Local adaptation has been demonstrated in many organisms, but the traits involved, and the temporal and spatial scales at which selection acts are generally unknown. We carried out a multi-year study of 200 accessions (natural inbred lines) of Swedish Arabidopsis thaliana using local field sites and a combination of common-garden experiments that measured adult survival and fecundity, and selection experiments that measured fitness over the full life cycle. We found evidence of strong and variable selection, with particular genotypes favored more than five-fold in certain years and locations. Fecundity showed evidence of classical local adaptation, with accessions generally performing better close to their home. However, southern accessions usually had the highest fecundity-but were far more sensitive to harsh winters and slug herbivory, which strongly decreased both survival and fecundity. Accessions originally sampled on beaches had low fecundity in all environments, but massively outperformed all other accessions in the selection experiments, presumably due to an advantage during seedling establishment associated with their very large seeds. We conclude that local adaptation in A. thaliana reflects strong temporally and spatially varying selection on multiple traits, generally involving trade-offs and different life-history strategies, making fitness difficult to predict and measure.

How evolutionary forces interact to maintain genetic variation within populations has been a matter of extensive theoretical debates. While mutation and exogenous gene flow increase genetic variation, stabilizing selection and genetic drift are expected to deplete it. To date, levels of genetic variation observed in natural populations are hard to predict without accounting for other processes, such as balancing selection in heterogeneous environments. We aimed to empirically test three hypotheses: (i) admixed populations have higher quantitative genetic variation due to introgression from other gene pools, (ii) quantitative genetic variation is lower in populations from harsher environments (i.e., experiencing stronger selection), and (iii) quantitative genetic variation is higher in populations from heterogeneous environments. Using growth, phenological and functional trait data from three clonal common gardens and 33 populations (522 clones) of maritime pine (Pinus pinaster Aiton), we estimated the association between the population-specific total genetic variances (i.e., among-clone variances) for these traits and ten population-specific indices related to admixture levels (estimated based on 5165 SNPs), environmental temporal and spatial heterogeneity and climate harshness. Populations experiencing colder winters showed consistently lower genetic variation for early height growth (a fitness-related trait in forest trees) in the three common gardens. Within-population quantitative genetic variation was not associated with environmental heterogeneity or population admixture for any trait. Our results provide empirical support for the potential role of natural selection in reducing genetic variation for early height growth within populations, which indirectly gives insight into the adaptive potential of populations to changing environments.

Summary In a context of climate change, it is necessary to decipher the strategies established by plants to cope with limited water supply. Transcriptome, methylome and small RNA data were generated for two oak species with contrasting levels of drought tolerance ( Quercus robur and Quercus petraea ), under control and drought stress conditions All data are in line with a species-specific response to drought stress consistent with their ecological preferences. The biological processes associated with genomic regions identified in all datasets were mainly associated with parietal processes in Q. petraea, which may explain in part its better tolerance to water deprivation. A significant proportion of DNA methylation differences observed in control conditions between the two oak species were maintained during DS which may constitute a pool of epigenetic markers discriminating these two oak species. These markers were enriched in highly differentiated SNPs suggesting that some of them may be associated both with the ecological differences or intrinsic barriers to reproduction between the two species. An integrative approach of the three datasets revealed genomic co-locations of potential importance for forest three adaptation to drought stress.

Specialized or secondary metabolites play a key role in plant resistance against abiotic stresses and defences against bioaggressors. For example, in sessile oaks Quercus petraea , phenolics contribute to reduce herbivore damage and improve drought resistance. Here, we explored the natural variation of specialized metabolites in nine European provenances of sessile oaks and aimed to detect its underlying genetic bases. We sampled mature leaves from high and low branches on 225 sessile oak trees located in a common garden and used untargeted metabolomics to characterise the variation of 217 specialized metabolites. In addition, we used whole genome low-depth sequencing to genotype individuals for 1.4M genetic markers. We found that leaf specialized metabolites displayed extensive within-provenance variation, but very little differentiation between provenances. In addition, a genome-wide association study allowed detecting significant associations for 42% of these metabolites. Hence, our results suggest that genetic variation for most leaf specialized metabolites is unlikely to be locally adaptive, however lack of differentiation among populations suggests selection acts locally to maintain diversity at loci associated with leaf specialized metabolites variation.

Drought and waterlogging impede tree growth and may even lead to tree death. Oaks, an emblematic group of tree species, have evolved a range of adaptations to cope with these constraints. The two most widely distributed European species, pedunculate (PO; Quercus robur L.) and sessile oak (SO; Quercus petraea Matt. Lieb), have overlapping ranges, but their respective distribution are highly constrained by local soil conditions. These contrasting ecological preferences between two closely related and frequently hybridizing species constitute a powerful model to explore the functional bases of the adaptive responses in oak. We exposed oak seedlings to waterlogging and drought, conditions typically encountered by the two species in their respective habitats, and studied changes in gene expression in roots using RNA-seq. We identified genes that change in expression between treatments differentially depending on species. These "species × environment"-responsive genes revealed adaptive molecular strategies involving adventitious and lateral root formation, aerenchyma formation in PO, and osmoregulation and ABA regulation in SO. With this experimental design, we also identified genes with different expression between species independently of water conditions imposed. Surprisingly, this category included genes with functions consistent with a role in intrinsic reproductive barriers. Finally, we compared our findings with those for a genome scan of species divergence and found that the expressional candidate genes included numerous highly differentiated genetic markers between the two species. By combining transcriptomic analysis, gene annotation, pathway analyses, as well as genome scan for genetic differentiation among species, we were able to highlight loci likely involved in adaptation of the two species to their respective ecological niches.

Although complex interactions between hosts and microbial associates are increasingly well documented, we still know little about how and why hosts shape microbial communities in nature. In addition, host genetic effects on microbial communities vary widely depending on the environment, obscuring conclusions about which microbes are impacted and which plant functions are important. We characterized the leaf microbiota of 200 Arabidopsis thaliana genotypes in eight field experiments and detected consistent host effects on specific, broadly distributed microbial species (operational taxonomic unit [OTUs]). Host genetic effects disproportionately influenced central ecological hubs, with heritability of particular OTUs declining with their distance from the nearest hub within the microbial network. These host effects could reflect either OTUs preferentially associating with specific genotypes or differential microbial success within them. Host genetics associated with microbial hubs explained over 10% of the variation in lifetime seed production among host genotypes across sites and years. We successfully cultured one of these microbial hubs and demonstrated its growth-promoting effects on plants in sterile conditions. Finally, genome-wide association mapping identified many putatively causal genes with small effects on the relative abundance of microbial hubs across sites and years, and these genes were enriched for those involved in the synthesis of specialized metabolites, auxins, and the immune system. Using untargeted metabolomics, we corroborate the consistent association between variation in specialized metabolites and microbial hubs across field sites. Together, our results reveal that host genetic variation impacts the microbial communities in consistent ways across environments and that these effects contribute to fitness variation among host genotypes.

Drought and waterlogging impede tree growth and may even lead to tree death. With climate change, these environmental factors are a growing source of concern, particularly for temperate forests. Oaks, an emblematic group of tree species, have evolved a range of adaptations to cope with these constraints. The two most widely distributed European species — pedunculate oak (PO) and sessile oaks (SO) — have overlapping ranges, but are highly constrained locally by soil water content variation. These differences in local ecological requirements provide a powerful biological model for studying the role of ecological barriers in speciation. We used an experimental set-up mimicking the ecological preferences of these species, in which seedlings were subjected to waterlogging and drought. We studied gene expression in roots by RNA-seq and identified genes differentially expressed between treatments with different outcomes depending on species. These “species x environment”-responsive genes revealed adaptive molecular strategies involving adventitious and lateral root formation, aerenchyma formation in PO, and osmoregulation and ABA regulation in SO. With this experimental design, we also identified genes with expression profiles presenting a “species” effect regardless of imposed constraints with important roles in intrinsic reproductive barriers. Finally, we compared our findings with those for a genome scan of species divergence and found that the candidate genes were enriched in highly differentiated SNPs. This suggests that many of the genes involved in the contrasting transcriptomic responses are subject to natural selection and that gene regulation helps to maintain these two different oak species in sympatry.

Microbial communities are influenced by a complex system of host effects, including traits involved in physical barriers, immunity, hormones, metabolisms and nutrient homeostasis. Variation of host control within species is governed by many genes of small effect and is sensitive to biotic and abiotic environments. On the flip side, these host impacts seem targeted on particular microbial species, with that impact percolating through the microbial community. There is not yet evidence that the nature and strength of these interactions differs between fungal and bacterial communities, or among different compartments of the plant. The challenge of deciphering how systems of host traits impact systems of microbial associates is vast but holds promise for developing novel strategies to improve plant health.

How evolutionary forces interact to maintain quantitative genetic variation within populations has been a matter of extensive theoretical debates. While mutation and migration increase genetic variation, natural selection and genetic drift are expected to deplete it. To date, levels of genetic variation observed in natural populations are hard to predict without accounting for other processes, such as balancing selection in heterogeneous environments. We aimed to empirically test three hypotheses: (i) admixed populations have higher quantitative genetic variation due to introgression from other gene pools, (ii) quantitative genetic variation is lower in populations from harsher environments (i.e. experiencing stronger selection), and (iii) quantitative genetic variation is higher in populations from spatially heterogeneous environments. We used phenotypic measurements of five growth, phenological and functional traits from three clonal common gardens, consisting of 523 clones from 33 populations of maritime pine ( Pinus pinaster Aiton). Populations from harsher climates (mainly colder areas) showed lower genetic variation for height in the three common gardens. Surprisingly, we did not find any association between within-population genetic variation and environmental heterogeneity or population admixture for any trait. Our results suggest a predominant role of natural selection in driving within-population genetic variation, and therefore indirectly their adaptive potential.

Exotic pathogens cause severe damage in natural populations in the absence of coevolutionary dynamics with their hosts. However, some resistance to such pathogens may occur in naive populations. The objective of this study was to investigate the genetics of this so-called 'exapted' resistance to two pathogens of Asian origin (Erysiphe alphitoides and Phytophthora cinnamomi) in European oak. Host-pathogen compatibility was assessed by recording infection success and pathogen growth in a full-sib family of Quercus robur under controlled and natural conditions. Two high-resolution genetic maps anchored on the reference genome were used to study the genetic architecture of resistance and to identify positional candidate genes. Two genomic regions, each containing six strong and stable quantitative trait loci (QTLs) accounting for 12-19% of the phenotypic variation, were mainly associated with E. alphitoides infection. Candidate genes, especially genes encoding receptor-like-kinases and galactinol synthases, were identified in these regions. The three QTLs associated with P. cinnamomi infection did not colocate with QTLs found for E. alphitoides. These findings provide evidence that exapted resistance to E. alphitoides and P. cinnamomi is present in Q. robur and suggest that the underlying molecular mechanisms involve genes encoding proteins with extracellular signaling functions.

Environmental sequencing shows that plants harbor complex communities of microbes that vary across environments. However, many approaches for mapping plant genetic variation to microbe-related traits were developed in the relatively simple context of binary host-microbe interactions under controlled conditions. Recent advances in sequencing and statistics make genome-wide association studies (GWAS) an increasingly promising approach for identifying the plant genetic variation associated with microbes in a community context. This review discusses early efforts on GWAS of the plant phyllosphere microbiome and the outlook for future studies based on human microbiome GWAS. A workflow for GWAS of the phyllosphere microbiome is then presented, with particular attention to how perspectives on the mechanisms, evolution and environmental dependence of plant-microbe interactions will influence the choice of traits to be mapped.

The Hengduan Mountains (HDM) biodiversity hotspot exhibits exceptional alpine plant diversity. Here, we investigate factors driving intraspecific divergence within a HDM alpine species Salix brachista (Cushion willow), a common component of subnival assemblages. We produce a high-quality genome assembly for this species and characterize its genetic diversity, population structure and pattern of evolution by resequencing individuals collected across its distribution. We detect population divergence that has been shaped by a landscape of isolated sky island-like habitats displaying strong environmental heterogeneity across elevational gradients, combined with population size fluctuations that have occurred since approximately the late Miocene. These factors are likely important drivers of intraspecific divergence within Cushion willow and possibly other alpine plants with a similar distribution. Since intraspecific divergence is often the first step toward speciation, the same factors can be important contributors to the high alpine species diversity in the HDM.

Genetic structure in broadly distributed herbs may be shaped by past climate events, leading to distinct genetic lineages, and possibly to different Evolutionary Significant Units (ESUs). We identified the potential glacial refugia, postglacial (re)colonization routes, contact zones and ESUs in the widespread but locally rare Silene nutans to propose appropriate conservation strategies related to migration history-related genetic structure. We sequenced four plastid DNA markers from 136 populations covering S. nutans European distribution range. We built a network on concatenated sequences using statistical parsimony and performed Bayesian and Maximum Likelihood analyses to build a phylogenetic tree. Haplotype genetic diversity and between-population differentiation (G(ST), N-ST) were calculated. Based on 23 plastid haplotypes, populations were ascribed to seven strongly differentiated genetic lineages. Potential refugia were identified: in the Iberian and Italian Peninsulas, expanding to south-western France and the Alps, respectively; in south-central France, reaching England and Belgium; an eastern refugium, following a north-westward expansion; in the Balkans, expanding to north-eastern Italy, and in the Carpathians. Several contact zones were identified. Silene nutans consists of a cryptic assemblage of genetically distinct taxa, with at least seven ESUs. Evaluating the distribution range, the abundance, and the conservation status of each ESU is necessary for the preservation of the S. nutans species complex at European and regional levels. Ex situ conservation and genetic restoration involving plant translocations must consider the phylogeographic history and the distinct genetic lineages to avoid the introduction of poorly adapted plants and outbreeding depression leading to non-viable hybrid progeny, especially in contact zones.

By following the evolution of populations that are initially genetically homogeneous, much can be learned about core biological principles. For example, it allows for detailed studies of the rate of emergence of de novo mutations and their change in frequency due to drift and selection. Unfortunately, in multicellular organisms with generation times of months or years, it is difficult to set up and carry out such experiments over many generations. An alternative is provided by "natural evolution experiments" that started from colonizations or invasions of new habitats by selfing lineages. With limited or missing gene flow from other lineages, new mutations and their effects can be easily detected. North America has been colonized in historic times by the plant Arabidopsis thaliana, and although multiple inter-crossing lineages are found today, many of the individuals belong to a single lineage, HPG1. To determine in this lineage the rate of substitutions-the subset of mutations that survived natural selection and drift-, we have sequenced genomes from plants collected between 1863 and 2006. We identified 73 modern and 27 herbarium specimens that belonged to HPG1. Using the estimated substitution rate, we infer that the last common HPG1 ancestor lived in the early 17 th century, when it was most likely introduced by chance from Europe. Mutations in coding regions are depleted in frequency compared to those in other portions of the genome, consistent with purifying selection. Nevertheless, a handful of mutations is found at high frequency in present-day populations. We link these to detectable phenotypic variance in traits of known ecological importance, life history and growth, which could reflect their adaptive value. Our work showcases how, by applying genomics methods to a combination of modern and historic samples from colonizing lineages, we can directly study new mutations and their potential evolutionary relevance.

Oaks are an important part of our natural and cultural heritage. Not only are they ubiquitous in our most common landscapes1 but they have also supplied human societies with invaluable services, including food and shelter, since prehistoric times2. With 450 species spread throughout Asia, Europe and America3, oaks constitute a critical global renewable resource. The longevity of oaks (several hundred years) probably underlies their emblematic cultural and historical importance. Such long-lived sessile organisms must persist in the face of a wide range of abiotic and biotic threats over their lifespans. We investigated the genomic features associated with such a long lifespan by sequencing, assembling and annotating the oak genome. We then used the growing number of whole-genome sequences for plants (including tree and herbaceous species) to investigate the parallel evolution of genomic characteristics potentially underpinning tree longevity. A further consequence of the long lifespan of trees is their accumulation of somatic mutations during mitotic divisions of stem cells present in the shoot apical meristems. Empirical4 and modelling5 approaches have shown that intra-organismal genetic heterogeneity can be selected for6 and provides direct fitness benefits in the arms race with short-lived pests and pathogens through a patchwork of intra-organismal phenotypes7. However, there is no clear proof that large-statured trees consist of a genetic mosaic of clonally distinct cell lineages within and between branches. Through this case study of oak, we demonstrate the accumulation and transmission of somatic mutations and the expansion of disease-resistance gene families in trees.

Research on plant-herbivore interactions has long recognized that plant genetic variation plays a central role in driving insect abundance and herbivory, as well as in determining plant defense. However, how plant genes influence herbivore feeding performances, and which plant defensive traits mediate these effects, remain poorly understood. Here we investigated the feeding performances of two insect leaf chewers with contrasting diet breadth (the generalist Lymantria dispar L. and the specialist Thaumetopoea processionea L.) on different genotypes of pedunculate oak (Quercus robur L.) and tested the role of leaf phenolics. We used leaves from four clones of 30 Q. robur full-sibs grown in a common garden to estimate the performance of both herbivores in laboratory feeding trials and to quantify the concentration of constitutive chemical defences (phenolic compounds). We found that tree genetics influenced leaf consumption by T. processionea but not by L. dispar. However genetic variation among trees did not explain growth rate variation in either herbivore nor in leaf phenolics. Interestingly, all phenolic compounds displayed a positive relationship with L. dispar growth rate, and leaf consumption by both herbivores displayed a positive relationship with the concentrations of condensed tannins, suggesting that highly defended leaves could induce a compensatory feeding response. While genetic variation in oaks did not explain herbivore growth rate, we found positive genetic correlations between the two herbivores for leaf consumption and digestion. Overall, we found that oak genotype and phenolic compounds partly and independently contribute to variability in herbivore performance. We challenged the current view of plant-insect interaction and provided little support to the idea that the effect of plant genotype on associated organisms is driven by plant defences. Together, our results point to the existence of genetically determined resistance traits in oaks whose effects differ between herbivores and motivate further research on mechanisms governing oak-herbivore interactions.

Although the complex interactions between hosts and microbial associates are increasingly well documented, we still know little about how and why hosts shape microbial communities in nature. We characterized the leaf microbiota within 200 clonal accessions in eight field experiments and detected effects of both local environment and host genotype on community structure. Within environments, hosts′ genetics preferentially associate with a core of ubiquitous microbial hubs that, in turn, structure the community. These microbial hubs correlate with host performance, and a GWAS revealed strong candidate genes for the host factors impacting heritable hubs. Our results reveal how selection may act to enhance fitness through microbial associations and bolster the possibility of enhancing crop performance through these host factors.

Although the complex interactions between hosts and microbial associates are increasingly well documented, we still know little about how and why hosts shape microbial communities in nature. We characterized the leaf microbiota within 200 clonal accessions in eight field experiments and detected effects of both local environment and host genotype on community structure. Within environments, hosts′ genetics preferentially associate with a core of ubiquitous microbial hubs that, in turn, structure the community. These microbial hubs correlate with host performance, and a GWAS revealed strong candidate genes for the host factors impacting heritable hubs. Our results reveal how selection may act to enhance fitness through microbial associations and bolster the possibility of enhancing crop performance through these host factors.