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Cultivated sunflower (Helianthus annuus L.) exhibits numerous phenotypic and transcriptomic responses to drought. However, the ways in which these responses vary with differences in drought timing and severity are insufficiently understood. We used phenotypic and transcriptomic data to evaluate the response of sunflower to drought scenarios of different timing and severity in a common garden experiment. Using a semi-automated outdoor high-throughput phenotyping platform, we grew six oilseed sunflower lines under control and drought conditions. Our results reveal that similar transcriptomic responses can have disparate phenotypic effects when triggered at different developmental time points. Leaf transcriptomic responses, however, share similarities despite timing and severity differences (e.g., 523 differentially expressed genes (DEGs) were shared across all treatments), though increased severity elicited greater differences in expression, particularly during vegetative growth. Across treatments, DEGs were highly enriched for genes related to photosynthesis and plastid maintenance. A co-expression analysis identified a single module (M8) enriched in all drought stress treatments. Genes related to drought, temperature, proline biosynthesis, and other stress responses were overrepresented in this module. In contrast to transcriptomic responses, phenotypic responses were largely divergent between early and late drought. Early-stressed sunflowers responded to drought with reduced overall growth, but became highly water-acquisitive during recovery irrigation, resulting in overcompensation (higher aboveground biomass and leaf area) and a greater overall shift in phenotypic correlations, whereas late-stressed sunflowers were smaller and more water use-efficient. Taken together, these results suggest that drought stress at an earlier growth stage elicits a change in development that enables greater uptake and transpiration of water during recovery, resulting in higher growth rates despite similar initial transcriptomic responses.

Floral morphology and pigmentation are both charismatic and economically relevant traits associated with cultivated sunflower (Helianthus annuus L.). Recent work has linked floral morphology and pigmentation to pollinator efficiency and seed yield. Understanding the genetic architecture of such traits is essential for crop improvement, and gives insight into the role of genetic constraints in shaping floral diversity. A diversity panel of 288 sunflower genotypes was phenotyped for a variety of morphological, phenological, and color traits in both a greenhouse and a field setting. Association mapping was performed using 5788 SNP markers using a mixed linear model approach. Several dozen markers across 10 linkage groups were significantly associated with variation in morphological and color trait variation. Substantial trait plasticity was observed between greenhouse and field phenotyping, and associations differed between environments. Color traits mapped more strongly than morphology in both settings, with markers together explaining 16% of petal carotenoid content in the greenhouse, and 17% and 24% of variation in disc anthocyanin presence in the field and greenhouse, respectively. Morphological traits like disc size mapped more strongly in the field, with markers together explaining up to 19% of disc size variation. Loci identified here through association mapping within cultivated germplasm differ from those identified through biparental crosses between modern cultivated sunflower and either its wild progenitor or domesticated landraces. Several loci lie within genomic regions involved in domestication. Differences between phenotype expression under greenhouse and field conditions highlight the importance of plasticity in determining floral morphology and pigmentation.

With rising food demands, crop production on salinized lands is increasingly necessary. Sunflower (Helianthus annuus), a moderately salt-tolerant crop, exhibits a tradeoff where more vigorous, high-performing genotypes have a greater proportional decline in biomass under salinity stress. Prior research has found deviations from this relationship across genotypes. Here, we identified the traits and genomic regions underlying variation in this expectation-deviation tolerance (the magnitude and direction of deviations from the expected effect of salinity). We grew a sunflower diversity panel under control and salt-stressed conditions and measured a suite of morphological (growth, mass allocation, plant and leaf morphology) and leaf ionomic traits. The genetic basis of variation and plasticity in these traits was investigated via genome-wide association, which also enabled the identification of genomic regions (i.e. haplotypic blocks) influencing multiple traits. We found that the magnitude and direction of plasticity in whole-root mass fraction, fine root mass fraction, and chlorophyll content, as well as leaf sodium and potassium content under saline conditions, were most strongly correlated with expectation-deviation tolerance. We identified multiple genomic regions underlying these traits as well as a single alpha-mannosidase gene directly associated with this tolerance metric. Our results show that, by taking the vigor-salinity effect tradeoff into account, we can identify unique traits and genes associated with salinity tolerance. Since these traits and genomic regions are distinct from those associated with high vigor (i.e. growth in benign conditions), they provide an avenue for increasing salinity tolerance in high-performing sunflower genotypes without compromising vigor.

Cultivated crops are expected to be less stress tolerant than their wild relatives, leading to efforts to mine wild relatives for traits to increase crop tolerance. However, empirical tests of this expectation often confound tolerance with plant vigor. We assessed whether wild and cultivated Helianthus annuus L. differed for salinity tolerance with 0 and 150 mM NaCl treatments. Salinity tolerance was assessed as the proportional reduction in biomass and as the deviation from expected performance based on vigor. Cultivated accessions had a greater proportional decline in biomass than wild accessions, but proportional decline was positively associated with vigor in both. Thus, wild and cultivated H. annuus did not differ for tolerance when variation in vigor was corrected for statistically. For traits potentially related to tolerance mechanisms, wild and cultivated accessions differed for elemental content and allocation of N, P, K, Mg, Ca, S, Na, Fe, Mn, B, Cu, and Zn for some tissues, biomass allocation, specific leaf area, and leaf succulence. However, these traits were generally unrelated to tolerance corrected for vigor. Osmotic adjustment was associated with tolerance corrected for vigor only in wild accessions where more osmotic adjustment was associated with greater tolerance. Our results for H. annuus suggest that efforts to use wild relatives to enhance crop abiotic stress tolerance will benefit from greater knowledge of traits related to plant growth responses decoupled from vigor, in order to get beyond potential growth-tolerance trade-offs.

With rising food demands, crop production on salinized lands is increasingly necessary. Sunflower, a moderately salt tolerant crop, exhibits a trade-off where more vigorous, high-performing genotypes have a greater proportional decline in biomass under salinity stress. Prior research has found deviations from this relationship across genotypes; the magnitude and direction of these deviations provides a useful metric of tolerance. Here, we identified the traits and genomic regions underlying variation in this expectation-deviation tolerance. We grew a diversity panel under control and salt-stressed conditions and measured a suite of morphological (growth, allocation, plant and leaf morphology) and leaf ionomic traits. The genetic basis of variation in these traits and their plasticity was investigated via genome-wide association studies, which also enabled the identification of genomic regions (i.e., haplotypic blocks) influencing multiple traits. We found that the magnitude of plasticity in whole root mass fraction, fine root mass fraction, and chlorophyll content, as well as leaf Na and K content under saline conditions, were the traits most strongly correlated with expectation-deviation tolerance. Additionally, we identified multiple genomic regions underlying these traits as well as a single gene directly associated with this tolerance metric. Our results show that, by taking the vigor-salinity effect trade-off into account, we can identify unique traits and genes associated with salinity tolerance. Since these traits and genomic regions are distinct from those associated with high vigor (i.e., growth in benign conditions), they provide an avenue for increasing salinity tolerance in high-performing sunflower genotypes without compromising vigor.Single sentence summaryDespite a trade-off between vigor and salinity-induced decline in biomass, distinct traits and genomic regions exist that could modulate this trade-off in cultivated sunflower.

Species often include multiple ecotypes that are adapted to different environments 1. However, it is unclear how ecotypes arise and how their distinctive combinations of adaptive alleles are maintained despite hybridization with non-adapted populations 2-4. Here, by resequencing 1,506 wild sunflowers from 3 species (Helianthus annuus, Helianthus petiolaris and Helianthus argophyllus), we identify 37 large (1-100 Mbp in size), non-recombining haplotype blocks that are associated with numerous ecologically relevant traits, as well as soil and climate characteristics. Limited recombination in these haplotype blocks keeps adaptive alleles together, and these regions differentiate sunflower ecotypes. For example, haplotype blocks control a 77-day difference in flowering between ecotypes of the silverleaf sunflower H. argophyllus (probably through deletion of a homologue of FLOWERING LOCUS T (FT)), and are associated with seed size, flowering time and soil fertility in dune-adapted sunflowers. These haplotypes are highly divergent, frequently associated with structural variants and often appear to represent introgressions from other-possibly now-extinct-congeners. These results highlight a pervasive role of structural variation in ecotypic adaptation. Local adaptation is common in species that experience different environments across their range, often resulting in the formation of ecotypes-ecological races with distinct morphological and/or physiological characteristics that provide an environment-specific fitness advantage. Despite the prevalence of ecotypic differentiation, much remains to be understood about the genetic basis and evolutionary mechanisms that underlie its establishment and maintenance. In particular , a longstanding evolutionary question-dating to criticisms of Darwin's theories by his contemporaries 4-concerns how such ecological divergence can occur when challenged by hybridization with non-adapted populations 2. Local adaptation typically requires alleles at multiple loci that contribute to increased fitness in the same environment ; however, different ecotypes are often geographically close and interfertile, and hybridization between them should break up adaptive allelic combinations 3. To better understand the genetic basis of local adaptation and ecotypic differentiation, we conducted an in-depth study of genetic, phenotypic and environmental variation in three annual sunflower species, each of which includes multiple reproductively compatible ecotypes. Two species (H. annuus and H. petiolaris) have broad, overlapping distributions across North America. Helianthus annuus, the common sunflower, is generally found on mesic soils, but can grow in a variety of disturbed or extreme habitats, including semi-desert or frequently flooded areas. An especially well-characterized ecotype (formally known as H. annuus subsp. texanus) is adapted to the higher temperatures and herbivore pressures in Texas (USA) 5. Helianthus petiolaris, the prairie sunflower, prefers sandier soils; ecotypes of this species are adapted to sand sheets and dunes 6. The third species-H. argophyllus, the silverleaf sunflower-is endemic to southern Texas and includes both an early-flowering, coastal-island ecotype and a late-flowering inland ecotype 7. Population structure of wild sunflowers In a common garden experiment, we grew 10 plants from each of 151 populations of the 3 species, selected from across their native range (Fig. 1a); for each of these populations, we collected corresponding soil samples. We generated extensive records of developmental and morphological traits, and resequenced the genomes of 1,401 individual plants. We resequenced an additional 105 H. annuus plants to fill gaps in geographical coverage, as well as 12 outgroup taxa (Supplementary Table 1). Sunflower genomes are relatively large (H. annuus, 3.5 Gbp; https://doi.

Foliar nutrient resorption is a key modulator of plant nutrient use. However, evolutionary patterns for nutrient resorption remain unclear, especially in herbs. We measured nitrogen and phosphorus resorption on preselected leaves across the Helianthus (sunflower) genus in a common garden in Athens, GA. We analyzed our data with published leaf traits and native habitat environmental data. Using phylogenetically controlled analyses, we tested if (1) nutrient resorption correlates with leaf economic, vasculature, and defense traits through evolutionary time, and (2) native habitat environment predicts nutrient resorption evolution. For Helianthus, nutrient resorption capacity is greater in resource-conservative species, as previously defined for Helianthus with a principle components analysis of leaf economic spectrum traits (photosynthetic rate, respiration rate, leaf lifespan, leaf mass per area, and green-leaf N and P concentrations). Nutrient resorption capacity also evolutionarily correlates with individual leaf economic traits, though not always as expected based on broad species surveys. Greater nutrient resorption is also positively associated with leaf chemical defenses, but not leaf vasculature or senescence rate. Finally, nitrogen resorption evolution increases with native habitat precipitation, but native habitat soil fertility does not predict nitrogen or phosphorus resorption. Our results suggest nutrient resorption evolution is more closely tied to resource economic strategy than native habitat.

Background There is a growing appreciation that the evolution of floral traits is shaped by selection not only from pollinators, but also factors like abiotic stress, florivory, and disease. Many suc

Background and Aims Trait-based plant ecology attempts to use small numbers of functional traits to predict plant ecological strategies. However, a major gap exists between our understanding of organ-level ecophysiological traits and our understanding of whole-plant fitness and environmental adaptation. In this gap lie whole-plant organizational traits, including those that describe how plant biomass is allocated among organs and the timing of plant reproduction. This study explores the role of whole-plant organizational traits in adaptation to diverse environments in the context of life history, growth form and leaf economic strategy in a well-studied herbaceous system. Methods A phylogenetic comparative approach was used in conjunction with common garden phenotyping to assess the evolution of biomass allocation and reproductive timing across 83 populations of 27 species of the diverse genus Helianthus (the sunflowers). Key Results Broad diversity exists among species in both relative biomass allocation and reproductive timing. Early reproduction is strongly associated with resource-acquisitive leaf economic strategy, while biomass allocation is less integrated with either reproductive timing or leaf economics. Both biomass allocation and reproductive timing are strongly related to source site environmental characteristics, including length of the growing season, temperature, precipitation and soil fertility. Conclusions Herbaceous taxa can adapt to diverse environments in many ways, including modulation of phenology, plant architecture and organ-level ecophysiology. Although leaf economic strategy captures one key aspect of plant physiology, on their own leaf traits are not particularly predictive of ecological strategies in Helianthus outside of the context of growth form, life history and whole-plant organization. These results highlight the importance of including data on whole-plant organization alongside organ-level ecophysiological traits when attempting to bridge the gap between functional traits and plant fitness and environmental adaptation.

Disruption of ion homeostasis is a major component of salinity stress's effect on crop yield. In cultivated sunflower prior work revealed a negative relationship between vigor and salinity tolerance. Here, we determined the association of elemental content/distribution traits with salinity tolerance, both with and without taking vigor (biomass in control treatment) into account. We grew seedlings of 12 Helianthus annuus genotypes in two treatments (0, 100 mM NaCl). Plants were measured for biomass (+allocation), and element content (Na, P, K, Ca, Mg, S, Fe, B, Mn, Cu, Zn) in leaves (young and mature), stem, and roots. Genotype tolerance was assessed as both proportional decline of biomass and as expectation deviation (deviation from the observed relationship between vigor and proportional decline in biomass). Genotype rankings on these metrics were not the same. Elemental content and allocation/distribution were highly correlated both at the plant and organ level. Suggestive associations between tolerance and elemental traits were fewer and weaker than expected and differed by tolerance metric. Given the highly correlated nature of elemental content, it remains difficult to pinpoint specific traits underpinning tolerance. Results do show that taking vigor into account is important when seeking to determining traits that can be targeted to increase tolerance independent of vigor, and that the multivariate nature of associated traits should additionally be considered.

Species often include multiple ecotypes that are adapted to different environments. But how do ecotypes arise, and how are their distinctive combinations of adaptive alleles maintained despite hybridization with non-adapted populations? Re-sequencing of 1506 wild sunflowers from three species identified 37 large (1-100 Mbp), non-recombining haplotype blocks associated with numerous ecologically relevant traits, and soil and climate characteristics. Limited recombination in these regions keeps adaptive alleles together, and we find that they differentiate several sunflower ecotypes; for example, they control a 77 day difference in flowering between ecotypes of silverleaf sunflower (likely through deletion of a FLOWERING LOCUS T homolog), and are associated with seed size, flowering time and soil fertility in dune-adapted sunflowers. These haplotypes are highly divergent, associated with polymorphic structural variants, and often appear to represent introgressions from other, possibly extinct, congeners. This work highlights a pervasive role of structural variation in maintaining complex ecotypic adaptation.

Abiotic stress limits crop productivity worldwide, making stress-resistant crops a major goal for improving food security. Stress resistance is often assessed by comparing genotypes under a single control and stressed level of an abiotic factor, then ranking the genotypes’ overall performance, using a metric like geometric mean productivity (GMP). However, abiotic stress is rarely a presence/absence phenomenon, but often varies continuously. Here, we tested whether the common practice of assessing performance using a single control and stress level is predictive of performance across a range of stress levels. Twelve sunflower genotypes were grown in the greenhouse under one of eight nutrient levels (from non-limiting to severely limiting) and assessed for performance (shoot dry mass at the budding stage) and related traits at both seedling and budding stages. Contrary to expectations, genotype rankings for GMP were consistent regardless of the two nutrient levels used to calculate GMP. This suggests that, for nutrient stress in sunflower, studies using an arbitrary control and stress level are broadly representative: genotypes with high GMP exhibit high GMP regardless of the nutrient levels used. In addition, GMP values were highly correlated with performance in the respective nutrient levels from which they were calculated; thus, higher GMP for a given genotype indicates that it performed well (relative to other genotypes) under both stressed and non-stressed conditions. Together, these findings strongly support the current paradigm of comparing genotypes at a single control and stress level as a valid strategy for screening for nutrient stress resistance in sunflower. However, few traits were consistently associated with performance (shoot dry mass) at either high- or low-nutrient availability, and seedling traits were rarely predictive of budding-stage traits and performance, suggesting further studies are needed to determine the mechanisms underlying nutrient stress resistance in this important oilseed crop.

Recent work suggests variation in plant growth strategies is governed by a tradeoff in resource acquisition and use, ranging from a rapid resource acquisition strategy to a resource‐conservative strategy. While evidence for this tradeoff has been found in leaves, knowledge of root trait strategies, and whether they reflect adaptive differentiation across environments, is limited. In the greenhouse, we investigated variation in fine root morphology (specific root length and tissue density), chemistry (nitrogen concentration and carbon:nitrogen), and anatomy (root cross‐sectional traits) in populations of 26 Helianthus species and sister Phoebanthus tenuifolius. We also compared root trait variation in this study with leaf trait variation previously reported in a parallel study of these populations. Root traits varied widely and exhibited little phylogenetic signal, suggesting high evolutionary lability. Specific root length and root tissue density were weakly negatively correlated, but neither was associated with root nitrogen, providing little support for a single axis of root trait covariation. Correlations between traits measured in the greenhouse and native site characteristics were generally weak, suggesting a variety of equally viable root trait combinations exist within and across environments. However, high root nitrogen was associated with lower xylem vessel number and cross‐sectional area, suggesting a tradeoff between nutrient investment and water transport capacity. This led to correlations between root and leaf traits that were not always consistent with an acquisition–conservation tradeoff at the whole‐plant level. Given that roots must balance acquisition of water and nutrients with functions like anchorage, exudation, and microbial symbioses, the varied evidence for root trait covariation likely reflects the complexity of interacting selection pressures belowground. Similarly, the lack of evidence for a single acquisition–conservation tradeoff at the whole‐plant level likely reflects the vastly different selection pressures shaping roots and leaves, and the resources they are optimized to obtain.

Developing more stress-tolerant crops will require greater knowledge of the physiological basis of stress tolerance. Here we explore how biomass declines in response to salinity relate to leaf traits across twenty genotypes of cultivated sunflower (Helianthus annuus). Plant growth, leaf physiological traits, and leaf elemental composition were assessed after 21 days of salinity treatments (0, 50, 100, 150, or 200 mM NaCl) in a greenhouse study. There was a trade-off in performance such that vigorous genotypes, those with higher biomass at zero mM NaCl, had both a larger absolute decrease and proportional decrease in biomass due to increased salinity. More vigorous genotypes at control were less tolerant to salinity. Contrary to expectation, genotypes with a low increase in leaf Na and decrease in K:Na were not better at maintaining biomass with increasing salinity. Rather, genotypes with a greater reduction in leaf S and K content were better at maintaining biomass at increased salinity. While we found an overall trade-off between sunflower vigour and salt tolerance, some genotypes were more tolerant than expected. Further analysis of the traits and mechanisms underlying this trade-off may allow us to breed these into high vigour genotypes in order to increase their salt tolerance.

Leaf defenses are widely recognized as key adaptations and drivers of plant evolution. Across environmentally diverse habitats, the macroevolution of leaf defenses can be predicted by the univariate trade-off model, which predicts that defenses are functionally redundant and thus trade off, and the resource availability hypothesis, which predicts that defense investment is determined by inherent growth rate and that higher defense will evolve in lower resource environments. Here, we examined the evolution of leaf physical and chemical defenses and secondary metabolites in relation to environmental characteristics and leaf economic strategy across 28 species of Helianthus (the sunflowers). Using a phylogenetic comparative approach, we found few evolutionary trade-offs among defenses and no evidence for defense syndromes. We also found that leaf defenses are strongly related to leaf economic strategy, with higher defense in more resource-conservative species, although there is little support for the evolution of higher defense in low-resource habitats. A wide variety of physical and chemical defenses predict resistance to different insect herbivores, fungal pathogens, and a parasitic plant, suggesting that most sunflower defenses are not redundant in function and that wild Helianthus represents a rich source of variation for the improvement of crop sunflower.

The leaf economics spectrum (LES) is a prominent ecophysiological paradigm that describes global variation in leaf physiology across plant ecological strategies using a handful of key traits. Nearly a decade ago, Shipley et al. (2006) used structural equation modelling to explore the causal functional relationships among LES traits that give rise to their strong global covariation. They concluded that an unmeasured trait drives LES covariation, sparking efforts to identify the latent physiological trait underlying the 'origin' of the LES. Here, we use newly developed phylogenetic structural equation modelling approaches to reassess these conclusions using both global LES data as well as data collected across scales in the genus Helianthus. For global LES data, accounting for phylogenetic non-independence indicates that no additional unmeasured traits are required to explain LES covariation. Across datasets in Helianthus, trait relationships are highly variable, indicating that global-scale models may poorly describe LES covariation at non-global scales.

The leaf economics spectrum (LES) describes a major axis of plant functional trait variation worldwide, defining suites of leaf traits aligned with resource-acquisitive to resource-conservative ecological strategies. The LES has been interpreted to arise from leaf-level trade-offs among ecophysiological traits common to all plants. However, it has been suggested that the defining leaf-level trade-offs of the LES may not hold within specific functional groups (e.g., herbs) nor within many groups of closely related species, which challenges the usefulness of the LES paradigm across evolutionary scales. Here, we examine the evolution of the LES across 28 species of the diverse herbaceous genus Helianthus (the sunflowers), which occupies a wide range of habitats and climate variation across North America. Using a phylogenetic comparative approach, we find repeated evolution of more resource-acquisitive LES strategies in cooler, drier, and more fertile environments. We also find macroevolutionary correlations among LES traits that recapitulate aspects of the global LES, but with one major difference: leaf mass per area is uncorrelated with leaf lifespan. This indicates that whole-plant processes likely drive variation in leaf lifespan across Helianthus, rather than leaf-level trade-offs. These results suggest that LES patterns do not reflect universal physiological trade-offs at small evolutionary scales.

Summary 1. Efforts to understand the effects of plant traits on carbon and nutrient cycling have recently focused on species variation and the potential for species data to improve predictions of past, present and future variation in ecosystems. However, the evolutionary lability of relevant traits among closely related species and the extent of intraspecific variation warrant further consideration. 2. Here, we examine interspecific and intraspecific variation in leaf N, LMA, root N and SRL at multiple scales, using Helianthus as a representative study system. 3. Substantial evolutionary lability of traits is demonstrated by interspecific variation in phylogenetically explicit analyses of closely related Helianthus species, and population differentiation of wild H. anomalus and cultivated H. annuus. 4. Intraspecific variation in leaf N and LMA, including genetic, environmental and ontogenetic responses, demonstrates that trait values for a single species can encompass a surprisingly large portion of the range encompassed by species in the leaf economics spectrum. 5. Synthesis. We recommend using data from selected natural populations to model effects of leaf N and LMA on decomposition, while using data from common garden experiments to determine evolutionary lability and thus inform potential for evolutionary change. If the high evolutionary lability of traits demonstrated for Helianthus is found for other important genera, this would suggest that these key ecophysiological traits are likely to respond to the selective pressures of global climate and land-use change.

Most C3 plant species have partially open stomata during the night especially in the 3-5 h before dawn. This pre-dawn stomatal opening has been hypothesized to enhance early-morning photosynthesis (A) by reducing diffusion limitations to CO2 at dawn. We tested this hypothesis in cultivated Helianthus annuus using whole-shoot gas exchange, leaf level gas exchange and modelling approaches. One hour pre-dawn low-humidity treatments were used to reduce pre-dawn stomatal conductance (g). At the whole-shoot level, a difference of pre-dawn g (0.40 versus 0.17 mol m(-2) s(-1)) did not significantly affect A during the first hour after dawn. Shorter term effects were investigated with leaf level gas exchange measurements and a difference of pre-dawn g (0.10 versus 0.04 mol m(-2) s(-1)) affected g and A for only 5 min after dawn. The potential effects of a wider range of stomatal apertures were explored with an empirical model of the relationship between A and intercellular CO2 concentration during the half-hour after dawn. Modelling results demonstrated that even extremely low pre-dawn stomatal conductance values have only a minimal effect on early-morning A for a few minutes after dawn. Thus, we found no evidence that pre-dawn stomatal opening enhances A.

• Plant resource-use traits are generally hypothesized to be adaptively differentiated for populations distributed along resource gradients. Although nutrient limitations are expected to select for resource-conservative strategies, water limitations may select for either resource-conservative or -acquisitive strategies. We test whether population differentiation reflects local adaptation for traits associated with resource-use strategies in a desert annual (Helianthus anomalus) distributed along a gradient of positively covarying water and nutrient availability. • We compared quantitative trait variation (Q(ST)) with neutral genetic differentiation (F(ST)), in a common garden glasshouse study, for leaf economics spectrum (LES) and related traits: photosynthesis (A(mass), A(area)), leaf nitrogen (N(mass), N(area)), leaf lifetime (LL), leaf mass per area (LMA), leaf water content (LWC), water-use efficiency (WUE, estimated as δ(13)C) and days to first flower (DFF). • Q(ST)-F(ST) differences support adaptive differentiation for Amass , N(mass), N(area), LWC and DFF. The trait combinations associated with drier and lower fertility sites represent correlated trait evolution consistent with the more resource-acquisitive end of the LES. There was no evidence for adaptive differentiation for A(area), LMA and WUE. • These results demonstrate that hot dry environments can selectively favor correlated evolution of traits contributing to a resource-acquisitive and earlier reproduction 'escape' strategy, despite lower fertility.

Leaf vein traits are implicated in the determination of gas exchange rates and plant performance. These traits are increasingly considered as causal factors affecting the 'leaf economic spectrum' (LES), which includes the light-saturated rate of photosynthesis, dark respiration, foliar nitrogen concentration, leaf dry mass per area (LMA) and leaf longevity. This article reviews the support for two contrasting hypotheses regarding a key vein trait, vein length per unit leaf area (VLA). Recently, Blonder et al. (2011, 2013) proposed that vein traits, including VLA, can be described as the 'origin' of the LES by structurally determining LMA and leaf thickness, and thereby vein traits would predict LES traits according to specific equations. Careful re-examination of leaf anatomy, published datasets, and a newly compiled global database for diverse species did not support the 'vein origin' hypothesis, and moreover showed that the apparent power of those equations to predict LES traits arose from circularity. This review provides a 'flux trait network' hypothesis for the effects of vein traits on the LES and on plant performance, based on a synthesis of the previous literature. According to this hypothesis, VLA, while virtually independent of LMA, strongly influences hydraulic conductance, and thus stomatal conductance and photosynthetic rate. We also review (i) the specific physiological roles of VLA; (ii) the role of leaf major veins in influencing LES traits; and (iii) the role of VLA in determining photosynthetic rate per leaf dry mass and plant relative growth rate. A clear understanding of leaf vein traits provides a new perspective on plant function independently of the LES and can enhance the ability to explain and predict whole plant performance under dynamic conditions, with applications towards breeding improved crop varieties.

Granite outcrops in the southeastern United States are rare and isolated habitats that support edaphically controlled communities dominated by herbaceous plants. They harbor rare and endemic species that are expected to have low genetic variability and high population structure due to small population sizes and their disjunct habitat. We test this expectation for an annual outcrop endemic, Helianthus porteri (Porter's sunflower). Contrary to expectation, H. porteri has relatively high genetic diversity (H e = 0.681) and relatively low genetic structure among the native populations (F ST = 0.077) when compared to 5 other Helianthus species (N = 288; 18 expressed sequence tag-SSR markers). These findings suggest greater gene flow than expected. The potential for gene flow is supported by the analysis of transplant populations established with propagules from a common source in 1959. One population established close to a native population (1.5 km) at the edge of the natural range is genetically similar to and shares rare alleles with the adjacent native population and is distinct from the central source population. In contrast, a transplant population established north of the native range has remained similar to the source population. The relatively high genetic diversity and low population structure of this species, combined with the long-term success of transplanted populations, bode well for its persistence as long as the habitat persists.

Premise of the study Patterns of plant stem traits are expected to align with a "fast-slow" plant economic spectrum across taxa. Although broad patterns support such tradeoffs in field studies, tests of hypothesized correlated trait evolution and adaptive differentiation are more robust when taxa relatedness and environment are taken into consideration. Here we test for correlated evolution of stem and leaf traits and their adaptive differentiation across environments in the herbaceous genus, Helianthus. Methods Stem and leaf traits of 14 species of Helianthus (28 populations) were assessed in a common garden greenhouse study. Phylogenetically independent contrasts were used to test for evidence of correlated evolution of stem hydraulic and biomechanical properties, correlated evolution of stem and leaf traits, and adaptive differentiation associated with source habitat environments. Key results Among stem traits, there was evidence for correlated evolution of some hydraulic and biomechanical properties, supporting an expected tradeoff between stem theoretical hydraulic efficiency and resistance to bending stress. Population differentiation for suites of stem and leaf traits was found to be consistent with a "fast-slow" resource-use axis for traits related to water transport and use. Associations of population traits with source habitat characteristics supported repeated evolution of a resource-acquisitive "drought-escape" strategy in arid environments. Conclusions This study provides evidence of correlated evolution of stem and leaf traits consistent with the fast-slow spectrum of trait combinations related to water transport and use along the stem-to-leaf pathway. Correlations of traits with source habitat characteristics further indicate that the correlated evolution is associated, at least in part, with adaptive differentiation of Helianthus populations among native habitats differing in climate.

Plant roots exude numerous metabolites into the soil that influence nutrient availability. Although root exudate composition is hypothesized to be under selection in low fertility soils, few studies have tested this hypothesis in a phylogenetic framework. In this study, we examined root exudates of three pairs of Helianthus species chosen as phylogenetically-independent contrasts with respect to native soil nutrient availability. Under controlled environmental conditions, seedlings were grown to the three-leaf-pair stage, then transferred to either high or low nutrient treatments. After five days of nutrient treatments, we used gas chromatography-mass spectrometry for analysis of root exudates, and detected 37 metabolites across species. When compared in the high nutrient treatment, species native to low nutrient soils exhibited overall higher exudation than their sister species native to high nutrient soils in all three species pairs, providing support for repeated evolutionary shifts in response to native soil fertility. Species native to low nutrient soils and those native to high nutrient soils responded similarly to low nutrient treatments with increased exudation of organic acids (fumaric, citric, malic acids) and glucose, potentially as a mechanism to enhance nutrition acquisition. However, species native to low nutrient soils also responded to low nutrient treatments with a larger decrease in exudation of amino acids than species native to high nutrient soils in all three species pairs. This indicates that species native to low nutrient soils have evolved a unique sensitivity to changes in nutrient availability for some, but not all, root exudates. Overall, these repeated evolutionary divergences between species native to low nutrient soils and those native to high nutrient soils provide evidence for the adaptive value of root exudation, and its plasticity, in contrasting soil environments.

Water plays a central role in plant biology and the efficiency of water transport throughout the plant affects both photosynthetic rate and growth, an influence that scales up deterministically to the productivity of terrestrial ecosystems. Moreover, hydraulic traits mediate the ways in which plants interact with their abiotic and biotic environment. At landscape to global scale, plant hydraulic traits are important in describing the function of ecological communities and ecosystems. Plant hydraulics is increasingly recognized as a central hub within a network by which plant biology is connected to palaeobiology, agronomy, climatology, forestry, community and ecosystem ecology and earth-system science. Such grand challenges as anticipating and mitigating the impacts of climate change, and improving the security and sustainability of our food supply rely on our fundamental knowledge of how water behaves in the cells, tissues, organs, bodies and diverse communities of plants. A workshop, 'Emerging Frontiers in Plant Hydraulics' supported by the National Science Foundation, was held in Washington DC, 2015 to promote open discussion of new ideas, controversies regarding measurements and analyses, and especially, the potential for expansion of up-scaled and down-scaled inter-disciplinary research, and the strengthening of connections between plant hydraulic research, allied fields and global modelling efforts.

Studies of local adaptation generally investigate plants growing in relatively stable habitats. We asked whether populations of the long-lived clonal grass Uniola paniculata (Sea Oats) are locally adapted to microhabitats in the southeastern US coastal dunes, a habitat characterized by dynamic environmental gradients spanning relatively small distances. Although vegetative zonation is well characterized across these gradients, little is known about intraspecific evolutionary responses of species spanning the gradients. Plants from the foredune and backdune areas of the gradient (

Hybrid species provide a unique opportunity to examine trait evolution. We previously demonstrated that the hybrid species Helianthus anomalus is more fit than its ancestral parents, Helianthus annuus and Helianthus petiolaris, in its habitat and hypothesized that nutrient limitations have been a major selective agent in that adaptive evolution. In this glasshouse study, we asked whether H. anomalus has more conservative growth and resource use traits than its ancestral parents, as expected for low-nutrient-adapted species. Helianthus anomalus had a lower maximum relative growth rate, leaf area ratio, and specific leaf area as well as less of a decrease in relative growth rate with nitrogen limitation than its ancestral parents. Helianthus anomalus also had higher nitrogen use efficiency, largely because of a longer mean residence time, as well as greater leaf toughness and longer leaf life span. However, H. anomalus did not have a lower photosynthetic rate, higher ratio of fine root length to leaf area, ...

Plant uptake of soil organic N in addition to inorganic N could play an important role in ecosystem N cycling as well as plant nutrition. We measured in situ plant uptake of organic and inorganic N by the dominant canopy species in two contrasting temperate forest ecosystems (bottomland floodplain forest, subxeric sandhills long-leaf pine forest). Seedlings of Acer rubrum L. and Pinus palustris Mill. in floodplain and sandhills forests, respectively, were treated with isotopically-enriched organic N (15N-[2]-13C-glycine) or inorganic N (15NH4+) to examine in situ uptake. We also measured soil organic and inorganic N concentrations to assess the availability of N for plant uptake. Neither species took up organic N as intact 15N-[2]-13C-glycine, but significant root 15N enrichment in both species indicated that N mineralized from labeled glycine was taken up. Free amino-N dominated the total 2 M KCl-extractable N in floodplain (57 ± 3%) and sandhills soils (75 ± 3%), followed by NH4+ then NO3− in b...

Woody plants native to mesic habitats tend to be more vulnerable to drought-induced cavitation than those in xeric habitats. Cavitation resistance in herbaceous plants, however, is rarely studied and whether or not annual plants in arid habitats conform to the trends observed in woody plants is unknown. This question is addressed by comparing the hydraulic properties of annual plants endemic to relatively mesic and seasonally xeric habitats in the Great Basin Desert, in both native and experimental settings. Vulnerability to cavitation between species differed as predicted when vulnerability curves of similar-sized native individuals were compared. Contrary to expectations, Helianthus anomalus from the relatively mesic dune sites, on average, exhibited higher native embolism, lower soil-to-leaf hydraulic conductance (k(L)) and lower transpiration rates, than its xeric analogue, H. deserticola. In transplant gardens, H. anomalus' vulnerability to cavitation was unaffected by transplant location or watering treatment. In H. deserticola, however, vulnerability to cavitation varied significantly in response to watering in transplant gardens and varied as a function of stem water potential (Psi(stem)). H. deserticola largely avoided cavitation through its higher water status and generally more resistant xylem, traits consistent with a short life cycle and typical drought-escape strategy. By contrast, H. anomalus' higher native embolism is likely to be adaptive by lowering plant conductance and transpiration rate, thus preventing the loss of root-to-soil hydraulic contact in the coarse sand dune soils. For H. anomalus this dehydration avoidance strategy is consistent with its relatively long 3-4 month life cycle and low-competition habitat. We conclude that variance of hydraulic parameters in herbaceous plants is a function of soil moisture heterogeneity and is consistent with the notion that trait plasticity to fine-grained environmental variation can be adaptive.

Nighttime transpirational water loss from C(3) trees occurs without carbon gain and is both common and substantial. However, the magnitude of this water loss varies and a better understanding of the environmental factors driving this variation is needed. We investigated the response of nighttime conductance (g(night)) and transpiration (E(night)) to soil nitrogen limitation. We used instantaneous gas exchange measurements in greenhouse studies of Populus angustifolia James (narrowleaf cottonwood) and Populus balsamifera L. spp. trichocarpa (Torr.A. Gray ex Hook.) Brayshaw (black cottonwood). g(night) for sufficiently watered plants ranged from 0.045 to 0.308 mol m(-2) s(-1) for P. balsamifera and 0.037 to 0.188 mol m(-2) s(-1) for P. angustifolia, which was much larger than minimum leaf conductance (g(min); up to 0.005 mol m(-2) s(-1) in the dark). Long-term nitrogen limitation sufficient to substantially reduce biomass did not affect g(night) or E(night) when potentially confounding water stress effects were eliminated. We conclude that nighttime water loss from two Populus species is large and although it is under stomatal control is not regulated at night in response to soil nitrogen availability.

C(3) plants dominate many landscapes and are critically important for ecosystem water cycling. At night, plant water losses can include transpiration (E(night)) from the canopy and hydraulic redistribution (HR) from roots. We tested whether E(night) limits the magnitude of HR in a greenhouse study using Artemisia tridentata, Helianthus anomalus and Quercus laevis. Plants were grown with their roots split between two compartments. HR was initiated by briefly withholding all water, followed by watering only one rooting compartment. Under study conditions, all species showed substantial E(night) and HR (highest minus lowest soil water potential [Psi(s)] during a specified diel period). Suppressing E(night) by canopy bagging increased HR during the nightly bagging period (HR(N)) for A. tridentata and H. anomalus by 73 and 33% respectively, but did not affect HR(N) by Q. laevis. Total daily HR (HR(T)) was positively correlated with the Psi(s) gradient between the rooting compartments, which was correlated with light and/or atmospheric vapour pressure deficit (VPDa) the prior day. For A. tridentata, HR(T) was negatively correlated with night-time VPDa. Ecological implications of the impact of E(night) on HR may include decreased plant productivity during dry seasons, altered ecosystem water flux patterns and reduced nutrient cycling in drying soils.

Habitats that differ in soil resource availability are expected to differ for selection on resource-related plant traits. Here, we examined spatial and temporal variation in phenotypic selection on leaf ecophysiological traits for 10 Helianthus populations, including two species of hybrid origin, Helianthus anomalus and Helianthus deserticola, and artificial hybrids of their ancestral parents. Leaf traits assessed were leaf size, succulence, nitrogen (N) concentration and water-use efficiency (WUE). Biomass and leaf traits of artificial hybrids indicate that the actively moving dune habitat of H. anomalus was more growth limiting, with lower N availability but higher relative water availability than the stabilized dune habitat of H. deserticola. Habitats differed for direct selection on leaf N and WUE, but not size or succulence, for the artificial hybrids. However, within the H. anomalus habitat, direct selection on WUE also differed among populations. Across years, direct selection on leaf traits did not differ. Leaf N was the only trait for which direct selection differed between habitats but not within the H. anomalus habitat, suggesting that nutrient limitation is an important selective force driving adaptation of H. anomalus to the active dune habitat.

Evolutionary ecologists have long been intrigued by the fact that many plant species can inhabit a broad range of environmental conditions and that plants often exhibit dramatic differences in phenotype across environmental gradients. We investigated responses to salinity treatments in the salt marsh plant Borrichia frutescens to determine if the species is responding to variation in edaphic salt content through phenotypic plasticity or specialized trait response. We grew seedlings from fruits collected in high- and low-salt microhabitats, assigned seedlings to high- and low-salt treatments in a greenhouse, and measured traits related to salt tolerance. All traits were highly plastic in response to salinity. Plants from the two microhabitats did not differ in trait means or respond differently to the treatments. These results suggest that environmental differences between the two microhabitats are not creating genotypes adapted to high and low salt levels. In addition, despite evidence for variation in allozyme markers in this population, there was no significant genotypic variation (family effect) in any of the trait means measured across microhabitats. There was variation in plasticity for only leaf Na and leaf B concentration. The high degree of plasticity for all traits and the lack of differences among microhabitats across the salinity gradient suggest plasticity in many traits may be fixed for this species.

Night-time leaf conductance (g(night)) and transpiration may have several adaptive benefits related to plant water, nutrient and carbon relations. Little is known, however, about genetic variation in g(night) and whether this variation correlates with other gas exchange traits related to water use and/or native habitat climate. We investigated g(night) in 12 natural accessions and three near isogenic lines (NILs) of Arabidopsis thaliana. Genetic variation in g(night) was found for the natural accessions, and g(night) was negatively correlated with native habitat atmospheric vapour pressure deficit (VPD(air)), suggesting lower g(night) may be favoured by natural selection in drier habitats. However, there were also significant genetic correlations of g(night) with daytime gas exchange traits expected to affect plant fitness [i.e. daytime leaf conductance, photosynthesis and intrinsic water-use efficiency (WUE(i))], indicating that selection on daytime gas exchange traits may result in indirect selection on g(night). The comparison of three NILs to their parental genotypes identified one quantitative trait locus (QTL) contributing to variation in g(night). Further characterization of genetic variation in g(night) within and among populations and species, and of associations with other traits and native habitats will be needed to understand g(night) as a putatively adaptive trait.

Water movement from roots to soil at night in the process of hydraulic lift (redistribution) rehydrates the rhizosphere and has been proposed to improve plant nutrient acquisition. Another process that has now been found in many plant species is nighttime transpiration and this could also affect nutrient relations by influencing supply of mobile nutrients to roots at night. The effects of these soil/root water relations interactions have not been adequately tested. We chose ten Sarcobatus vermiculatus (Hook.) Torrey shrubs with different magnitudes of hydraulic lift (i.e. diel range in soil water potential) to test the hypothesis that the magnitude of lift would be positively related to the amount of nitrogen (N) uptake over a period of days. A 15N tracer was injected in the 20–30 cm soil layer at locations with hydraulic lift to determine plant 15N acquisition by shallow roots conducting hydraulic lift. Half of the plants were also placed in large humidified tents (i.e. “bagged”), which suppressed nighttime transpiration, and thus were expected to have greater magnitudes of hydraulic lift, although they did not. All plants took up the 15N tracer, but contrary to our hypothesis the magnitude of hydraulic lift had no significant effect on the amount of 15N acquired over a 9-day period following labeling. However, plants that were bagged tended to have lower 15N acquisition (P = 0.07). These data indicate that decreased nighttime transpirational water loss or some other effect of bagging may decrease nutrient acquisition by these nutrient-limited phreatophytic shrubs and more generally suggests a possible nutritional benefit of nighttime transpiration by plants. This suggestion needs more thorough testing to elucidate an important potential link between plant water and nutrient relations.

Plant species with separate genders often exhibit gender differences in traits related to reproductive allocation. In gynodioecious species, females often produce more seeds than do hermaphrodites, leading to a higher reproductive cost. The mechanisms that allow females to meet the high costs of reproduction are currently under debate. In this study, we test the hypothesis that there are genetically based gender differences in physiological traits that enable females to finance these costs through higher photosynthetic carbon gain in the gynodioecious perennial Geranium maculatum. Females and hermaphrodites were compared in a greenhouse study that minimized environmental and selfing rate differences between the genders. We found that females produced smaller flowers but more of them and more fruits than did hermaphrodites. However, genders did not differ in their seed number, seed mass, fruit set, and reproductive allocation. In addition, genders did not differ in photosynthetic rate (A), leaf N, and wate...

Natural hybridization accompanied by a shift in niche preference by hybrid genotypes can lead to hybrid speciation. Natural selection may cause the fixation of advantageous alleles in the ecologically diverged hybrids, and the loci experiencing selection should exhibit a reduction in allelic diversity relative to neutral loci. Here, we analyzed patterns of genetic diversity at 59 microsatellite loci associated with expressed sequence tags (ESTs) in a homoploid hybrid sunflower species, Helianthus anomalus. We used two indices, ln RV and ln RH, to compare variation and heterozygosity (respectively) at each locus between the hybrid species and its two parental species, H. annuus and H. petiolaris. Mean values of ln RV and ln RH were significantly lower than zero, which implies that H. anomalus experienced a population bottleneck during its recent evolutionary history. After correcting for the apparent bottleneck, we found six loci with a significant reduction in variation or with heterozygosity in the hybrid species, compared to one or both of the parental species. These loci should be viewed as a ranked list of candidate loci, pending further sequencing and functional analyses. Sequence data were generated for two of the candidate loci, but population genetics tests failed to detect deviations from neutral evolution at either locus. Nonetheless, a greater than eight-fold excess of nonsynonymous substitutions was found near a putative N-myristoylation motif at the second locus (HT998), and likelihood-based models indicated that the protein has been under selection in H. anomalus in the past and, perhaps, in one or both parental species. Finally, our data suggest that selective sweeps may have united populations of H. anomalus isolated by a mountain range, indicating that even low gene-flow species may be held together by the spread of advantageous alleles.

Incomplete stomatal closure during the night is observed in a diverse range of C3 and C4 species ([Fig. 1][1] ; Supplemental Table S1) and can lead to substantial nighttime transpirational water loss. Although water loss is an inevitable consequence of stomatal opening for photosynthetic carbon gain

The leaf economics spectrum (LES) describes a major axis of plant functional trait variation worldwide, defining suites of leaf traits aligned with resource-acquisitive to resource-conservative ecological strategies. The LES has been interpreted to arise from leaf-level trade-offs among ecophysiological traits common to all plants. However, it has been suggested that the defining leaf-level trade-offs of the LES may not hold within specific functional groups (e.g., herbs) nor within many groups of closely related species, which challenges the usefulness of the LES paradigm across evolutionary scales. Here, we examine the evolution of the LES across 28 species of the diverse herbaceous genus Helianthus (the sunflowers), which occupies a wide range of habitats and climate variation across North America. Using a phylogenetic comparative approach, we find repeated evolution of more resource-acquisitive LES strategies in cooler, drier, and more fertile environments. We also find macroevolutionary correlations among LES traits that recapitulate aspects of the global LES, but with one major difference: leaf mass per area is uncorrelated with leaf lifespan. This indicates that whole-plant processes likely drive variation in leaf lifespan across Helianthus, rather than leaf-level trade-offs. These results suggest that LES patterns do not reflect universal physiological trade-offs at small evolutionary scales.

We investigated the response of Helianthus species nighttime conductance (g night) and transpiration (E night) to soil nutrient and water limitations in nine greenhouse studies. The studies primarily used wild Helianthus annuus, but also included a commercial and early domesticate of H. annuus and three additional wild species (Helianthus petiolaris Nutt., Helianthus deserticola Heiser, and Helianthus anomalus Blake). Well-watered plants of all species showed substantial g night (0.023–0.225 mol m−2 s−1) and E night (0.29–2.46 mmol m−2 s−1) measured as instantaneous gas exchange. Based on the potential for transpiration to increase mass flow of mobile nutrients to roots, we hypothesized that g night and E night would increase under limiting soil nutrients but found no evidence of responses in all six studies testing this. Based on known daytime responses to water limitation, we hypothesized that g night and E night would decrease when soil water availability was limited, and results from all four studies testing this supported our hypothesis. We also established that stomatal conductance at night was on average 5 times greater than cuticular conductance. Additionally, g night and E night varied nocturnally and across plant reproductive stages while remaining relatively constant as leaves aged. Our results further the ability to predict conditions under which nighttime water loss will be biologically significant and demonstrate that for Helianthus, g night can be regulated.

We examined how plant traits related to growth and resource use have evolved during hybrid speciation and specialization into stressful habitats. Two desert sunflower species of homoploid hybrid origin are endemic to habitats with lower soil nutrient levels than those of their ancestral parent species. We hypothesized that the hybrid species would exhibit greater tolerance to low levels of soil nutrients than their parental species. The 2 hybrid species, Helianthus anomalus and H. deserticola, and their parental species, H. annuus and H. petiolaris, were compared for plant traits and growth through reproduction under 3 nutrient levels in a greenhouse study. An additional seedling study compared species for maximum seedling relative growth rate under optimum conditions. The hybrid species did have greater tolerance of nutrient limitation than the parental species, demonstrated by a resistance to change in stem height and diameter growth across treatments. A similar trend was observed in total biom...

Can the complex phenotypes that characterize naturally occurring hybrid species be re-created in early-generation artificial hybrids? We address this question with three homoploid hybrid species (Helianthus anomalus, Helianthus deserticola, Helianthus paradoxus) and their ancestral parents (Helianthus annuus, Helianthus petiolaris) that are phenotypically distinct and ecologically differentiated. These species, and two synthetic hybrid populations of the ancestral parents, were characterized for morphological, physiological, and life-history traits in greenhouse studies. Among the synthetic hybrids, discriminant analysis identified a few individuals with the multitrait phenotype of the natural hybrid species: 0.7%–1.1% were H. anomalus-like, 0.5%–13% were H. deserticola-like, and only 0.4% were H. paradoxus-like. These relative frequencies mirror previous findings that genetic correlations are favorable for generating the hybrid species’ phenotypes, and they correspond well with phylogeographic evidence that demonstrates multiple natural origins of H. deserticola and H. anomalus but a single origin for H. paradoxus. Even though synthetic hybrids with hybrid species phenotypes are rare, their phenotypic correlation matrices share most of the same principal components (eigenvectors), setting the stage for predictable recovery of hybrid species’ phenotypes from different hybrid populations. Our results demonstrate past hybridization could have generated hybrid species-like multitrait phenotypes suitable for persistence in their respective environments in just three generations after initial hybridization.

Summary 1 Recent studies of hybridization have shown that extreme (e.g. transgressive) phenotypes can be generated during massive genetic recombination events such as interspecific hybridization. Extreme hybrid phenotypes appear to have the potential to create new lineages and lead to evolutionary divergence. 2 Hybrid genotypes of some salt marsh species have proven capable of reengineering their ecosystems, while in other cases, hybridization has transformed plant communities by facilitating species invasion. 3 Using the Louisiana irises, a naturally hybridizing group of species, we assessed the potential for hybrids of Iris brevicaulis and I. fulva to possess extreme physiological and growth traits. Our glasshouse experiment used two soil water environments (wet and flood) to measure hybrid trait expression at both the genotypic class and individual genotype level. 4 At the genotypic class level, two of the three hybrid classes (F1s, and backcrosses towards each parental species) were transgressive for at least one physiological trait (F1s for root mass proportion, specific leaf area and final leaf area; backcrosses to I. brevicaulis for leaf area ratio). Three fitness components (total biomass, ramet production and flower production) were measured on each plant in the experiment. All three hybrid classes had greater clonal fitness (total biomass) but not greater sexual fitness (flower number) than parent species. 5 At the individual genotype level, two F1 hybrids showed extreme physiological trait expression: one for specific leaf area, and the other for shallow root allocation. Flower production was the only fitness component that showed some degree of environment-dependence at the individual genotype level. 6 Although early generation hybrid classes can contain rare genotypes with extreme phenotypic trait expression, and hybrid fitness components were equal or superior to both parental species in most cases, there was not a strong association between transgressive traits and elevated hybrid fitness.

In salt marsh habitats, noted for their extreme environments, a widely held assumption is that a few large clones dominate plant populations. Using a large number of polymorphic genetic markers, we were able to test this assumption for two salt marsh plants known to span extreme salinity gradients. For both species, clonal diversity was surprisingly high across populations: Simpson's diversity indices were 0.96 and 0.99. Although clonal diversity was high, there was no pattern of association between specific clones or alleles with salt microhabitat. Our findings suggest that sexual reproduction and recruitment from seeds may have been generally underappreciated as an important ecological force in the salt marsh. Furthermore, clonal diversity has implications for conservation and restoration of these critical coastal habitats, particularly with regard to buffering environmental change or disease. Recent studies also suggest that high levels of intraspecific diversity can affect a variety of community and ecosystem processes.