The role of anthropogenic dispersal in shaping the distribution and genetic composition of a widespread North American tree species.

Dispersal and colonization are among the most important ecological processes for species persistence as they allow species to track changing environmental conditions. During the last glacial maximum (LGM), many cold-intolerant Northern Hemisphere plants retreated to southern glacial refugia. During subsequent warming periods, these species expanded their ranges northward. Interestingly, some tree species with limited seed dispersal migrated considerable distances after the LGM ~19,000 years before present (YBP). It has been hypothesized that indigenous peoples may have dispersed valued species, in some cases beyond the southern limits of the Laurentide Ice Sheet. To investigate this question, we employed a molecular genetics approach on a widespread North American understory tree species whose fruit was valued by indigenous peoples. Twenty putative anthropogenic (near pre-Columbian habitations) and 62 wild populations of Asimina triloba (pawpaw), which produces the largest edible fruit of any North American tree, were genetically assayed with nine microsatellite loci. Putative anthropogenic populations were characterized by reduced genetic diversity and greater excess heterozygosity relative to wild populations. Anthropogenic populations in regions that were glaciated during the LGM had profiles consistent with founder effects and reduced gene flow, and shared rare alleles with wild populations hundreds of kilometers away (mean = 723 km). Some of the most compelling evidence for human-mediated dispersal is that putative anthropogenic and wild populations sharing rare alleles were separated by significantly greater distances (mean = 695 km) than wild populations sharing rare alleles (mean = 607 km; p = .014). Collectively, the genetic data suggest that long-distance dispersal played an important role in the distribution of pawpaw and is consistent with the hypothesized role of indigenous peoples.

Phylogeography of the Neotropical epiphytic orchid, Brassavola nodosa: evidence for a secondary contact zone in northwestern Costa Rica.

Spatial patterns of genetic variation can reveal otherwise cryptic evolutionary and landscape processes. In northwestern Costa Rica, an approximately concordant genetic discontinuity occurs among populations of several plant species. We conducted phylogeographic analyses of an epiphytic orchid, Brassavola nodosa, to test for genetic discontinuity and to explore its underlying causes. We genotyped 18 populations with 19 nuclear loci and two non-coding chloroplast sequence regions. We estimated genetic diversity and structure, relative importance of pollen and seed dispersal, and divergence time to understand how genetic diversity was spatially partitioned. Nuclear genetic diversity was high with little differentiation among populations (GSTn = 0.065). In contrast, chloroplast haplotypes were highly structured (GSTc = 0.570) and reveal a discontinuity between northwestern and southeastern populations within Costa Rica. Haplotype differences suggest two formerly isolated lineages that diverged ~10,000-100,000 YBP. Haplotype mixing and greater genetic diversity occur in an intermediate transition zone. Patterns of nuclear and chloroplast data were consistent. Different levels of genetic differentiation for the two genomes reflect the relative effectiveness of biotic versus abiotic dispersers of pollen and seeds, respectively. Isolation of the two lineages likely resulted from the complex environmental and geophysical history of the region. Our results suggest a recent cryptic seed dispersal barrier and/or zone of secondary contact. We hypothesize that powerful northeasterly trade winds hinder movement of wind-borne seeds between the two regions, while the multi-directional dispersal of pollen by strong-flying sphinx moths resulted in lower differentiation of nuclear loci.

Heterogeneity of clonal patterns among patches of kudzu, Pueraria montana var. lobata, an invasive plant.

BACKGROUND AND AIMS: Viny species are among the most serious invasive plants, and better knowledge of how vines grow to dominate landscapes is needed. Patches may contain a single genotype (i.e. genet), a competitively dominant genet or many independent but interacting genets, yet the clonal structure of vining species is often not apparent. Molecular markers can discriminate among the genetic identities of entwined vines to reveal the number and spatial distribution of genets. This study investigated how genets are spatially distributed within and among discrete patches of the invasive vine kudzu, Pueraria montana var. lobata, in the United States. It was expected that ramets of genets would be spatially clustered within patches, and that an increase in the number of genets within a patch would be associated with a decrease in the average size of each genet. METHODS: Six discrete kudzu patches were sampled across 2 years, and 1257 samples were genotyped at 21 polymorphic allozyme loci. Variation in genotypic and genetic diversity among patches was quantified and patterns of genet interdigitation were analysed. KEY RESULTS: Substantial genotypic and genetic variation occurred within and among patches. As few as ten overlapping genets spanned up to 68 m(2) in one patch, while >90 % of samples were genetically unique in another patch. Genotypic diversity within patches increased as mean clone size decreased, although spatially widespread genets did not preclude interdigitation. Eight genets were shared across ≥2 patches, suggesting that vegetative dispersal can occur among patches. CONCLUSIONS: Genetically unique kudzu vines are highly interdigitated. Multiple vegetative propagules have become established in spatially discrete patches, probably through the movement of highway construction or maintenance machinery. The results suggest that common methods for controlling invasive vines (e.g. mowing) may inadvertently increase genotypic diversity. Thus, understanding vine architecture and growth has practical implications.

High but variable outcrossing rates in the invasive Geranium carolinianum (Geraniaceae).

• Premise of the study: Mating system plays an important role in population establishment and persistence, maintenance of genetic variation, and adaptive ability, especially for invasive species that colonize new environments to which they may be poorly adapted. In mixed-mating species, population differences in self-fertilization rates often arise due to variation in local ecological conditions (e.g., pollinator or mate availability) or genetic variation in traits promoting selfing or outcrossing. Knowledge of how and why selfing rates vary can help us understand how populations respond to different environments, how this affects patterns of genetic variation, and the role of mating systems in biological invasions.• Methods: We determined outcrossing rates in invasive (China) and native (US) populations of the weedy annual Geranium carolinianum in natural populations and an open-pollinated common garden to answer the following questions: To what extent do populations vary in mating system? Do invasive populations differ from native populations? Is interpopulation variation in mating system under genetic control?• Key results: Despite having many characteristics of selfing species, we found high variation in outcrossing rates (tm from 0.1 to 1.0) resulting from both environmental and genetic variation. Outcrossing rates were generally high and plastic in the Chinese populations.• Conclusions: A delayed selfing mechanism provides reproductive assurance while allowing facultative outcrossing when mates are not limiting. In invasive populations, high outcrossing rates were facilitated by large admixed founding populations, promoting local adaptation and the maintenance of genetic diversity.

Mating system contributes only slightly to female maintenance in gynodioecious Geranium maculatum (Geraniaceae).

Gynodioecy, the co-occurrence of female and hermaphroditic individuals within a population, is an important intermediate in the evolution of separate sexes. The first step, female maintenance, requires females to have higher seed fitness compared with hermaphrodites. A common mechanism thought to increase relative female fitness is inbreeding depression avoidance, the magnitude of which depends on hermaphroditic selfing rates and the strength of inbreeding depression. Less well studied is the effect of biparental inbreeding on female fitness. Biparental inbreeding can affect relative female fitness only if its consequence or frequency differs between sexes, which could occur if sex structure and genetic structure both occur within populations. To determine whether inbreeding avoidance and/or biparental inbreeding can account for female persistence in Geranium maculatum, we measured selfing and biparental inbreeding rates in four populations and the spatial genetic structure in six populations. Selfing rates of hermaphrodites were low and did not differ significantly from zero in any population, leading to females gaining at most a 1-14% increase in seed fitness from inbreeding avoidance. Additionally, although significant spatial genetic structure was found in all populations, biparental inbreeding rates were low and only differed between sexes in one population, thereby having little influence on female fitness. A review of the literature revealed few sexual differences in biparental inbreeding among other gynodioecious species. Our results show that mating system differences may not fully account for female maintenance in this species, suggesting other mechanisms may be involved.

Genetic and morphological contrasts between wild and anthropogenic populations of Agave parryi var. huachucensis in south-eastern Arizona.

BACKGROUND AND AIMS: At least seven species of Agave, including A. parryi, were cultivated prehistorically in Arizona, serving as important sources of food and fibre. Many relict populations from ancient cultivation remain in the modern landscape, offering a unique opportunity to study pre-Columbian plant manipulation practices. This study examined genetic and morphological variation in six A. p. var. huachucensis populations of unknown origin to compare them with previous work on A. parryi populations of known origin, to infer their cultivation history and to determine whether artificial selection is evident in populations potentially managed by early agriculturalists. METHODS: Six A. p. var. huachucensis and 17 A. parryi populations were sampled, and morphometric, allozyme and microsatellite data were used to compare morphology and genetic structure in purportedly anthropogenic and wild populations, as well as in the two taxa. Analysis of molecular variance and Bayesian clustering were performed to partition variation associated with taxonomic identity and hypothesized evolutionary history, to highlight patterns of similarity among populations and to identify potential wild sources for the planting stock. KEY RESULTS: A p. var. huachucensis and A. parryi populations differed significantly both morphologically and genetically. Like A. parryi, wild A. p. var. huachucensis populations were more genetically diverse than the inferred anthropogenic populations, with greater expected heterozygosity, percentage of polymorphic loci and number of alleles. Inferred anthropogenic populations exhibited many traits indicative of past active cultivation: greater morphological uniformity, fixed heterozygosity for several loci (non-existent in wild populations), fewer multilocus genotypes and strong differentiation among populations. CONCLUSIONS: Where archaeological information is lacking, the genetic signature of many Agave populations in Arizona can be used to infer their evolutionary history and to identify potentially fruitful sites for archaeological investigation of ancient settlements and cultivation practices. The same approach can clearly be adopted for other species in similar situations.

Patterns of genetic diversity reveal multiple introductions and recurrent founder effects during range expansion in invasive populations of Geranium carolinianum (Geraniaceae).

Genetic diversity, and thus the adaptive potential of invasive populations, is largely based on three factors: patterns of genetic diversity in the species' native range, the number and location of introductions and the number of founding individuals per introduction. Specifically, reductions in genetic diversity ('founder effects') should be stronger for species with low within-population diversity in their native range and few introductions of few individuals to the invasive range. We test these predictions with Geranium carolinianum, a winter annual herb native to North America and invasive in China. We measure the extent of founder effects using allozymes and microsatellites, and ask whether this is consistent with its colonization history and patterns of diversity in the native range. In the native range, genetic diversity is higher and structure is lower than expected based on life history traits. In China, our results provide evidence for multiple introductions near Nanjing, Jiangsu province, with subsequent range expansion to the west and south. Patterns of genetic diversity across China reveal weak founder effects that are driven largely by low-diversity populations at the expansion front, away from the introduction location. This suggests that reduced diversity in China has resulted from successive founder events during range expansion, and that the loss of genetic diversity in the Nanjing area was mitigated by multiple introductions from diverse source populations. This has implications for the future of G. carolinianum in China, as continued gene flow among populations should eventually increase genetic diversity within the more recently founded populations.

Genetic inference of epiphytic orchid colonization; it may only take one.

Colonization of vacant habitat is a fundamental ecological process that affects the ability of species to persist and undergo range modifications in continually shifting landscapes. Thus, understanding factors that affect and limit colonization has important ecological and conservation implications. Epiphytic orchids are increasingly threatened by various factors, including anthropogenic habitat disturbance. As cleared areas (e.g. pastures) are recolonized by suitable host trees, the establishment and genetic composition of epiphytic orchid populations are likely a function of their colonization patterns. We used genetic analyses to infer the prevailing colonization pattern of the epiphytic orchid, Brassavola nodosa. Samples from three populations (i.e. individuals within a tree) from each of five pastures in the dry forest of Costa Rica were genotyped with neutral nuclear and chloroplast markers. Spatial autocorrelation and hierarchical genetic structure analyses were used to assess the relatedness of individuals within populations, among populations within pastures and among populations in different pastures. The results showed significant relatedness within populations (mean r=0.166) and significant but lower relatedness among populations within a pasture (mean r=0.058). Our data suggest that colonization of available habitats is by few individuals with subsequent population expansion resulting from in situ reproduction, and that individuals within a tree are not a random sample of the regional seed pool. Furthermore, populations within a pasture were likely colonized by seeds produced by founders of a neighbouring population within that pasture. These results have important ramifications for understanding conservation measures needed for this species and other epiphytic orchids.

Development and characterization of microsatellite primers in Geranium carolinianum (Geraniaceae) with 454 sequencing.

PREMISE OF THE STUDY: Microsatellite primers were developed for Geranium carolinanum, a North American winter annual herb, for use in population genetic analyses. • METHODS AND RESULTS: Genomic DNA enriched for repeat-containing fragments was sequenced on the Roche 454 Titanium platform, resulting in 470 primer pairs developed from 1115 microsatellite-containing sequences. A subset of 37 primer pairs was screened for polymorphism across three native and three invasive populations. We identified four monomorphic and eight polymorphic loci. Polymorphic loci contained between two and seven alleles per locus, and mean within-population expected heterozygosity ranged from 0.100 to 0.290. Within populations, observed heterozygosity for individual loci ranged from zero to 0.857, and expected heterozygosity ranged from 0.046 to 0.559. • CONCLUSIONS: These microsatellite markers will be useful for future studies of genetic diversity, structure, and mating systems across the geographic range of G. carolinianum, and may be transferable to other closely related species.

Differential effects of landscape-level environmental features on genetic structure in three codistributed tree species in Central America.

Landscape genetic studies use spatially explicit population genetic information to determine the physical and environmental causes of population genetic structure on regional scales. Comparative studies that identify common barriers to gene flow across multiple species within a community are important to both understand the evolutionary trajectories of populations and prioritize habitat conservation. Here, we use a comparative landscape genetic approach to ask whether gradients in temperature or precipitation seasonality structure genetic variation across three codistributed tree species in Central America, or whether a simpler (geographic distance) or more complex, species-specific environmental niche model is necessary to individually explain population genetic structure. Using descriptive statistics and causal modelling, we find that different factors best explain genetic distance in each of the three species: environmental niche distance in Bursera simaruba, geographic distance in Ficus insipida and historical barriers to gene flow or cryptic reproductive barriers for Brosimum alicastrum. This study confirms suggestions from previous studies of Central American tree species that imply that population genetic structure of trees in this region is determined by complex interactions of both historical and current barriers to gene flow.

Evaluating the utility of microsatellites for investigations of autopolyploid taxa.

Autopolyploid taxa present numerous challenges for population genetic analyses due to difficulties determining allele dosage. Dosage ambiguity hinders accurate assessment of allele frequencies, multilocus genotypes (MLGTs), as well as levels and patterns of clonality. The pervasiveness of polyploidy in the evolutionary history of plant taxa makes this a recurring problem. Whereas diploidization of loci may occur over time, duplication of at least some loci is still frequently evident. Fortunately, with high-quality allozyme gels, it is possible to accurately infer allele dosage and, thus, determine exact MLGTs. However, accurately assessing dosage of microsatellite peaks is nearly impossible when studying wild populations with a large number of alleles per locus. Even if precise knowledge of genotypes is not required, for comparable numbers of alleles per locus and loci, the number of "phenotypes" is always lower with microsatellites than allozymes due to the inability to assess allele dosage. Microsatellite loci typically have more alleles per locus relative to allozymes although fewer loci are generally employed. Here, we present a mathematical model for comparing the relative utility of simple sequence repeat (SSR) versus allozyme markers to discriminate MLGTs. For example, the average plant allozyme study (2.6 alleles per locus, 10 polymorphic loci) has better discriminating power than SSR markers with 10 alleles at each of 3 loci, 9 alleles at 4 loci, 6 alleles at 5 loci, 5 alleles at 6 loci, and 4 alleles at 8 loci, demonstrating the value of assessing the relative discriminating power of these markers.

Genetic diversity and structure in two species of Leavenworthia with self-incompatible and self-compatible populations.

Self-fertilization is a common mating system in plants and is known to reduce genetic diversity, increase genetic structure and potentially put populations at greater risk of extinction. In this study, we measured the genetic diversity and structure of two cedar glade endemic species, Leavenworthia alabamica and L. crassa. These species have self-incompatible (SI) and self-compatible (SC) populations and are therefore ideal for understanding how the mating system affects genetic diversity and structure. We found that L. alabamica and L. crassa had high species-level genetic diversity (H(e)=0.229 and 0.183, respectively) and high genetic structure among their populations (F(ST)=0.45 and 0.36, respectively), but that mean genetic diversity was significantly lower in SC compared with SI populations (SC vs SI, H(e) for L. alabamica was 0.065 vs 0.206 and for L. crassa was 0.084 vs 0.189). We also found significant genetic structure using maximum-likelihood clustering methods. These data indicate that the loss of SI leads to the loss of genetic diversity within populations. In addition, we examined genetic distance relationships between SI and SC populations to analyze possible population history and origins of self-compatibility. We find there may have been multiple origins of self-compatibility in L. alabamica and L. crassa. However, further work is required to test this hypothesis. Finally, given their high genetic structure and that individual populations harbor unique alleles, conservation strategies seeking to maximize species-level genetic diversity for these or similar species should protect multiple populations.

Inferring ancient Agave cultivation practices from contemporary genetic patterns.

Several Agave species have played an important ethnobotanical role since prehistory in Mesoamerica and semiarid areas to the north, including central Arizona. We examined genetic variation in relict Agave parryi populations northeast of the Mogollon Rim in Arizona, remnants from anthropogenic manipulation over 600 years ago. We used both allozymes and microsatellites to compare genetic variability and structure in anthropogenically manipulated populations with putative wild populations, to assess whether they were actively cultivated or the result of inadvertent manipulation, and to determine probable source locations for anthropogenic populations. Wild populations were more genetically diverse than anthropogenic populations, with greater expected heterozygosity, polymorphic loci, effective number of alleles and allelic richness. Anthropogenic populations exhibited many traits indicative of past active cultivation: fixed heterozygosity for several loci in all populations (nonexistent in wild populations); fewer multilocus genotypes, which differed by fewer alleles; and greater differentiation among populations than was characteristic of wild populations. Furthermore, manipulated populations date from a period when changes in the cultural context may have favoured active cultivation near dwellings. Patterns of genetic similarity among populations suggest a complex anthropogenic history. Anthropogenic populations were not simply derived from the closest wild A. parryi stock; instead they evidently came from more distant, often more diverse, wild populations, perhaps obtained through trade networks in existence at the time of cultivation.

Inferring recruitment history from spatial genetic structure within populations of the colonizing tree Albizia julibrissin (Fabaceae).

Comparative analyses of spatial genetic structure (SGS) among species, populations, or cohorts give insight into the genetic consequences of seed dispersal in plants. We analysed SGS of a weedy tree in populations with known and unknown recruitment histories to first establish patterns in populations with single vs. multiple founders, and then to infer possible recruitment scenarios in populations with unknown histories. We analysed SGS in six populations of the colonizing tree Albizia julibrissin Durazz. (Fabaceae) in Athens, Georgia. Study sites included two large populations with multiple, known founders, two small populations with a single, known founder, and two large populations with unknown recruitment histories. Eleven allozyme loci were used to genotype 1385 individuals. Insights about the effects of colonization history from the SGS analyses were obtained from correlograms and Sp statistics. Distinct differences in patterns of SGS were identified between populations with multiple founders vs. a single founder. We observed significant, positive SGS, which decayed with increasing distance in the populations with multiple colonists, but little to no SGS in populations founded by one colonist. Because relatedness among individuals is estimated relative to a local reference population, which usually consists of those individuals sampled in the study population, SGS in populations with high background relatedness, such as those with a single founder, may be obscured. We performed additional analyses using a regional reference population and, in populations with a single founder, detected significant, positive SGS at all distances, indicating that these populations consist of highly related descendants and receive little seed immigration. Subsequent analyses of SGS in size cohorts in the four large study populations showed significant SGS in both juveniles and adults, probably because of a relative lack of intraspecific demographic thinning. SGS in populations of this colonizing tree is pronounced and persistent and is determined by the number and relatedness of founding individuals and adjacent seed sources. Patterns of SGS in populations with known histories may be used to indirectly infer possible colonization scenarios for populations where it is unknown.

Spatial genetic structure of the southeastern North American endemic, Ceratiola ericoides (Empetraceae).

Understanding the spatial distribution of genetic diversity (i.e., spatial genetic structure [SGS]) within plant populations can elucidate mechanisms of seed dispersal and patterns of recruitment that may play an important role in shaping the demography and spatial distribution of individuals in subsequent generations. Here we investigate the SGS of allozyme diversity in 2 populations of the southeastern North American endemic shrub, Ceratiola ericoides. The data suggest that the 2 populations have similar patterns of SGS at distances of 0-45 m that likely reflect the isolation by distance (IBD) model of seed dispersal. However, at distances >or=50 m, the pattern of SGS differs substantially between the 2 populations. Whereas one population continues to reflect the classical IBD pattern, the second population shows a marked increase in autocorrelation coefficient (r) values at 50-75 m. Furthermore, r values at these distances are as much as 33% higher than at 0-5 m where the highest r value would be predicted by IBD. A likely explanation is the differing frequencies of 2 fruit morphologies in these populations and the greater role that birds play in seed dispersal in the second population.

Isolation and characterization of microsatellite loci in the Guanacaste tree, Enterolobium cyclocarpum.

We isolated nine microsatellite loci from the Guanacaste tree (Enterolobium cyclocarpum) and optimized them for future research on breeding populations of this species. Loci were screened across 53 individuals from one population and were shown to be variable with the number of alleles per locus ranging from five to 15. Polymorphic information content ranged from 0.420 to 0.900 and observed heterozygosity ranged from 0.547 to 0.906.

Comparison of clonal diversity in mountain and Piedmont populations of Trillium cuneatum (Melanthiaceae-Trilliaceae), a forest understory species.

The balance between clonal and sexual reproduction can vary widely among plant populations, and the extent of clonality may be influenced by the combined effects of historical land use and variation in environmental conditions. We investigated patterns of clonal spread in five Trillium cuneatum populations, two in the Appalachian Mountains characterized by mesic, cooler conditions, and three at lower elevations experiencing warmer, drier conditions and greater disturbance. Using a new measure of the genet effective number and innovative orthogonal contrast methods, we quantified genet structure, contrasting clonal growth in the mountains with that in the Piedmont. Asexual propagation was more common in the Piedmont, where 25% of the sampled ramets were clonally derived, but was much less frequent in the mountains (7% clonal replicates). Hierarchical partitioning of variation in genet diversity showed that the majority (75.8%) of the variation resulted from more vegetative replication in the Piedmont. Most of the remaining variation (21.6%) was attributable to differences between urban and rural Piedmont populations, and a small, statistically nonsignificant fraction of the variation (2.6%) was due to interpopulation differences within the mountains. Higher frequency of cloning may enhance both genetic and demographic population viability in fragmented Piedmont habitats.

Genetic consequences of pre-Columbian cultivation for Agave murpheyi and A. delamateri (Agavaceae).

Pre-Columbian farmers cultivated several species of agave in central Arizona from ca. A.D. 600-1350. Because of the longevity and primarily asexual reproduction of these species, relict agave clones remain in the landscape and provide insights into pre-Columbian agricultural practices. We analyzed variation in allozyme allele frequencies to infer genetic effects of prehistoric cultivation on Agave murpheyi and A. delamateri, specifically to estimate genetic diversity and structure, to determine whether cultivated populations descended from a single clone, and to examine regional-scale genetic variation. Agave murpheyi maintained more genetic diversity at the species and population levels than A. delamateri, and A. murpheyi populations typically included more multilocus genotypes. Relict plants from prehistoric fields reflect a more complex history than descent from a single clone; A. murpheyi populations may have included more diversity initially because bulbils (produced routinely in A. murpheyi but not A. delamateri) and possibly seed would have facilitated transport of genetically diverse planting stock. Genetic variation in both cultigens was lower than in most contemporary commercial crops but similar to that observed in modern traditional agricultural systems.

Hierarchical patterns of paternity within crowns of Albizia julibrissin (Fabaceae).

The floral architecture and phenology of the tree species Albizia julibrissin (Fabaceae) offer the potential for flowers within inflorescences to share common pollen donors. Patterns of paternity within individual tree crowns may differ among isolated individuals and those in populations due to differences in pollinator foraging behavior. To determine how genetic diversity is partitioned within individual seed pools and whether these patterns differ among isolated and population trees, we obtained all fruits from three inflorescences from four clusters from three isolated trees and from three population trees in Athens, Georgia. We assayed 14 polymorphic allozymes to genotype all progeny within singly sired fruits to determine the multilocus genotype of each fruit's pollen donor. Inflorescences had multiple pollen donors, but simulation analyses revealed that redundancy of pollen donors tended to be more likely within inflorescences than randomly across the crown. Analysis of genetic and genotypic diversity indicated that individual maternal trees received pollen from many donors in uneven frequencies. Results suggest that isolated trees receive pollen from slightly fewer pollen donors and experience more within-plant pollinator movement than trees in populations. However, isolated trees receive qualitatively similar pollen from many sources, suggesting that these trees are not effectively isolated and that pollen moves long distances in this species.