bhutan, an ancient kingdom enshrouded in the Himalayas, hosts largely untapped agrobiodiversity that may harness genetic variation useful for adaptation to local climates and user needs. Here, we genotyped‐by‐sequencing 351 pooled samples of local maize (Zea mays L.) landraces, the entire collection of the Bhutan National Gene Bank, comparing their genomic diversity with maize from other countries in the Himalayan range. We reconstructed the adaptation of Bhutanese maize to historical and projected climates, identifying areas of future maladaptation. We then run a common garden experiment involving local smallholder farmers in a participatory evaluation of landraces' performance, aiming at the identification of quantitative trait nucleotides (QTNs) contributing to adaptation, performance, and farmers' choice. We found that Bhutanese maize agrobiodiversity is unique in the Himalayan range, and a locus on Chromosome 5 supports the differentiation of three distinct genetic clusters. We found that a portion of current genomic diversity can be associated with the Bhutanese landscape and that maize cultivation in the southwest of the country may be negatively impacted by projected climates. We also found that Bhutanese maize agrobiodiversity is large and may contribute to adaptation and improvement. A genome‐wide association study identified 117 QTNs for climatic adaptation, agronomic performance, and farmers' preferences. Our results show that Bhutanese maize landraces are a unique source of genetic agrobiodiversity for local adaptation. We found that the integration of genomics, climate science, and participatory methods can speed up the identification of genetic factors contributing to the sustainable intensification of maize cultivation in the Himalayas and beyond.

Polysaccharides of Aspergillus cristatus attenuate obesity by regulating gut microbiota and gut microbiota-related metabolites

Golden-flower fungus, the only dominant microorganism determining the Fu-brick tea quality through fermentation and the important microbe in Liupao tea, is considered a potential probiotic fungus based on its anti-obesity effect. However, the classification of golden-flower fungi is still controversial; the anti-obesity effect of golden-flower fungus polysaccharides remains unknown. In this study, we identify a golden-flower strain as Aspergillus cristatus based on morphological characteristics and multigene phylogeny analysis, which resolves the controversy of classification. Moreover, we find A. cristatus polysaccharides (ACPS) attenuate obesity in rats. ACPS modulate gut bacterial composition, in which Akkermansia, Akkermansia muciniphila, Bacteroides, Romboutsia, Blautia, and Desulfovibrio are considered the core microbes regulated by ACPS. ACPS increase fecal total short-chain fatty acid content and serum, hepatic, and fecal total bile acid content. Furthermore, ACPS-induced gut microbiota alteration plays a causal role in the protection from obesity, according to a fecal transplantation experiment. Thus, ACPS ameliorate obesity by regulating gut microbiota and gut microbiota-related metabolites.

Reprint of: Prelude to molecularization: The double gradient model of Sulo Toivonen and Lauri Saxén

The present molecular investigations of Organizer phenomena show a remarkable connection to the earlier classical embryological studies that used transplantation as a method for making mechanistic models of induction. One of the most prominent of these connections is the dual gradient model for anterior-posterior and dorsal-ventral polarity. This paper will discuss some of the history of how transplantation experiments provided data that could be interpreted in terms of two gradients of biologically active materials. It will highlight how the attempts to discover the elusive Induktionsstoffen gave rise to the double gradient model of Sulo Toivonen and Lauri Saxén in the 1950s and 1960s. This paper will also document how this research into the identity of these molecules gave rise to the developmental genetics that eventually would find the molecules responsible for primary embryonic induction.

Developing a Gel-Col Heterogeneous Network Hydrogel Scaffold for Tissue-Engineered Skin with Enhanced Basement Membrane Formation

Tissue-engineered skin is constructed utilizing biocompatible matrix materials as scaffolds, and incorporating human epidermal keratinocytes (HEKs), dermal fibroblasts (HDFs), and bioactive molecules, addressing clinical issues such as skin graft rejection and donor scarcity. Conventional fabrication methods of tissue-engineered skin typically rely on homogeneous single-network scaffolds, which may not adequately address the distinct requirements of epidermal and dermal cells or the critical formation of the basement membrane, potentially impacting the development and functional integrity of the skin tissue. Cell culture studies employing GelMA hydrogels of varied concentrations have elucidated that epidermal cells exhibit increased viability on scaffolds with higher concentrations (15%), whereas fibroblasts show increased proliferation within scaffolds of lower concentrations (5%). Consequently, we have engineered a GelMA-collagen (Gel-Col) hydrogel scaffold with a heterogeneous network of dense and sparse matrices, conducive to the growth of both fibroblasts and epidermal cells. Culturing epidermal cells on this innovative scaffold, coupled with histological section staining, RT-PCR, and Western blot analyses, has demonstrated that the scaffold significantly enhances cellular adhesion and promotes the development of the epidermal basement membrane compared to single-network hydrogels. Mouse wound transplantation experiments have further confirmed the scaffold’s effectiveness in accelerating wound repair, supporting the development of the basement membrane, and enhancing epidermal regeneration. This research provides a reference for constructing tissue-engineered skin that more precisely emulates the intricate architecture and cellular dynamics of natural skin, propelling the advancement of tissue-engineered skin grafts in the clinical treatment of skin injuries.

Single-tube Ptprc SNP genotyping of JAXBoy (CD45.1) and C57BL/6J (CD45.2) mice by endpoint PCR and gel electrophoresis

Allelic variation at the Ptprc gene, which encodes the pan leukocyte marker CD45/Ly5, is commonly exploited to track hematopoietic reconstitution by flow cytometry in mixed bone marrow chimera transplant experiments. Historically, this was accomplished using bone marrow from C57BL/6 (Ptprcb/CD45.2/Ly5.2) and congenic B6.SJL-PtprcaPepcb/Boy (Ptprca/CD45.1/Ly5.1) mice. Recently, the Jackson Laboratory directly CRISPR-engineered the Ptprca allele in C57BL/6J mice. This new isogenic strain, termed JAXBoy, differs from wild-type C57BL/6J mice by two nucleotides, compared to the biologically significant 37 megabase (Mb) SJL interval retained in B6.SJL-PtprcaPepcb/Boy/J mice. Currently, Ptprc/CD45 variants are identified by flow cytometry or allele-specific real-time PCR, both of which require specialized workflows and equipment compared to standard genotyping of endpoint PCR products by gel electrophoresis. Here, we employed allele-specific oligonucleotides in conjunction with differential incorporation of a long non-specific oligo 5’-tail to allow for simultaneous identification of the Ptprca and Ptprcb alleles using endpoint PCR and gel electrophoresis. This method allows for integration of Ptprc genotyping into standard genotyping workflows, which use a single set of thermocycling and gel electrophoresis conditions. Importantly, the strategy of primer placement and tail addition described here can be adapted to discriminate similar single- or multi-nucleotide polymorphisms at other genomic loci.

Sex‐specific consequences of host shift for morphology and fluctuating asymmetry in a seed beetle: an experimental evolution approach

AbstractMorphological plasticity may be induced by altering host plants and commonly it is sex‐specific in phytophagous insects. It is hypothesized that stress in insects caused by a host shift leads to morphological changes and developmental destabilization, which may be identified by fluctuating asymmetry. We performed reciprocal transplant experiments in eight replicated populations of the seed beetle Acanthoscelides obtectus (Say) (Coleoptera: Chrysomelidae, Bruchinae) adapted to common bean (Phaseolus vulgaris L., Fabaceae) as its ancestral host and chickpea (Cicer arietinum L., Fabaceae) as a suboptimal host. Using methods of geometric morphometrics we assessed the effects of the short‐ and long‐term host shift on size and shape variation of beetles of both sexes. We also tested the hypothesis that fluctuating asymmetry is lower when the beetles develop within seeds of the ancestral host. Our results showed that females respond more plastically related to their body size and shape of the abdomen, particularly during a short‐term host shift, suggesting that females have a greater importance in maintaining the population on a new host. The level of fluctuating asymmetry is lowest in males that have evolved for generations on bean indicating that they have the most canalized development, so we suggested that symmetry in A. obtectus is maintained by sexual selection. Even if the beetles are adapted to a suboptimal host over many generations, they stabilize their development after returning to the ancestral host. Our work indicates that host shifts may change morphological aspects of the beetles in a sex‐specific manner and consequently influence their developmental trajectories.

Intra- and interspecific diversity in a tropical plant clade alter herbivory and ecosystem resilience

Declines in biodiversity generated by anthropogenic stressors at both species and population levels can alter emergent processes instrumental to ecosystem function and resilience. As such, understanding the role of biodiversity in ecosystem function and its response to climate perturbation is increasingly important, especially in tropical systems where responses to changes in biodiversity are less predictable and more challenging to assess experimentally. Using large-scale transplant experiments conducted at five neotropical sites, we documented the impacts of changes in intraspecific and interspecific plant richness in the genus Piper on insect herbivory, insect richness, and ecosystem resilience to perturbations in water availability. We found that reductions of both intraspecific and interspecific Piper diversity had measurable and site-specific effects on herbivory, herbivorous insect richness, and plant mortality. The responses of these ecosystem-relevant processes to reduced intraspecific Piper richness were often similar in magnitude to the effects of reduced interspecific richness. Increased water availability reduced herbivory by 4.2% overall, and the response of herbivorous insect richness and herbivory to water availability were altered by both intra- and interspecific richness in a site-dependent manner. Our results underscore the role of intraspecific and interspecific richness as foundations of ecosystem function and the importance of community and location-specific contingencies in controlling function in complex tropical systems.

Intra- and interspecific diversity in a tropical plant clade alter herbivory and ecosystem resilience.

Declines in biodiversity generated by anthropogenic stressors at both species and population levels can alter emergent processes instrumental to ecosystem function and resilience. As such, understanding the role of biodiversity in ecosystem function and its response to climate perturbation is increasingly important, especially in tropical systems where responses to changes in biodiversity are less predictable and more challenging to assess experimentally. Using large-scale transplant experiments conducted at five neotropical sites, we documented the impacts of changes in intraspecific and interspecific plant richness in the genus Piper on insect herbivory, insect richness, and ecosystem resilience to perturbations in water availability. We found that reductions of both intraspecific and interspecific Piper diversity had measurable and site-specific effects on herbivory, herbivorous insect richness, and plant mortality. The responses of these ecosystem-relevant processes to reduced intraspecific Piper richness were often similar in magnitude to the effects of reduced interspecific richness. Increased water availability reduced herbivory by 4.2% overall, and the response of herbivorous insect richness and herbivory to water availability were altered by both intra- and interspecific richness in a site-dependent manner. Our results underscore the role of intraspecific and interspecific richness as foundations of ecosystem function and the importance of community and location-specific contingencies in controlling function in complex tropical systems.

Mixed-Species Stands Improve the Coordination between Leaf and Fine Root Traits in a Common Garden Experiment

The coordination between leaf and root traits is conducive to an integrated understanding of whole-plant ecological strategies and reveals how community composition and diversity contribute to defining the functions and services of ecosystems. However, there is limited understanding regarding the impact of species richness and trait categories on the coordination between leaf and root traits. Based on a 9-year common garden experiment, we investigated the leaf and fine root traits of 56 plots (25.8 m × 25.8 m) encompassing various trait categories (trait categories were defined according to the root depth, leaf habit, and mycorrhizal type) and different levels of species richness (1, 2, 4, 8) in the context of a forest biodiversity and ecosystem functioning experiment conducted in subtropical China (BEF-China). We found the following: (1) Our findings indicate that there was generally a significant difference in leaf traits, occasionally in absorptive root traits, and no difference in transport root traits between different trait categories. (2) Conversely, species richness significantly influenced all transport root traits except root nitrogen and most leaf and absorptive root traits. (3) The results demonstrated that trait categories played a crucial role in the coordination between leaf and fine root traits. Additionally, the coordination between leaf and fine root traits increased with higher species richness, particularly in deep-rooted, evergreen, and ectomycorrhizal fungi species. Furthermore, the coordination between leaf and fine root traits was significantly lower in monocultures compared to four- and eight-species mixtures. These results suggest that a significant mixture effect exists in the coordination between leaf and fine root traits due to the comprehensive and divergent capture of above- and belowground resources and reduced intraspecific competition. Therefore, compared to monocultures, mixed-species stands can enhance the coordination of leaf and fine root traits, and it is advisable to establish forests with mixtures of more than four species, dominated by deep-rooted, evergreen, and ectomycorrhizal fungi species, to maintain ecosystem stability and functional integrity.

Ash seedlings in a reciprocal transplant experiment—the extent of damage of mature forest stands affects ash offspring performance

In past decades, ash dieback has caused a rapid decline of European ash (Fraxinus excelsior) in temperate forests of Europe. Numerous studies focus on mitigating the negative impacts of ash dieback to forest ecosystems or identifying resistant genotypes. The role of natural selection toward genotypes withstanding ash dieback for ash regeneration has been less frequently studied with experimental means to date. This is, however, necessary in times of global change, because the preservation of ash in Europe’s forests will depend, above all, on the adaptability of the future generations of ash trees. To quantify the extent and effects of ash dieback severity for ash regeneration we selected five forest stands moderately damaged and five forest stands highly damaged by ash dieback, in Schleswig-Holstein, Germany. We reciprocally transplanted naturally regenerated ash seedlings sampled in the field between these 10 sites. A shading treatment added to each half of the plots per site was meant to test for effects of altered light conditions in the herb layer due to canopy opening caused by ash dieback. With this approach, we tested seedling survival, performance and fungal infection for an interacting effect of origin and target site in regard to ash dieback severity and environmental factors over 2 years and recorded leaf traits (specific leaf area, leaf dry matter content) in the second year. Reduced light conditions under the shading nets had strong effects, influencing first year performance and infection probability as well as second year survival, growth and leaf trait characteristics. Soil conditions had only a marginal influence on transplanted seedlings. Transplantation direction between moderately and highly damaged sites affected infection marginally during the first year and survival as well as leaf traits significantly during the second year. Most notably, seedlings transplanted from moderately damaged to severely damaged sites exhibited the highest infection probability and lowest SLA, while seedlings transplanted vice versa were least likely to be infected and exhibited the highest SLA. Results hint at a first filtering effect by the ash dieback history of a forest stand and might indicate a transition from ecologically to evolutionary driven differentiation of ash seedling responses.