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.     

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.     

Development of microsatellite markers for Anadenanthera colubrina (Leguminosae), a neotropical tree species

? Premise of the study: We developed and characterized nuclear microsatellite markers for Anadenanthera colubrina, a tropical tree species widely distributed in South America. ? Methods and Results: Leaf samples of mature A. colubrina trees, popularly called "angico," were collected from an area that is greatly impacted by agricultural practices in the region of Ribeir?o Preto in S?o Paulo State in southeastern Brazil. Twenty simple sequence repeat (SSR) markers were developed, 14 of which had polymorphic loci. A total of 96 alleles were detected with an average of 6.86 alleles per polymorphic locus. The expected heterozygosity, calculated at polymorphic loci, ranged from 0.18 to 0.83. Finally, we demonstrated that 18 loci were cross-amplified in A. peregrina. ? Conclusions: A total of 14 polymorphic markers suggest a high potential for genetic diversity, gene flow, and mating system analyses in A. colubrina.     

Hallucinogenic snuff from Northwestern Argentina: Microscopical identification of anadenanthera colubrina var. cebil (fabaceae) in powdered archaeological material

Two mummified bodies have been recovered from the Alero I or La Matanza site (15 km from Cusi-Cusi, Dpto. Santa Catalina, Puna of Jujuy, Argentina) in association with characteristic snuffing paraphernalia. The purpose of this work is to identify the powdered material contained in one of the tubes in order to confirm the hypothesis generated from ethnographical sources, that the rape (snuff) complex was associated with the consumption of Anadenanthera colubrina var. cebil, Fabaceae, the cebil, or, if not, which materials were inhaled. Following analysis with a light microscope, petrographic microscope and scanning electron microscope the unknown material was identified as cebil by comparison with reference material and specialized literature.

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Rapé AlucinóGeno del Noroeste de Argentina: identificación microscópica de Anadenanthera colubrina VAR. cEBIL (fABACEAE) en material arqueológico pulverizado

Resumen  CEBIL (FABACEAE) EN MATERIAL ARQUEOLóGICO PULVERIZADO. En 1991 se recuperaron dos cuerpos momificados en el Alero I La Matanza (distante 15 km de Cusi-Cusi, Dpto. Santa Catalina, en la Puna de Jujuy, Argentina) en asociación con el ajuar caracter#x00ED;istico del complejo del rap#x00E9; o complejo alucinógeno. Uno de los tubos de hueso hallados contenta material pulverizado. El objeto de este trabajo es identificar esta muestra con el objeto de contrastar la hipótesis generada a partir de estudios etnogr#x00E1;fico que asocia el complejo del rapé con el consuma de Anadenanthera colubrina var. cebil, Fabaceae, el cebil, o en caso negativo, identificar el material empleado. Mediante análisis con microscopio fotónico, microscopio petrogr#x00E1;fico y microscopio electrónico de barrido se identific#x20F3; la muestra como cebil por comparación con material de referenda y bibliograf#x00ED;a especializada.

     

Genetic structure of Alhagi (Hedysareae,Fabaceae) populations using ISSR data in Iran

Molecular Biology Reports - Alhagi Gagnebin (Fabaceae: Hedysareae) is a small genus of shrubs or subshrubs distributed in temperate and tropical regions of Asia, Europe, and Africa. The exact...     

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.     

Genetic diversity within and among two Argentinean and one Mexican species of Acacia (Fabaceae)

Acacia is a pantropical genus comprising > 1450 species. Following Vassal's treatment Acacia is considered as a single genus with three subgenera (Acacia, Aculeiferum and Phyllodineae). Acacia caven, A. curvifructa and A. farnesiana belong to subgenus Acacia and the relationship between them is controversial. The aim of this study was to elucidate the relationship between the three species using amplified fragment length polymorphism, analysing 15 populations of these species, and to compare the results obtained with those from a morphological analysis. Genetic diversity indices (percentage of polymorphic loci, genetic diversity) showed that genetic variation in A. caven is higher than that in A. curvifructa and A. farnesiana. Of the total genetic diversity in A. caven and A. farnesiana, most is found within populations (∼70%). Analysis with STRUCTURE showed that the optimal number of clusters (K) was ten, and in all cases where populations were grouped they were geographically close and/or belong to the same variety. The morphological canonical discriminant analysis did not result in a separation between all individuals, indicating that they do not harbour consistent morphological discontinuities. Altogether, the results of our molecular analyses showed the existence of significant differences between A. caven, A. curvifructa and A. farnesiana, which argues for recognizing them as different species. © 2015 The Linnean Society of London, Botanical Journal of the Linnean Society, 2015, 177 , 593–606.     

Isolation and biodiversity in Cytisus villosus Pourret (Fabaceae,Genisteae): enzyme polymorphism in disjunct populations

ABSTRACT

The genetic diversity of isolated populations of Cytisus villosus has been studied by means of enzyme polymorphism analysis. Two types of isolated populations were studied: “terrestrial islands” in Sicily, and “true islands” in the Aeolian archipelago. In the populations of “true islands” the number of alleles and the heterozygosity are lower than in “terrestrial islands”. Isolation amongst Sicilian populations seems to be more recent than isolation of the Aeolian populations, and may be attributed to climatic changes which occurred during the Holocene and/or to human activities. The disjunction of the Aeolian populations seems much more recent than the origin of the isles themselves; the colonization of the archipelago is attributed to a single, recent dispersal event not followed by local evolution. In view of the biological structure of the Aeolian populations, C. villosus must be regarded as a locally endangered species.     

Genetic structure and mating systems in wild and cultivated populations of Phaseolus coccineus and P. vulgaris (Fabaceae)

We present a study of the genetic variation, genetic structure, and outcrossing rate of two wild populations of Phaseolus coccineus ssp. formosus (both from Central Mexico), two cultivated populations of P. coccineus ssp. coccineus (one from Central Mexico, the other from Chiapas), and one cultivated population of P. vulgaris (from Central Mexico), using seven electrophorectic loci. All the populations of P. coccineus showed high and similar levels of genetic variation, without differences among wild and cultivated populations (H ranged between 0.187 to 0.271). In contrast, P. vulgaris has low levels of genetic variation (H = 0.041). The three P. coccineus populations from Central Mexico were genetically similar, while the cultivated P. coccineus from Chiapas and the cultivated P. vulgaris from Central Mexico were very different. In all the loci and all the populations for both species, the fixation indices (F) were positive and differed significantly from random mating expectation in 59% of the cases for P. coccineus. The highest F was the estimated for P. vulgaris. For P. coccineus, the multilocus outcrossing rate estimates ranged from 0.592 to 0.698, and these did not differ significantly among populations. For P. coccineus, we also estimated the outcrossing rate for each of 60 different families, showing great variation, with a majority of families with a t between 0.8 and 1, but a substantial fraction of the families with a t as low as 0.12–0.20. The domestication process has neither eroded the levels of genetic variation of the cultivated populations of P. coccineus nor changed its mating system.     

Genetic structure of refugial populations of the temperate plant Shortia rotundifolia (Diapensiaceae) on a subtropical island

Continental island systems harbour relict biota and populations that might have migrated during glacial periods due to the formation of landbridges. Here we analysed the genetic structure of relict populations of the temperate plant Shortia rotundifolia on the subtropical island of Iriomotejima, Japan. This plant, which inhabits riparian environments, is designated “near threatened”. Only five extant populations have been found on the island. Our analyses of 10 nuclear microsatellite loci detected genetic diversity of H _E = 0.488 and H _O = 0.358 for all populations of S. rotundifolia on the island. A high inbreeding coefficient for all populations together (F _IS = 0.316) and each population separately (F _IS = 0.258–0.497) might be attributable to crossing among closely related descendants within a population, an idea that is supported by the relatedness coefficient. These results and an examination of the populations’ demographic histories suggest that the extant populations on Iriomotejima have not experienced a recent population bottleneck. The five extant populations were genetically differentiated (F _ST = 0.283; P < 0.001), suggesting low seed dispersal by gravity and/or low pollen flow via pollinators in the riparian environment. In addition, population differentiation was not related to genetic distance, implying that at one time, ancestral populations might have been distributed over a wider area of the island. However, population fragmentation and range contraction might have occurred at random during the postglacial period.     

Offspring performance and recruitment of the pioneer tree Acacia caven (Fabaceae) in a fragmented subtropical dry forest

The process of habitat fragmentation results in the breaking apart of originally continuous habitats, causing multiple changes in biotic and abiotic interactions. Alterations in resource availability and in mutualistic and antagonistic plant–animal interactions may impact plant offspring quantity and quality. Currently, several old fragmented systems evidence a process of flora homogenization, where shade‐tolerant species are replaced by pioneer light‐demanding species. Notably, the relationship between quantity and quality parameters of plant offspring production and the successful recruitment of pioneer species in fragmented forests has been poorly explored. Here, we assess population size, sapling recruitment and offspring performance of one of the most widespread tree species of subtropical South America, the native pioneer Acacia caven (Fabaceae). Population size of adults and saplings increased from small to continuous forests, whereas the sapling recruitment per adult tree (sapling/adult ratio) showed no significant differences among forests of different size. Seedling performance was negatively related to forest area and population size, implying potential superior competitive ability of seedlings produced in smaller populations compared to larger ones. Our results show that A. caven is resilient to habitat fragmentation effects, which may be ascribed to a set of advantageous ecological traits such as outcrossing, massive flowering, generalist pollination, drought resistance, rapid growth and re‐sprouting. Thus, this pioneer tree benefits from the availability of vacant sites and resources released by declining plant populations of other species, eventually becoming the dominant species in fragmented habitats. Pioneer native plant species with ecological traits such as A. caven may represent the silent successful survivors and new colonizers of fragmented habitats, the ubiquitous landscapes of the future.     

Genetic diversity in fragmented populations of Berchemiella wilsonii var. pubipetiolata (Rhamnaceae)

BACKGROUD AND AIMS: Berchemiella wilsonii var. pubipetiolata (Rhamnaceae) is distributed in fragmented habitat patches in eastern China. It is highly endangered because of severe disturbance by anthropogenic activities. Information on genetic variation and structure is critical for developing successful conservation strategies for this species. METHODS: Allozyme variation of population genetic diversity and structure was investigated for a total of 98 individuals sampled from four extant populations using isoelectric focusing in thin-layer polyacrylamide slab gels. KEY RESULTS: Based on 20 loci scored from the nine enzymes examined, a high genetic diversity was detected at both the species and population level, while there was a loss of low frequency alleles (<0.1) in all populations. Most loci showed deviation from Hardy-Weinberg equilibrium due to excess of heterozygotes in all populations, suggesting that selection for heterozygotes has occurred in this species. The genetic diversity was mainly found within populations with a moderate genetic differentiation (F(ST) = 0.13), but the two geographically discontinuous population groups showed significant differences, with F-statistic values of 0.078 for the Zhejiang populations and 0.014 for the Anhui populations, respectively. CONCLUSIONS: It appears most likely that this species has experienced a recent decrease in population size, and genetic drift in small populations has resulted in a loss of alleles occurring at low frequency. The differentiation into two population groups reflects a population genetic consequence that has been influenced by the different land-use in the two regions. Some conservation concerns are discussed together with possible strategies for implementing in situ and ex situ conservation.     

Genetic relationships betweenLupinus pilosus andL. palaestinus (Fabaceae)

Lupinus pilosus Murr. andL. palaestinus Boiss., are both East Mediterranean annuals, greatly resemble each other in morphology, and have the same chromosome number (2n = 42). Some fertile hybrids can be formed by artificial hybridization or under unusual habitat conditions. Isolation in nature between the species is, however, maintained by prevalent selfing, partial reproductive barriers and genic imbalance, together with differences in ecological requirements. It is suggested that this speciation pattern which includes little morphological and hardly any chromosomal divergence might have occurred in other cases of edaphic vicariants too.     

Genetic variation and biogeography of the disjunct Vitis subg. Vitis (Vitaceae)

Aim Vitis subg. Vitis provides an example of a plant disjunction occurring in the Northern Hemisphere. It shows broad morphological variation but is assumed to be a species complex with limited genetic differentiation. Based on a comprehensive sampling of taxa and polymorphism in both chloroplast and nuclear DNA, we assessed genetic variation within this subgenus. Our aims were to clarify the relationships among species and to examine their historical biogeography. Location Asia, Europe, North America. Methods We analysed a total of 30 species and putative hybrids from subgenus Vitis and examined the infra‐specific variation in some species. Polymorphism in chloroplast DNA was assessed in trnL and trnH–psbA–trnK sequences (c. 2170 bp) and in 15 microsatellite loci. We also obtained nuclear data for size variation at 24 microsatellite loci. Phylogenetic inference was performed with Bayesian analyses. A maximum parsimony network was constructed to depict the evolutionary relationships among haplotypes, and microsatellite data were also subjected to hierarchical clustering analysis using the Ward distance. In addition, we assessed size homoplasy by sequencing both chloroplast and nuclear microsatellite loci. Results Chloroplast polymorphisms resolved subgenus Vitis as a monophyletic group with limited genetic variation. The ancestral haplotypes were found in Eurasia. American taxa all harboured derived haplotypes. Most of them formed a monophyletic group that did not include Vitis californica. The four main haplotypes in Vitis vinifera corresponded to two different origins. Nuclear microsatellites indicated that genetic variation was especially large in North America. Asian species exhibited a lower level of nuclear divergence and the European V. vinifera corresponded to a differentiated nuclear lineage. Main conclusions We obtained some evidence that subgenus Vitis has an Asian origin and then dispersed to Europe and North America. Geographic separation was followed by diversification, presumably during the Pleistocene, resulting in phylogeographic patterns similar to other biota. In contrast to chloroplast DNA, nuclear DNA shows a larger than expected genetic variation. Our molecular data also highlight the need to re‐examine certain aspects of the current subgeneric classification.     

Genetic similarity of disjunct populations of the giant sea bass Stereolepis gigas

Sequence variation in nuclear and mitochondrial genes of the giant sea bass Stereolepis gigas collected from the Pacific coast and the northern Sea of Cortez was examined. Restriction fragment length polymorphism analysis and direct sequencing showed extremely low mtDNA sequence diversity (13 closely related haplotypes with no evidence of geographical population subdivision). The mitochondrial haplotype mismatch distribution is consistent with a population expansion following the Last Pleistocene glaciation. Differences in single nucleotide polymorphism frequencies between Pacific and Sea of Cortez populations were detected at two of four nuclear loci, which may reflect natural selection or genetic drift in populations with low effective numbers of males. Although Pacific coast and Sea of Cortez populations of giant sea bass do not exhibit the mitochondrial phylogenetic break characteristic of many species with disjunct Pacific and Gulf populations, the possibility of genetic differentiation at nuclear loci suggests that a cautious approach to broodstock selection for captive breeding and restoration programmes be exercised.     

The free reducing oligosaccharides of angico branco (Anadenanthera colubrina) gum exudate: an aid for structural assignments in the heteropolysaccharide

A novel method is described for the determination of sequential side-chain structures in the complex, high-arabinose polysaccharide of the gum exudate of angico branco (Anadenanthera colubrina), using as basis the structurally similar reducing oligosaccharides present in small quantities. Of the ten detected, eight were characterized as disaccharides (2, 3, and 9), linear trisaccharides (1 and 4), branched pentasaccharides (5 and 6), and a doubly branched heptasaccharide (8). The oligosaccharides are substituents of the polysaccharide, which has a (1-->3)-linked beta-D-galactopyranosyl main chain, and with two exceptions they had 6-O-substituted galactopyranosyl reducing ends, probably corresponding to its main-chain units. Characterization was effected through their 1D and 2D NMR correlation spectra, which were better resolved and more readily interpretable than those of the polysaccharide. These spectral data were supported by monosaccharide composition and rotation values. Controlled Smith degradations and methylation analyses were carried out when it was necessary. These data were confirmed by field-desorption MS.     

Large-scale phylogeography of the disjunct Neotropical tree species Schizolobium parahyba (Fabaceae-Caesalpinioideae)

Neotropical rainforests exhibit high levels of endemism and diversity. Although the evolutionary genetics of plant diversification has garnered increased interest, phylogeographic studies of widely distributed species remain scarce. Here we describe chloroplast and nuclear variation patterns in Schizolobium parahyba (Fabaceae), a widespread tree in Neotropical rainforests that harbor two varieties with a disjunct distribution. Chloroplast and nuclear sequence analyses yielded 21 and 4 haplotypes, respectively. Two genetic diversity centers that correlate with the two known varieties were identified: the Southeastern Atlantic forest and the Amazonian basin. In contrast, the populations from southern and northeastern Atlantic forests and Andean-Central American forests exhibited low levels of genetic diversity and divergent haplotypes, likely related to historical processes that impact the flora and fauna in these regions, such as a founder's effect after dispersion and demographic expansion. Phylogeographic and demographic patterns suggest that episodes of genetic isolation and dispersal events have shaped the evolutionary history for this species, and different patterns have guided the evolution of S. parahyba. Moreover, the results of this study suggest that the dry corridor formed by Cerrado and Caatinga ecoregions and the Andean uplift acted as barriers to this species' gene flow, a picture that may be generalized to most of the plant biodiversity tropical woodlands and forests. These results also reinforce the importance of evaluating multiple genetic markers for a more comprehensive understanding of population structure and history. Our results provide insight into the conservation efforts and ongoing work on the genetics of population divergence and speciation in these Neotropical rainforests.     

Population Genetic structure ofPotentilla fragariodes var.major (Rosaceae) in Korea

The genetic diversity and population structure of eighteenPotentilla fragariodes var.major (Rosaceae) populations in Korea were determined using genetic variations at 22 allozyme loci. The percent of polymorphic loci within the enzymes was 66.7%. Genetic diversity at the species level and at the population level was high (Hes = 0.203; Hep = 0.185, respectively), whereas the extent of the population divergence was relatively low (G_ST = 0.069). F_IS, a measure of the deviation from random mating within the 18 populations, was 0.075. An indirect estimate of the number of migrants per generation (Nm = 3.36) indicated that gene flow was high among Korean populations of the species. In addition, analysis of fixation indices revealed a slight heterozygote deficiency in some populations and at some loci. Wide geographic ranges, perennial herbaceous nature and the persistence of multiple generations are associated with the high level of genetic variation. AlthoughP. fragariodes var.major usually propagated by asexually-produced ramets, we could not rule out the possibility that sexual reproduction occurred at a low rate because each ramet may produce terminal flowers. Mean genetic identity between populations was 0.983. It is highly probable that directional movement toward genetic uniformity in a relatively homogeneous habitat operates among Korean populations ofP. fragariodes var.major.