European colonization of eastern North America resulted in widespread loss of old-growth eastern deciduous forest, altering native plant and animal biodiversity. Though recent studies have revealed remnant old-growth forests, it remains uncertain how widespread forests with old growth characteristics are throughout the southeastern United States. This is especially true near urban centers where historical disturbance was likely most pronounced. Yet, small patches of forest with old growth characteristics persist as urban or suburban forested natural areas where they are important reservoirs of biodiversity and provide crucial ecosystem services. Here, we investigate several upland deciduous forest communities in Shelby County, Tennessee, to assess whether urban and suburban forested natural areas exhibit species assemblage and structural features consistent with commonly-accepted criteria for old-growth forest, and how they may have been shaped by diverse land use histories across an urban to suburban gradient. We found community composition broadly similar between urban and suburban forested natural areas, but also significant differences in species assemblage based on tree densities and basal areas, likely due to unique land use and disturbance histories. Forests also meet some, but not all, commonly employed criteria for western and mixed mesophytic old-growth forest, including individual tree ages >200 years, consistent with ongoing successional trajectories toward old-growth forest. Nevertheless, the urban and suburban forested natural areas of southwest Tennessee harbor important plant biodiversity in a region that has experienced intensive post-colonial anthropogenic disturbance and likely warrant continued thoughtful management and restoration to attain "new" old-growth status.

Polyploidy – whole genome duplication – is common in plants. Studies over the last several decades have documented numerous mixed‐ploidy populations. Whether arising via recurrent whole genome duplication events within a population, or from secondary contact, the persistence of mixed populations is possible by niche differentiation. Specifically, one mechanism facilitating ploidy co‐occurrence is microbially‐mediated niche differentiation (MMND), wherein cytotypes occupy different niches via interactions with different sets of microbes. Inherently cryptic, MMND is underexplored in polyploid plant populations. Here, we search for evidence of MMND in creosotebush Larrea tridentata, a dominant desert shrub of the southwestern US and northern Mexico. We sequenced root‐associated fungal taxa in soils of diploid, autotetraploid, and autohexaploid plants growing in two naturally‐occurring mixed‐cytotype populations. Within populations, we found substantial fungal assemblage overlap across host plant cytotypes. However, using indicator species analysis, we identified some fungi that are differentiated by host plant cytotype, satisfying a necessarycondition for MMND. Future study is needed to determine the degree of niche differentiation conferred, if any, and whether the identified fungi play a role in the long‐term persistence of multiple cytotypes within populations.

Premise: Theory predicts that mixed ploidy populations should be short-lived due to strong fitness disadvantages for the rare ploidy. However, mixed ploidy populations are common, suggesting that the fitness costs for rare ploidies are counterbalanced by ecological benefits that emerge when rare. We investigated whether differences in ecological interactions with soil microbes help to maintain a tetraploid-hexaploid population of Larrea tridentata (creosote bush) in the Sonoran Desert, California, United States, where prior work documented ploidy-specific root-associated microbes. Methods: We used a plant-soil feedback (PSF) experiment to test whether host-specific soil microbes can alter the outcomes of intraploidy vs. interploidy competition. Host-specific soil microbes can build up over time; thus, distance from a host plant can affect the fitness of nearby plants. Results: Seedlings grown in soils from near plants of a different ploidy produced greater biomass relative to seedlings grown in soils from near plants of the same ploidy. Moreover, seedlings grown in soils from near plants of a different ploidy produced more biomass than those grown in soils that were farther from plants of a different ploidy. These results suggest that the ecological consequences of PSF may facilitate the persistence of mixed ploidy populations. Conclusions: This is the first evidence, to our knowledge, that is consistent with plant-soil microbe feedback as a viable mechanism to maintain the coexistence of multiple ploidy levels in a single population.

PremiseTraditional methods of ploidal-level estimation are tedious; using DNA sequence data for cytotype estimation is an ideal alternative. Multiple statistical approaches to leverage sequence data for ploidy inference based on site-based heterozygosity have been developed. However, these approaches may require high-coverage sequence data, use inappropriate probability distributions, or have additional statistical shortcomings that limit inference abilities. We introduce nQuack, an open-source R package that addresses the main shortcomings of current methods.Methods and ResultsnQuack performs model selection for improved ploidy predictions. Here, we implement expectation maximization algorithms with normal, beta, and beta-binomial distributions. Using extensive computer simulations that account for variability in sequencing depth, as well as real data sets, we demonstrate the utility and limitations of nQuack.ConclusionsInferring ploidy based on site-based heterozygosity alone is difficult. Even though nQuack is more accurate than similar methods, we suggest caution when relying on any site-based heterozygosity method to infer ploidy.

The unintentional introduction and rapid spread of chestnut blight (caused by Cryphonectria parasitica (Murr.) Barr) in the early 20th century resulted in the demise of American chestnut (Castanea dentata (Marsh.) Borkh.; Fagaceae) as a major component of forest canopies resulting in negative impacts on eastern forest communities. Research efforts over the last century have documented the persistence of occasional trees and root crown/stump sprouts throughout much of the species' historic range, providing the basis for ongoing breeding of blight-resistant trees and restoration efforts. Here we use environmental niche modeling to investigate whether environmentally suitable habitat remains for remnant trees throughout the southwestern historical range, and to evaluate the reintroduction potential of this relatively understudied part of the historical distribution. We also use stage-structured matrix projection models to investigate the potential demographic future of C. dentata near the historical southwestern range limit based on observations of American chestnut in these areas over the last several decades. We found suitable upland habitat with areas of high forest canopy cover occurs throughout much of the southwestern portion of the historical range and that populations of American chestnut in these areas are predicted to drastically decline over the coming decades. These results highlight the continued presence of suitable C. dentata habitat throughout the southwestern extent of its historical distribution, which should be incorporated into evaluations for future reintroduction, and emphasize the need for efforts to locate, conserve, and introduce genetic material from individuals with locally adapted genotypes into active restoration programs.

PremiseWhole‐genome duplication (polyploidy) is an important force shaping flowering‐plant evolution. Ploidy‐specific plant–pollinator interactions represent important community‐level biotic interactions that can lead to nonrandom mating and the persistence of mixed‐ploidy populations.MethodsAt a naturally occurring diploid–tetraploid contact zone of the autopolyploid desert shrubLarrea tridentata, we combined flower phenology analyses, collections of bees on plants of known cytotype, and flow cytometry analyses of bee‐collected pollen loads to investigate whether (1) diploid and tetraploid plants have unique bee pollinator assemblages, (2) bee taxa exhibit ploidy‐specific visitation and pollen collection biases, and (3) specialist and generalist bee taxa have ploidy‐specific visitation and pollen collection biases.ResultsAlthough bee assemblages overlapped, we found significant differences in bee visitation to co‐occurring diploids and tetraploids, with the introduced honeybee (Apis mellifera) and one native species (Andrenaspecies 12) more frequently visiting tetraploids. Consistent with bee assemblage differences, we found that diploid pollen was overrepresented among pollen loads on native bees, while pollen loads onA. melliferadid not deviate from the random expectation. However, mismatches between the ploidy of pollen loads and plants were common, consistent with ongoing intercytotype gene flow.ConclusionsOur data are consistent with cytotype‐specific bee visitation and suggest that pollinator behavior contributes to reduced diploid–tetraploid mating. Differences in bee visitation and pollen movement potentially contribute to an easing of minority cytotype exclusion and the facilitation of cytotype co‐occurrence.

ABSTRACT The American chestnut (Castanea dentata) was once widespread in eastern North America and an ecologically important hardwood tree of deciduous forest communities prior to its near-eradication by chestnut blight (Cryphonectria parasitica). Remnant populations occur across much of its historical range, especially in older forests of the Appalachians and northeastern U.S. However, broad swaths of the southwestern portion of the species' historical range remain poorly documented, potentially limiting the representation of genetic variation important for local adaptation in restoration efforts. Ongoing discovery and life-history characterization efforts for remnant C. dentata remains a priority to better understand the distribution and ecological status of this once important species, while identifying potential genetic sources of locally adapted or blight resistant trees. Here, we report the discovery of 22 C. dentata at four sites in southwestern Tennessee, adding novel observations that extend the range of known extant occurrences to the extreme western edge of the historical distribution in Fayette County. These observations include potentially reproductive individuals that should be revisited to assess reproductive and blight status, and that should be evaluated for current germplasm collection and restoration efforts.

Premise Obtaining phenotypic data from herbarium specimens can provide important insights into plant evolution and ecology but requires significant manual effort and time. Here, we present LeafMachine, an application designed to autonomously measure leaves from digitized herbarium specimens or leaf images using an ensemble of machine learning algorithms. Methods and Results We trained LeafMachine on 2685 randomly sampled specimens from 138 herbaria and evaluated its performance on specimens spanning 20 diverse families and varying widely in resolution, quality, and layout. LeafMachine successfully extracted at least one leaf measurement from 82.0% and 60.8% of high‐ and low‐resolution images, respectively. Of the unmeasured specimens, only 0.9% and 2.1% of high‐ and low‐resolution images, respectively, were visually judged to have measurable leaves. Conclusions This flexible autonomous tool has the potential to vastly increase available trait information from herbarium specimens, and inform a multitude of evolutionary and ecological studies.

Aim Whole-genome duplication (polyploidy) can influence the biogeography and ecology of plants that differ in ploidy level (cytotype). Here, we address how two consequences of plant polyploidy (parapatry of cytotypes and altered species interactions) shape the biogeography of herbivorous insects. Location Warm deserts of North America. Taxa Gall midges (Asphondylia auripila group, Diptera: Cecidomyiidae) that attack three parapatric cytotypes of creosote bush (Larrea tridentata, Zygophyllaceae). Methods We surveyed Asphondylia species diversity at 177 sites across a 2300-km extent. After noting a correspondence between the distributions of eight Asphondylia species and L. tridentata cytotypes, we fine-mapped Asphondylia species range limits with transects spanning cytotype contact zones. We then tested whether plant-insect interactions and/or abiotic factors explain this coincidence by (a) comparing attack rates and gall midge communities on alternative cytotypes in a narrow zone of sympatry and (b) using species distribution models (SDMs) to determine if climatically suitable habitat for each midge species extended beyond cytotype contact zones. Results The range limits of 6/17 Asphondylia species (including two novel putative species confirmed with COI sequencing) perfectly coincided with the contact zone of diploid and tetraploid L. tridentata. One midge species was restricted to diploid host plants while five were restricted to tetraploid and hexaploid host plants. Where diploid and tetraploid L. tridentata are sympatric, cytotype-restricted midge species more frequently attacked their typical host and Asphondylia community structure differed markedly between cytotypes. SDMs predicted that distributions of cytotype-restricted midge species were not constrained by climatic conditions near cytotype contact zones. Main conclusions Contact zones between plant cytotypes are dispersal barriers for many Asphondylia species due to plant-insect interactions. The distribution of L. tridentata cytotypes therefore shapes herbivore species ranges and herbivore community structure across North American deserts. Our results demonstrate that polyploidy in plants can affect the biogeography of ecological communities.

Aim Alternative hypotheses of Darwin's Naturalization Conundrum (DNC) predict that the non-native species that successfully establish within a community are those either more closely or more distantly related to the resident native species. Despite the increasing number of studies using phylogenetic data to test DNC and evaluate community assembly, it remains unknown whether phylogenetic relationships alone can be used to predict invasion susceptibility across communities differing environmentally and in disturbance history. In this study, we evaluate whether phylogenetic structure of diverse native communities predicts the occurrence of non-native species and offers insight into community assembly. Location Eastern United States of America. Methods We examine multiple communities across a north-south transect of the eastern United States to test whether non-native species richness and abundance are associated with phylogenetic diversity measures of the native community. We also test whether non-native species are consistently closely or distantly related to native species using two approaches differing in phylogenetic scale and whether this differs with ecologically successful species. Results Our analyses did not unambiguously resolve DNC. Non-native species richness and abundance decreased with increasing native species phylogenetic diversity. Within some communities, non-native species were significantly more closely related to native species than expected by chance, and tended to be more often closely related to a native species than that native species was to other native relatives. When considering species abundance, only one community showed that ecologically successful non-native species were closely related to resident species. Main conclusions Phylogenetic relationships can reveal important details about community assembly in diverse ecological settings. However, given the multifaceted nature of community assembly, phylogenetic metrics alone have limited utility as a general predictive tool for community invasion. Our study highlights a need to incorporate additional types of data to better understand why some communities are more susceptible to non-native species establishment.

Polyploidy is widely acknowledged to have played an important role in the evolution and diversification of vascular plants. However, the influence of genome duplication on population-level dynamics and its cascading effects at the community level remain unclear. In part, this is due to persistent uncertainties over the extent of polyploid phenotypic variation, and the interactions between polyploids and co-occurring species, and highlights the need to integrate polyploid research at the population and community level. Here, we investigate how community-level patterns of phylogenetic relatedness might influence escape from minority cytotype exclusion, a classic population genetics hypothesis about polyploid establishment, and population-level species interactions. Focusing on two plant families in which polyploidy has evolved multiple times, Brassicaceae and Rosaceae, we build upon the hypothesis that the greater allelic and phenotypic diversity of polyploids allow them to successfully inhabit a different geographic range compared to their diploid progenitor and close relatives. Using a phylogenetic framework, we specifically test (1) whether polyploid species are more distantly related to diploids within the same community than co-occurring diploids are to one another, and (2) if polyploid species tend to exhibit greater ecological success than diploids, using species abundance in communities as an indicator of successful establishment. Overall, our results suggest that the effects of genome duplication on community structure are not clear-cut. We find that polyploid species tend to be more distantly related to co-occurring diploids than diploids are to each other. However, we do not find a consistent pattern of polyploid species being more abundant than diploid species, suggesting polyploids are not uniformly more ecologically successful than diploids. While polyploidy appears to have some important influences on species co-occurrence in Brassicaceae and Rosaceae communities, our study highlights the paucity of available geographically explicit data on intraspecific ploidal variation. The increased use of high-throughput methods to identify ploidal variation, such as flow cytometry and whole genome sequencing, will greatly aid our understanding of how such a widespread, radical genomic mutation influences the evolution of species and those around them.

The history of species concepts is fraught with confusion and heated exchanges. This is no less true for how best to treat polyploid plant taxa. Polyploidy—whole-genome duplication—is an important source of vascular plant diversification (Wood et al., 2009). Following the discovery of polyploidy just over a century ago, interest in genome duplication and the revelation that ploidy variation is more than an infrequent and curious phenomenon generated support for recognizing ploidy races as units of biological diversity (Ramsey and Ramsey, 2014). Nevertheless, chromosome number variation continues to challenge studies of plant ecology and evolution because of persistent uncertainties over polyploid phenotypic diversity and reproductive interactions with their diploid progenitors. Plant systematists have struggled with the classification of polyploid complexes because, in contrast to diploid populations where phenotypic differences are often evident and concomitant, polyploid complexes can exhibit phenotypic differences ranging from subtle to strikingly distinct, despite usually strong reproductive incompatibilities (Fig. 1). This disagreement over how genetic differentiation without corresponding morphological differences should be interpreted in the context of speciation has resulted in relatively few polyploids being widely recognized as taxonomic entities (Soltis et al., 2007). As a result, polyploids have been left stranded in a taxonomic no-man's land where polyploidization is simultaneously considered a major mechanism of plant speciation and biodiversity, yet insufficient for species recognition (Mayr, 1992). Whole-genome duplication can produce an array of ecologically relevant phenotypic differences. (A) Natural interspecific crosses between diploid Nicotiana sylvestris and N. tomentosiformis produce allotetraploid N. tabacum, which is distinct in floral morphology and transgressive in floral pigmentation and coloration from the perspective of their pollinators (McCarthy et al., 2017). (B) Closely related European and Asian Hedera species (Green et al., 2011) exhibit staggering leaf variation at multiple ploidal levels, including diploid H. helix and its autotetraploid H. hibernica, which are invasive in North America. (C) The geographic distributions of diploid, autotetraploid, and autohexaploid Larrea tridentata in the southwestern United States and northern Mexico are mostly non-overlapping, but in areas where they do co-occur, environmental niche predictions (warmer colors indicate higher suitability), here shown for tetraploids (gray circles) and hexaploids (black triangles), suggest the cytotypes exhibit some degree of ecological niche differentiation (Laport et al., 2013). Scale bars: (A, B) = 1 cm, (C) = 200 km. Images provided by Elizabeth McCarthy (A) and by Adam Green, Justin Ramsey, and Tara Ramsey (B). Although arguments over the tacit or formal recognition of polyploids as taxonomic species are likely to continue, more widely recognizing polyploids as functional units of biological diversity in ecological studies would acknowledge the contemporary importance of genome duplication to ecological and population dynamics, and align with the recognition of its importance to plant evolution. For example, the inclusion of ploidy variation in ecological analyses will likely challenge our understanding of interspecific interactions and biodiversity across spatial and phylogenetic scales, and have significant consequences for ecological applications such as conservation efforts (Severns and Liston, 2008) and the management of invasive species (te Beest et al., 2012). Moving forward, polyploids meeting minimum criteria for species delimitation (de Queiroz, 2007) should be explicitly treated as units of biodiversity in ecological studies similar to diploid groups, perhaps relying on functional designations such as varieties or cryptic species. To do so, integrative approaches combining evolutionary genetics and ecological analyses should be extended to identify and evaluate polyploid populations for ecological novelty. Otherwise, the continued reluctance to recognize some polyploids in ecological studies will ignore an important source of phenotypic novelty and mask the influence of genome-scale mutations on the origins of new biodiversity. The confusing relationships between polyploids and their diploid progenitors (e.g., hybridization, recurrent formation) have justifiably fostered a conservative stance by evolutionary biologists and systematists when dealing with polyploid complexes. Polyploids often exhibit low rates of intercytotype gene flow via semifertile F1 hybrids (i.e., triploids, pentaploids, etc.) or unilateral sexual polyploidization (i.e., unions of reduced and unreduced gametes), especially among higher ploidies (e.g., tetraploids, hexaploids, etc.). Yet, intercytotype reproductive isolation is typically as strong as that between diverging diploid species (Husband et al., 2016), such that polyploids meet species criteria under the Biological Species Concept. However, ploidy remains a difficult phenotypic/genetic trait to identify in the field or herbarium, and despite the evolutionary implications of polyploidy, taxonomists have largely argued that it is impractical to broadly apply Linnean classification to populations differing in ploidy without corresponding morphological differences. Taxonomic decisions can therefore be biased by a polyploid lineage's mode of origin: allopolyploids (those formed via hybridization between closely related populations) often exhibit phenotypic and molecular intermediacy, or even transgressive phenotypes (differentiated from both parents), making them easier to identify in the field or with molecular markers than autopolyploids (those formed from genome duplication within a lineage), where morphological differences are often ambiguous or absent (Soltis et al., 2007). The current genomics revolution, however, is drastically altering the way we understand species and necessitates a re-evaluation of polyploid biodiversity. For example, molecular tools are being leveraged to identify cryptic species, and whole-genome sequencing has revealed that both plant and animal species hybridize much more commonly than previously appreciated. Therefore, while a conservative approach to polyploid taxonomy may remain prudent in light of disagreements over species concepts, simply decrying the complexity of diploid–polyploid relationships as a reason for not recognizing populations differing in ploidy as units of biodiversity is not. Failing to account for the phenotypic novelty that can accompany genome duplication in ecological studies obscures the fact that an increasing number of studies show that allo- and autopolyploid cytotypes often have unique responses to the abiotic environment and novel interspecific interactions (Segraves and Anneberg, 2016). Thus, it seems appropriate that there be judicious functional recognition of currently intraspecific polyploids (e.g., as varieties or cryptic species), just as has become common practice for diploid species where significant geographical or ecological structuring, genetic or phenotypic differentiation, and/or reproductive isolation is evident. Many studies clearly show that ecological and phenotypic differences associated with ploidy shifts are common and sometimes profound. These ploidy-specific changes have the potential to influence patterns of biodiversity across varying scales. For example, recent work has demonstrated that populations differing in ploidy can exhibit climatic niche differences (Laport et al., 2013), distinct physiological strategies for water use (Maherali et al., 2009), unique mycorrhizal associations (Těšitelová et al., 2013), shifts in pollinator visitation (Thompson and Merg, 2008), increased competitive or colonization potential (te Beest et al., 2012), and differences in secondary chemistry resulting in altered flower coloration (McCarthy et al., 2017). These ploidy-specific changes have the potential to influence patterns of biodiversity across varying scales, and including ploidal diversity in ecological studies will ensure that we are not misunderstanding the fundamental ecological processes influencing patterns of biodiversity. For example, given the frequent occurrence of polyploidy in herbaceous taxa of temperate and arctic regions, measures of diversity in these areas (e.g., phenotypic, phylogenetic, or species diversity) that do not account for intraspecific ploidal variation risk misrepresenting the diversity of regional floras. Furthermore, predicting the impact of climate change on species distributions may hinge upon whether a species' range comprises ecologically and geographically differentiated cytotypes (e.g., Hersch-Green, 2012; Laport et al., 2016). The ecological consequences of genome duplication on biotic interactions is perhaps most consequential at the community level. Given the potential for ploidy-specific phenotypes and ecological differences, ploidy information could be crucial in studies of community ecology when identifying the processes underlying why particular populations and species co-occur (e.g., niche partitioning, invasiveness), and for informing restoration or management strategies (e.g., community responses to human-caused disturbance or changing environments). Interpretations of species co-occurrence or invasion naïve to ploidal variation run the risk of misrepresenting biotic interactions within a community if, for example, they fail to recognize cytotype-specific pollinator visitation (e.g., Thompson and Merg, 2008). Shifts in flowering phenology associated with genome duplication could result in an extended flowering period where two cytotypes co-occur, relative to communities comprising a single ploidy. Such differences could have cascading ecological and evolutionary effects on the pollinators and co-occurring plant species, such as providing additional opportunities for pollinator resource collection or promoting cytotype-specific specialization. Thus, whether recognition of polyploids is formalized or not, recognizing polyploids as functional units of biodiversity has the potential to offer novel insights into ecological processes and patterns at both community-level and broader spatial scales. Polyploidy continues to challenge our understanding of speciation and patterns of biodiversity. However, significant headway in understanding the evolutionary and ecological aspects of genome duplication has been made, especially over the last ∼20 years. Technological advancements are facilitating the detection and characterization of previously unrecognized ploidal variation. For example, software-aided morphometric or phenotypic analyses, ecological niche models, and phylogenetic models incorporating chromosome numbers have streamlined studies of polyploids in the wild (e.g., Mandáková and Münzbergová, 2008; Glick and Mayrose, 2014). High-throughput flow cytometry screens for DNA content have especially aided in identifying the frequency of polyploids, as well as their phenotypic and geographical distributions, and should be leveraged more widely to better document polyploid complexes (Kron et al., 2007). Moreover, online repositories of chromosome information (e.g., Chromosome Counts Database, Index to Plant Chromosome Numbers) are providing a handy way to document cytogeographic information. Our clearer understanding of the importance of polyploidy in plant evolution, combined with the advent of such tools, suggests the time is right for ecological studies to treat (at least some) intraspecific ploidy variation as independent units of biodiversity to help address major ecological and evolutionary questions. For example, it would be illuminating to identify the frequency and ways genome duplication affects community structure and biotic interactions. Do ploidy changes typically result in novel species interactions? Are polyploids more likely to invade a community? How do polyploid-associated phenotypes influence range shifts and responses to climate change? By explicitly considering ploidy variation, we can only expand our understanding of how such a rampant, radical genomic mutation, which has played such an important role in plant evolution, also affects biodiversity over ecological timescales. The authors thank Justin Ramsey, Tara Ramsey, Brian Husband, Pamela Diggle, and three anonymous reviewers for comments that improved this manuscript. Elizabeth McCarthy, Adam Green, Justin Ramsey, and Tara Ramsey graciously provided images. The ideas presented in this paper were inspired by research funded by National Science Foundation grants NSF-EF 1550813 and NSF-DEB 1556371.

Genome duplication, or polyploidy, has played an important role in the diversification of flowering plants, but the ecological and evolutionary consequences of polyploidy still remain unclear. Polyploidy is known to either cause or facilitate phenotypic changes, and ploidy-specific phenotypic differences may lead to the exploitation of novel niche space. Many studies reporting phenotypic variation between cytotypes have been observational, and may not reflect ecological adaptation by diploids and polyploids to different habitat. One such trait, water use, may play an outsized role in survival and population expansion into novel habitat. To test whether water use differs among ploidy levels, we grew field-collected diploid, autotetraploid, and autohexaploid cytotypes of the characteristic North American desert plant, Larrea tridentata (DC.) Coville, under greenhouse conditions. We measured whole plant water use gravimetrically over six wk, standardizing water use with measures of total stomatal area. We found that water use was positively correlated with stomatal area, but the cytotypes had similar total stomatal areas and did not differ significantly in mean water use/total stomatal area. Cytotype-specific water use responses through time were also not significantly different. Taken together, these results suggest that the cytotypes have similar water relationships, and possibly fitness outcomes, with respect to water use in common environments.

PREMISE OF THE STUDY: Polyploidy is widely recognized as a mechanism of diversification. Contributions of polyploidy to specific pre‐ and postzygotic barriers—and classifications of polyploid speciation as “ecological” vs. “non‐ecological”—are more contentious. Evaluation of these issues requires comprehensive studies that test ecological characteristics of cytotypes as well as the coincidence of genetic structure with cytotype distributions. METHODS: We investigated a classical example of autopolyploid speciation, Larrea tridentata , at multiple areas of cytotype co‐occurrence. Habitat and phenological differences were compared between diploid, tetraploid, and hexaploid populations on the basis of edaphic, community composition, and flowering time surveys. Frequency of hybridization between diploids and tetraploids was investigated using a diploid‐specific chloroplast DNA (cpDNA) marker; genetic structure for all cytotypes was assessed using amplified fragment length polymorphisms (AFLPs). KEY RESULTS: Across contact zones, we found cytotypes in habitats distinguished by soil and vegetation. We observed modest differences in timing and production of flowers, indicating a degree of assortative mating that was asymmetric between cytotypes. Nonetheless, cpDNA analyses in diploid–tetraploid contact zones suggested that ∼5% of tetraploid plants had hybrid origins involving unilateral sexual polyploidization. Genetic structure of AFLPs largely coincided with cytotype distributions in diploid–tetraploid contact zones. In contrast, there was little structure in areas of contact between tetraploids and hexaploids, suggesting intercytotype gene flow or recurrent hexaploid formation. CONCLUSIONS: Diploid, tetraploid, and hexaploid cytotypes of L. tridentata are segregated by environmental distributions and flowering phenology in contact zones, with diploid and tetraploid populations having corresponding differences in genetic structure.

Abstract It is increasingly recognized that evolution may occur in ecological time. It is not clear, however, how fast evolution – or phenotypic change more generally – may be in comparison with the associated ecology, or whether systems with fast ecological dynamics generally have relatively fast rates of phenotypic change. We developed a new dataset on standardized rates of change in population size and phenotypic traits for a wide range of species and taxonomic groups. We show that rates of change in phenotypes are generally no more than 2/3, and on average about 1/4, the concurrent rates of change in population size. There was no relationship between rates of population change and rates of phenotypic change across systems. We also found that the variance of both phenotypic and ecological rates increased with the mean across studies following a power law with an exponent of two, while temporal variation in phenotypic rates was lower than in ecological rates. Our results are consistent with the view that ecology and evolution may occur at similar time scales, but clarify that only rarely do populations change as fast in traits as they do in abundance.

The American chestnut ( Castanea dentata [Marsh.] Borkh.; Fagaceae) was an historically important hardwood species in eastern deciduous forests of the United States and Canada prior to being nearly eradicated by chestnut blight ( Cryphonectria parasitica (Murr.) Barr). Several remnant populations have been identified persisting across fragmented parts of the historical range. The identification and characterization of remnant C. dentata populations is important for breeding and conservation efforts, as they may represent potential genetic sources of local adaptation or blight resistance, but much of the historical range remains unsurveyed. Here, I report the locations, blight infection status, and reproductive status of remnant American chestnut in upland forested areas of western New York, finding several reproductive/potentially reproductive trees.

The methods described in this chapter were developed to avoid toxic organic phase separation utilized in many low-cost DNA extraction protocols such as the CTAB method. The steps involve: (1) lysis of the plant material, (2) binding of DNA to silica powder under chaotropic conditions, (3) washing the bound DNA, and (4) elution of DNA from the silica powder. This method has been tested in several plant species and the applicability of such DNA preparations for molecular marker studies in barley is shown in Chap. 8 .

Standard agarose gel electrophoresis is a quick method for the evaluation of the quality and quantity of DNA. This chapter provides examples of genomic DNA produced using the low-cost extraction protocol, PCR amplification using the extracted genomic DNA, and enzymatic mismatch cleavage of PCR products with crude celery juice extract and weed juice extract to detect mutations.

Denaturation and annealing of PCR products allows DNA strands with small sequence differences to hybridize together. The result is heteroduplexed molecules that are single stranded in polymorphic sequence locations, but double stranded elsewhere. These molecules are the substrates for cleavage by single-strand-specific nucleases such as CEL I, crude Celery Juice Extract (CJE) containing CEL I, and other plant extracts containing single-strand-specific nucleases [Till et al. (Nucleic Acids Res, 32:2632–2641, 2004)]. Enzymatic cleavage initiates on a single strand and can result in double strand breaks. The products of cleavage can therefore be observed using native gel electrophoresis.

A crude celery extract containing the single-strand-specific nuclease CEL I, has been widely used in TILLING and Ecotilling projects around the world. Yet, celery is hard to come by in some countries. Sequences homologous to CEL I can be found in different plant species. Previous work showed that similar mismatch cleavage activities could be found in crude extracts of mung bean (Till BJ, Burtner C, Comai L, Henikoff S. Nucleic Acids Res 32:2632–2641, 2004). It is likely that the same activity can be recovered in many different plant species. Therefore, a protocol for the extraction of active enzyme was developed that uses plants common across the world, namely weeds. Monocotyledenous and dicotyledenous weedy plants from the grassland, field and waste grounds around crop fields are suitable for this protocol. Due to lower recovery of enzymatic activity compared to celery-based extractions, a centrifuge-based filter method is applied to concentrate the enzyme extract.