Genetic Diversity and Genetic Relationships of Purple Willow (Salix purpurea L.) from Natural Locations.

In this study, the genetic diversity and structure of 13 natural locations of Salix purpurea were determined with the use of AFLP (amplified length polymorphism), RAPD (randomly amplified polymorphic DNA) and ISSR (inter-simple sequence repeats). The genetic relationships between 91 examined S. purpurea genotypes were evaluated by analyses of molecular variance (AMOVA), principal coordinates analyses (PCoA) and UPGMA (unweighted pair group method with arithmetic mean) dendrograms for both single marker types and a combination of all marker systems. The locations were assigned to distinct regions and the analysis of AMOVA (analysis of molecular variance) revealed a high genetic diversity within locations. The genetic diversity between both regions and locations was relatively low, but typical for many woody plant species. The results noted for the analyzed marker types were generally comparable with few differences in the genetic relationships among S. purpurea locations. A combination of several marker systems could thus be ideally suited to understand genetic diversity patterns of the species. This study makes the first attempt to broaden our knowledge of the genetic parameters of the purple willow (S. purpurea) from natural location for research and several applications, inter alia breeding purposes.

Contrasting biodiversity-ecosystem functioning relationships in phylogenetic and functional diversity.

It is well known that ecosystem functioning is positively influenced by biodiversity. Most biodiversity-ecosystem functioning experiments have measured biodiversity based on species richness or phylogenetic relationships. However, theoretical and empirical evidence suggests that ecosystem functioning should be more closely related to functional diversity than to species richness. We applied different metrics of biodiversity in an artificial biodiversity-ecosystem functioning experiment using 64 species of green microalgae in combinations of two to 16 species. We found that phylogenetic and functional diversity were positively correlated with biomass overyield, driven by their strong correlation with species richness. At low species richness, no significant correlation between overyield and functional and phylogenetic diversity was found. However, at high species richness (16 species), we found a positive relationship of overyield with functional diversity and a negative relationship with phylogenetic diversity. We show that negative phylogenetic diversity-ecosystem functioning relationships can result from interspecific growth inhibition. The opposing performances of facilitation (functional diversity) and inhibition (phylogenetic diversity) we observed at the 16 species level suggest that phylogenetic diversity is not always a good proxy for functional diversity and that results from experiments with low species numbers may underestimate negative species interactions.

The African baobab (Adansonia digitata, Malvaceae): genetic resources in neglected populations of the Nuba Mountains, Sudan.

UNLABELLED: • PREMISE OF THE STUDY: Adansonia digitata L. is one of the most important indigenous fruit trees of mainland Africa. Despite its significance for subsistence and income generation of local communities, little is known about the genetic and morphological variability of East African populations of A. digitata, including those of Sudan. The aim of the current study, therefore, was to analyze genetic and morphological variability of different baobab populations in Kordofan, Sudan and to estimate the effect of human intervention on genetic differentiation and diversity.• METHODS: A total of 306 trees were randomly sampled from seven spatially separated locations in the Nuba Mountains, Sudan, to cover a wide range of differing environmental gradients and management regimes ('homesteads' and 'wild'). Genetic analyses were conducted using nine microsatellite markers. Because of the tetraploid nature of A. digitata, different approaches were applied to estimate patterns of genetic diversity. Investigations were completed by measurements of dendrometric and fruit morphological characters.• KEY RESULTS: Genetic diversity was balanced and did not differ between locations or management regimes, although tendencies of higher diversity in 'homesteads' were observed. A Bayesian cluster approach detected two distinct gene pools in the sample set, mainly caused by one highly diverse population close to a main road. The variability of tree characters and fruit morphometries was high, and significantly different between locations.• CONCLUSIONS: Results indicated a rather positive effect with human intervention. The observed populations provide a promising gene pool and likely comprise ecotypes well-adapted to environmental conditions at the northern distribution range of the species, which should be considered in conservation and management programs.

Habitat fragmentation and recent bottlenecks influence genetic diversity and differentiation of the Central European halophyte Suaeda maritima (Chenopodiaceae).

PREMISE OF THE STUDY: Central European salt habitats are mainly restricted to the maritime coast but scattered occurrences can also be found inland. In inland habitats, human activities have caused losses and reductions in the size of natural salt sites but have also created new anthropogenic habitats around potash mining dumps colonized by halophytic species in the last 30 yr. We aimed to investigate the effects of bottlenecks, isolation, and ongoing habitat fragmentation on the genetic variation of a species commonly growing in these special habitats. METHODS: We used 10 microsatellite markers to compare genetic diversity and differentiation of 31 populations of Suaeda maritima (Chenopodiaceae) from Central European coasts and inland habitats. Two approaches were applied to analyze the tetraploid data based on allele frequencies directly derived from microsatellite data and from transformed binary data. KEY RESULTS: In comparison to the coastal populations from the North Sea and the English Channel, significantly reduced genetic variation and increased between-population differentiation was revealed for populations from the German inland and the Baltic Sea coast. Genetic structure analyses clearly separated coastal and inland populations. CONCLUSIONS: Our results indicate that gene flow is restricted among populations from inland salt sites and the Baltic Sea coast, presumably due to their isolation, small sizes, genetic bottlenecks and/or founder events. Patterns of allele distribution indicate some occasional genetic exchange among habitat types in the past. Anthropogenic salt sites may facilitate gene flow among inland salt habitats preventing endangered inland halophyte populations from genetic erosion.

Population size affected by environmental variability impacts genetics, traits, and plant performance in Trifolium montanum L.

Population size, genetic diversity, and performance have fundamental importance for ecology, evolution, and nature conservation of plant species. Despite well-studied relationships among environmental, genetic, and intraspecific trait variation (ITV), the influence of population size on these aspects is less understood. To assess the sources of population size variation, but also its impact on genetic, functional trait, and performance aspects, we conducted detailed population size estimations, assessed 23 abiotic and biotic environmental habitat factors, performed population genetic analyses using nine microsatellite markers, and recorded nine functional traits based on 260 Trifolium montanum individuals from 13 semi-dry grassland locations of Central Europe. Modern statistical analyses based on a multivariate framework (path analysis) with preselected linear regression models revealed that the variation of abiotic factors (in contrast to factors per se) almost completely, significantly explained fluctuations in population size (R 2 = .93). In general, abiotic habitat variation (heterogeneity) was not affected by habitat area. Population size significantly explained genetic diversity (N A: R 2 = .42, H o: R 2 = .67, H e: R 2 = .43, and I: R 2 = .59), inbreeding (F IS: R 2 = .35), and differentiation (G ST: R 2 = .20). We also found that iFDCV (ITV) was significantly explained by abiotic habitat heterogeneity, and to a lesser extent by genetic diversity H e (R 2 = .81). Nevertheless, habitat heterogeneity did not statistically affect genetic diversity. This may be due to the use of selectively neutral microsatellite markers, and possibly by insufficient abiotic selective pressures on habitats examined. Small T. montanum populations in nonoptimal habitats were characterized by reduced genetic and functional trait diversity, and elevated genetic inbreeding and differentiation. This indicates reduced adaptability to current and future environmental changes. The long-term survival of small populations with reduced genetic diversity and beginning inbreeding will be highly dependent on habitat protection and adequate land-use actions.

Development of microsatellite markers for Anadenanthera colubrina var. cebil (Fabaceae), a native tree from South America.

PREMISE OF THE STUDY: Microsatellite primers were developed in the native legume tree Anadenanthera colubrina var. cebil to study the genetic diversity and genetic structure in natural populations in Argentina. METHODS AND RESULTS: Nine microsatellite markers were identified using a genomic library enriched for tandemly repeated motifs, eight of which markers were polymorphic. The polymorphism of these markers was assessed by investigating 20 individuals for fragment polymorphism; three to 13 alleles were observed for each locus. Observed and expected heterozygosities ranged from 0.300 to 1.000 and from 0.463 to 0.900, respectively. CONCLUSIONS: These results confirm that these primers will be useful for investigating the genetic diversity and genetic structure of natural populations of A. colubrina var. cebil in future studies.

Traditional medicinal knowledge of woody species across climatic zones in Benin (West Africa).

ETHNOPHARMACOLOGICAL RELEVANCE: Plant parts are often used by local people to treat their affections. This study addressed the Traditional Medicinal Knowledge of woody species in Benin and the dependence of medicinal use of woody species on climatic zones. AIM OF THE STUDY: It reports (i) the main diseases categories treated with medicinal use of woody species in Benin and changes across climatic zones by inferring their epidemiological status, and (ii) the woody species involved and their distribution according to climate conditions. MATERIALS AND METHODS: Ethnobotanical interviews were undertaken using a semi-structured questionnaire. Five hundred and ninety medicinal plant professionals (healers, traders …) were interviewed in the whole country. Frequency of citation and informant consensus factor were calculated to highlight the main human diseases categories and woody species used for their treatment. A principal component analysis was performed to determine the occurrence of diseases categories in different climatic zones. RESULTS: About 94% of international diseases categories were treated using medicinal woody species in Benin. Nighty-seven ailments in 16 diseases categories were identified. Among them, 5 diseases categories (General and unspecified, Digestive, Skin, Neurological, and Musculoskeletal) were highlighted as important. The Sudano-Guinean zone showed the highest diseases frequencies, whereas the Sudanian zone showed the lowest. The epidemiological status of some phytodistricts was worrisome. In our study, 123 woody species belonging to 93 genera and 35 families were reported, and among them, 16 were the most used as treatments. CONCLUSIONS: There is a lack of consensus among traditional healers about which woody species to use. Many different species were used to treat a given diseases category. Also, information concerning their organ composition was not available in the literature, for the majority of species. Biological and chemical investigations are thus needed for a better valorization of the most frequently used plants in the future.

Plant intraspecific functional trait variation is related to within-habitat heterogeneity and genetic diversity in Trifolium montanum L.

Intraspecific trait variation (ITV), based on available genetic diversity, is one of the major means plant populations can respond to environmental variability. The study of functional trait variation and diversity has become popular in ecological research, for example, as a proxy for plant performance influencing fitness. Up to now, it is unclear which aspects of intraspecific functional trait variation (iFDCV) can be attributed to the environment or genetics under natural conditions. Here, we examined 260 individuals from 13 locations of the rare (semi-)dry calcareous grassland species Trifolium montanum L. in terms of iFDCV, within-habitat heterogeneity, and genetic diversity. The iFDCV was assessed by measuring functional traits (releasing height, biomass, leaf area, specific leaf area, leaf dry matter content, Fv/Fm, performance index, stomatal pore surface, and stomatal pore area index). Abiotic within-habitat heterogeneity was derived from altitude, slope exposure, slope, leaf area index, soil depth, and further soil factors. Based on microsatellites, we calculated expected heterozygosity (He) because it best-explained, among other indices, iFDCV. We performed multiple linear regression models quantifying relationships among iFDCV, abiotic within-habitat heterogeneity and genetic diversity, and also between separate functional traits and abiotic within-habitat heterogeneity or genetic diversity. We found that abiotic within-habitat heterogeneity influenced iFDCV twice as strong compared to genetic diversity. Both aspects together explained 77% of variation in iFDCV ( R adj 2  = .77, F 2, 10 = 21.66, p < .001). The majority of functional traits (releasing height, biomass, specific leaf area, leaf dry matter content, Fv/Fm, and performance index) were related to abiotic habitat conditions indicating responses to environmental heterogeneity. In contrast, only morphology-related functional traits (releasing height, biomass, and leaf area) were related to genetics. Our results suggest that both within-habitat heterogeneity and genetic diversity affect iFDCV and are thus crucial to consider when aiming to understand or predict changes of plant species performance under changing environmental conditions.

Desert-like badlands and surrounding (semi-)dry grasslands of Central Germany promote small-scale phenotypic and genetic differentiation in Thymus praecox.

Environmental heterogeneity among sites can generate phenotypic and genetic variation facilitating differentiation and microevolution of plant populations. Badlands are desert-like, predominantly vegetation-poor habitats often embedded in (semi-)dry grasslands. The desert-like conditions of badlands demand extreme adaptation of plants, that is, phenotypic modifications in short-term and/or natural adaptation in long-term. However, detailed knowledge is missing about both plant phenotypic and genetic differentiation in this unique and widely occurring habitat type. The present study focused on the largest known badlands systems in Central Europe located in the "Drei Gleichen" region, a designated nature conservation area in Central Germany. Locations were suitable for this study in terms of having co-occurring badlands and (semi-)dry grassland habitats (sites) occupied by the pioneer plant Thymus praecox. Here, we studied the environmental preferences, morphological and functional trait variation, and genetic variation using microsatellite markers of T. praecox. Results revealed significant, mainly site-dependent environmental, phenotypic, and genetic differentiation. In general, individuals in badlands are shorter in height and have lower patch sizes (length × width), relative growth rates, and smaller stomata. The PCA additionally unveiled slightly increased leaf robustness, trichome density, decreased stomatal conductance, fewer females, and earlier phenology in badlands. We interpret differentiation patterns as adaptive responses to light, temperature, drought, and nutrient stress conditions supported by reviewed literature. Genetic differentiation was strongest between local badlands and grassland sites, and clearly weaker among locations and between sites (in total) as indicated by G ST, AMOVA, PCoA, and population structure. Our study supports the importance of small-scale microhabitat conditions as a driver of microevolutionary processes, and the population's need for sufficient phenotypic variation and genetic resources to deal with environmental changes. We demonstrated that badlands are an appropriate model system for testing plant response to extreme habitats and that more research is needed on these fascinating landscapes.

Characterization of 15 nuclear microsatellite markers for Afzelia africana (Fabaceae) and related species.

PREMISE: Afzelia africana (Fabaceae) is a valuable, internationally vulnerable tree species in tropical Africa. The development of specific simple sequence repeat (SSR) loci is necessary for population genetic studies in this tree species and its closest relatives. METHODS AND RESULTS: Fifteen new polymorphic microsatellite markers were developed for A. africana using Illumina next-generation sequencing. We tested the polymorphism of the 15 loci in three populations in Benin, West Africa. The number of expressed alleles per locus varied from one to 12. The levels of observed and expected heterozygosity ranged from 0.100 to 1.000 and from 0.095 to 0.882, respectively. Most markers successfully amplified in the closely related species A. quanzensis and A. bipindensis, but less so in A. bipindensis. CONCLUSIONS: Because of their cross-amplification ability, these newly developed loci will serve as useful tools for future molecular analyses on A. africana and related species.

Genetic diversity and structure of baobab (Adansonia digitata L.) in southeastern Kenya.

Baobab (Adansonia digitata L.) is an iconic tree of African savannahs. Its multipurpose character and nutritional composition of fruits and leaves offer high economic and social potential for local communities. There is an urgent need to characterize the genetic diversity of the Kenyan baobab populations in order to facilitate further conservation and domestication programmes. This study aims at documenting the genetic diversity and structure of baobab populations in southeastern Kenya. Leaf or bark samples were collected from 189 baobab trees in seven populations distributed in two geographical groups, i.e. four inland and three coastal populations. Nine microsatellite loci were used to assess genetic diversity. Overall, genetic diversity of the species was high and similarly distributed over the populations. Bayesian clustering and principal coordinate analysis congruently divided the populations into two distinct clusters, suggesting significant differences between inland and coastal populations. The genetic differentiation between coastal and inland populations suggests a limited possibility of gene flow between these populations. Further conservation and domestications studies should take into consideration thegeographical origin of trees and more attention should be paid to morphological characterization of fruits and leaves of the coastal and inland populations to understand the causes and the impact of the differentiation.

Characterization and transferability of microsatellite markers developed for Carpinus betulus (Betulaceae)(1).

PREMISE OF THE STUDY: Carpinus betulus (Betulaceae) is an octoploid, ecologically important, common tree species in European woodlands. We established 11 nuclear microsatellite loci allowing for detailed analyses of genetic diversity and structure. METHODS AND RESULTS: A microsatellite-enriched library was used to develop primers for 11 microsatellite loci that revealed high allele numbers and genetic diversity in a preliminary study. CONCLUSIONS: All of the loci developed here are informative for C. betulus. In addition, the loci are transferable to several species within the genus, and almost all loci cross-amplified in species of different genera of the Betulaceae.

Genetic diversity in aspen and its relation to arthropod abundance.

The ecological consequences of biodiversity have become a prominent public issue. Little is known on the effect of genetic diversity on ecosystem services. Here, a diversity experiment was established with European and North American aspen (Populus tremula, P. tremuloides) planted in plots representing either a single deme only or combinations of two, four and eight demes. The goals of this study were to explore the complex inter- and intraspecific genetic diversity of aspen and to then relate three measures for diversity (deme diversity, genetic diversity determined as Shannon index or as expected heterozygosity) to arthropod abundance. Microsatellite and AFLP markers were used to analyze the genetic variation patterns within and between the aspen demes and deme mixtures. Large differences were observed regarding the genetic diversity within demes. An analysis of molecular variance revealed that most of the total genetic diversity was found within demes, but the genetic differentiation among demes was also high. The complex patterns of genetic diversity and differentiation resulted in large differences of the genetic variation within plots. The average diversity increased from plots with only one deme to plots with two, four, and eight demes, respectively and separated plots with and without American aspen. To test whether intra- and interspecific diversity impacts on ecosystem services, arthropod abundance was determined. Increasing genetic diversity of aspen was related to increasing abundance of arthropods. However, the relationship was mainly driven by the presence of American aspen suggesting that species identity overrode the effect of intraspecific variation of European aspen.

A unifying study of phenotypic and molecular genetic variability in natural populations of Anadenanthera colubrina var. cebil from Yungas and Paranaense biogeographic provinces in Argentina.

Anadenanthera colubrina var. cebil is a discontinuously distributed native tree species in South American subtropical forests. Thirteen quantitative traits and eight nuclear microsatellite loci were examined in individuals from two biogeographic provinces of Argentina to determine the number and composition of genetically distinguishable groups of individuals and explore possible spatial patterns of the phenotypic and genetic variability. Means of reproductive traits were higher in the Yungas than in the Paranaense biogeographic province, whereas five out of eight nonreproductive quantitative traits showed higher mean values in the latter. Variance coefficients were moderate, and there were significant differences between and within provinces. Three clusters were defined based on spatial model for cluster membership for quantitative traits. One cluster grouped the individuals from the Paranaense biogeographic province whereas the individuals from the Yungas biogeographic province grouped regarding its population of origin. Parameters of molecular genetic variability showed higher values in the Yungas than in the Paranaense biogeographic province. Observed heterozygosity was lower than expected heterozygosity in both biogeographic provinces, indicating an excess of homozygosity. The homozygosity test by Watterson and the exact test by Slatkin suggested diversifying selection for locus Ac41.1. Bayesian clustering spatial model for microsatellites loci data were performed for both all loci and for all loci excluding locus Ac41.1. In both analyses two clusters were inferred. Analysis of molecular variance revealed similar results for all genotypes and for all genotypes defined excluding locus Ac41.1. Most of the total variance is attributable to genetic variation within clusters. The presence of homogeneous clusters was detected for both the phenotypic and molecular genetic variability. Two Bayesian clustering analyses were performed according to molecular genetic data, and two clusters were inferred. Individuals were assigned to their provinces of origin. Genetic molecular variation was higher in the populations of the Yungas biogeographic province which translates into highly qualified populations for conservation. Populations from the Paranaense biogeographic province showed the highest mean value of number of seeds per fruit making them valuable as well with regard to the exploitation of management strategies as a means to recover the impacted areas where these populations are located.

Microsatellite markers for the tetraploid halophyte Suaeda maritima (L.) Dumort. (Chenopodiaceae) and cross-species amplification in related taxa.

We developed 12 polymorphic microsatellite markers for the tetraploid halophyte Suaeda maritima (Chenopodiaceae). Population genetic parameters were estimated for three populations from different habitats (coastal and inland), using the program Tetrasat. Between two and 15 alleles per locus were observed. Mean expected heterozygosities (H(E) ) and Shannon-Wiener Diversity Indices (H') per locus and population ranged from zero to 0.852, and from zero to 2.990, respectively. The two inland populations were less diverse than the coastal one at most of the loci. All markers cross-amplified in the closely related Suaeda salsa, and all but one were transferable to Suaeda spicata and Suaeda salinaria.

Microsatellite markers for Spergularia media (L.) C. Presl. (Caryophyllaceae) and their cross-species transferability.

We developed 11 polymorphic microsatellite markers for Spergularia media (Caryophyllaceae), a perennial halophyte of coastal salt meadows and continental areas of western Eurasia. The number of alleles per locus observed in a single population of 20 individuals from the German North Sea coast ranged from 3 to 20. Observed and expected heterozygosities ranged from 0.200 to 0.850 and from 0.278 to 0.936 respectively. Observed heterozygosities were lower than expected heterozygosities at all loci, presumably as a consequence of inbreeding. All markers cross-amplified in the closely related S. salina. Of these, nine were polymorphic in the heterologous species. Only two primer pairs generated PCR products in S. diandra.

Isolation and characterization of 11 new microsatellite markers for Macaranga (Euphorbiaceae).

We provide primer sequences for 11 new polymorphic microsatellite markers developed in the tropical ant-plant genus Macaranga (Euphorbiaceae), after enrichment cloning of Macaranga tanarius and Macaranga hypoleuca. Allele numbers per locus ranged from two to 16 among 20 accessions of M. tanarius, and from three to 10 among 22 accessions of M. hypoleuca. Observed and expected heterozygosities ranged from 0.150 to 0.900 and from 0.375 to 0.894 in M. tanarius, and from 0.545 to 1.000 and from 0.434 to 0.870 in M. hypoleuca, respectively. Six of the 11 primer pairs successfully cross-amplified polymorphic polymerase chain reaction products in Macaranga winkleri.