Quantifying gene flow from spatial genetic structure data in a metapopulation of Chamaecrista fasciculata (Leguminosae)

Abstract An extensive allozyme survey was conducted within a natural "meta" population of the native North American annual legume, Chamaecrista fasciculata (Leguminosae) to quantify genetic structure at different spatial scales. Gene flow was then estimated by a recently developed indirect method based on a continuous population model, using pairwise kinship coefficients between individuals. The indirect estimates of gene flow, quantified in terms of neighborhood size, with an average value on the order of 150 individuals, were concordant among different spatial scales (subpopulation, population, metapopulation). This gene-flow value lies within the range of direct estimates previously documented from observations of pollen and seed dispersal for the same metapopulation. Monte Carlo simulations using the direct measures of gene flow as parameters further demonstrated that the observed spatial pattern of allozyme variation was congruent with a model of isolation by distance. Combining previously published estimates of pollen dispersal distances with kinship coefficients from this study, we quantified biparental inbreeding relative to either a single subpopulation or the whole metapopulation. At the level of a neighborhood, little biparental inbreeding was observed and most departure from Hardy-Weinberg genotypic proportions was explained by self-fertilization, whereas both selfing and biparental inbreeding contributed to nonrandom mating at the metapopulation level. Gene flow was also estimated from indirect methods based on a discontinuous population structure model. We discuss these results with respect to the effect of a patchy population structure on estimation of gene flow.     

Multiple pleurograms in Chamaecrista Moench (Leguminosae, Caesalpinioideae)

According to the literature, the seeds of Chamaecrista lack pleurograms and have pitted testas aligned in vertical lines. Preliminary observations have revealed that these small structures consist of superficial alterations of the testa that are coloured differently to the rest of the seed coat and stand out as pits. However, no anatomical inspection has yet determined the precise composition of these pits. The present work examined the morphology, surface micromorphology, anatomy, and histochemistry of the seed coat of mature seeds of C. desvauxii var. latistipula , C. flexuosa , and C. nictitans var. patellaria in order to elucidate the structural nature of the seed coat pits. For this, seed coats were studied using scanning electron microscopy and light microscopy. Structural modifications were examined in the seed coats of the studied species, and variations were noted in the mucilaginous layer, in the thickness of the palisade layer, and in the presence of larger hypodermic cells. Based on the anatomical features observed, and comparing these with published data on pleurograms in the Caesalpinioideae, it was determined that the structures referred to previously as tegumental pits on Chamaecrista seeds are actually small and numerous pleurograms, reported here for the first time.  © 2008 The Linnean Society of London, Botanical Journal of the Linnean Society , 2008, 157 , 487–492.     

Environmental contribution to floral trait variation in Chamaecrista fasciculata (Fabaceae: Caesalpinoideae)

Although intraspecific variation in plant floral traits has been documented for a number of plant species, the causes of such variation are largely unknown. We first quantified floral trait variation in an Illinois prairie population of Chamaecrista fasciculata Michx. We then used a field experiment to determine the contribution of leaf herbivory to this variation and a greenhouse experiment to determine the contribution of leaf herbivory, and variable soil nutrient and water content to floral trait variation. Variation in environmental factors explained a significant portion of the naturally occurring variation in corolla width, ovule number, ovule size, and anther length. In the field, manual removal of 25% or more leaf area reduced ovule size and anther length (and by inference, pollen production), and delayed flowering. In the greenhouse, plants from which we removed 25% or more of their leaf area or which were given limited water produced fewer ovules than control plants. Addition of nutrients interacted with soil moisture to affect corolla diameter and ovule number. Despite our demonstration of significant environmental impacts on reproductive traits, these impacts were relatively much smaller than those on plant size, suggesting that floral traits are buffered against variable resource availability.     

Two new Brazilian species of Chamaecrista Moench (Leguminosae: Caesalpinioideae)

Chamaecrista (sect.Apoucouita) onusta Irwin & Barneby from coastal Bahia andCh. (sect.Absus) hatschbachii Irwin & Barneby from north-central Minas Gerais are described, figured and systematically characterized. Four new combinations inChamaecrista incidental to the discussion are proposed.     

Seed ontogeny of Chamaecrista and its systematic implications in Cassiinae (Leguminosae, Caesalpinioideae)

The genus Chamaecrista originates from the dismembering of Cassia L., which was elevated to the level of subtribe Cassiinae, giving rise to Cassia sensu stricto, Chamaecrista, and Senna. Phylogenetic analysis has suggested that Cassiinae is not monophyletic. The relationships between its genera are not clear, because they sometimes appear close to Chamaecrista and Senna and at other times close to Senna and Cassia. We describe here development of the seeds of three species of Chamaecrista and compare our data with information published in the literature for Cassiinae and non-Cassiinae legumes. The ovules appear well conserved in Cassiinae, differing only in the length of the funiculus. In terms of seed development, the shape of the endosperm haustorium is similar in Senna and Chamaecrista. Chamaecrista shares more characteristics of its mature seeds with Senna than with Cassia. Chamaecrista and Senna have variable seed shapes, the presence of pleurograms, the absence of root cap primordia, a short epicotyl, and the cotyledonary mesophyll tends to be dorsiventral. These seed characteristics place Senna close to Chamaecrista and distance it from Cassia. We also observed characteristics unique to Chamaecrista, for example the occurrence of multiple pleurograms, a partially multiplicative seed coat, and the presence of embryonic colleters.     

Population variation in plant traits associated with ant attraction and herbivory in Chamaecrista fasciculata (Fabaceae)

The benefits of ant–plant–herbivore interactions for the plant depend on the abundance of ants and herbivores and the selective pressures these arthropods exert. In plants bearing extrafloral nectaries (EFN), different mean trait values may be selected for by different populations in response to local herbivore pressure, ultimately resulting in the evolution of differences in plant traits that attract ants as defensive agents against herbivory. To determine if variation in traits that mediate ant–plant interactions reflect herbivore selective pressures, we quantified intra- and inter-population variation in plant traits for eight populations of the EFN-bearing annual Chamaecrista fasciculata (Michx.) (Fabaceae). Censuses in rural and urban areas of Missouri and Illinois (USA) showed population differences in ant attendance and herbivore pressure. Seeds were collected from each population, and plants were grown in a common greenhouse environment to measure sugar production, nectar volume and composition, EFN size and time of emergence, leaf pubescence, and leaf quality throughout plant development. Populations varied mainly in terms of nectary size, sugar production, and nectar volume, but to a lesser degree in leaf pubescence. Populations of C. fasciculata within urban areas (low in insect abundance) had small nectaries and the lowest nectar production. There was a positive correlation across populations between herbivore density and leaf damage by those herbivores on the one hand and sugar production and nectar volume on the other. These results, in conjunction with lack of evidence for maternally based environmental effects, suggest that population differences in herbivore damage have promoted differential evolution of EFN-related traits among populations. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Inflated trichornes in flowers of Bauhinia (Leguminosae: Caesalpinioideae)

TUCKER, S. C., RUGENSTEIN, S. R. & DERSTINE, K. S., 1984. Inflated trichomes in flowers of Bauhinia (Leguminosae: Caesalpinioideae). An unusual kind of bifid multicellular inflated trichome is reported to occur on floral organs of II species of Bauhinia (Leguminosae-Caesalpinioideae), They occur on sepals, petals, stamens, carpels, receptacle, leaves, and stems, but not on all organs in each species. Their development in B. malabarica is described.     

Cytogenetics of species of Chamaecrista (Leguminosae – Caesalpinioideae) native to southern Brazil

Chromosome numbers, karyotypes, meiotic behaviour and pollen analysis are presented for species of Chamaecrista Moench (Leguminosae, Caesalpinioideae, Cassieae ) native to southern Brazil: C. nictitans ssp. patellaria , C. nictitans ssp. disadena , C. repens , C. rotundifolia , C. flexuosa , C. vestita and C. desvauxii . Meiotic behaviour is reported for the first time for all the taxa and was very regular; only bivalents were formed at diakinesis and metaphase I, chromosome disjunction and segregation were regular at anaphases I and II, meiotic indexes were over 99% and pollen fertility was over 92%. Pollen grains were subprolate in C. flexuosa and C. vestita and prolate–spheroidal in the other taxa. Karyotypes were symmetrical in all six species and the data are original, except for C. nictitans ssp. p atellaria. Chromosome number is presented for the first time for C. repens (2 n  = 16) and has been confirmed for the other taxa: 2 n  = 14 for C. desvauxii , 2 n  = 32 for the tetraploid C. nictitans ssp. patellaria and C. nictitans ssp. disadena , and 2 n  = 16 for the other species. These two basic numbers found in the genus, x  = 7 and x  = 8, point to chromosome evolution by dysploidy, which has also been accompanied by polyploidy.  © 2006 The Linnean Society of London, Botanical Journal of the Linnean Society , 2006, 150 , 429–439.     

Environmental variation for outcrossing rate in rapeseed (Brassica napus)

Summary Rapeseed (Brassica napus) is a predominantly selfpollinated crop with about one-third outcrossing. The outcrossing rate may be influenced by environmental factors, and hence changes in the heterozygosity level of a variety may occur during multiplication. In an investigation on environmental variation in outcrossing, we estimated the outcrossing rate in the Swedish spring rapeseed cv Topas by isozyme analysis and found that outcrossing varied from 12% to 47% over five locations in Sweden, Denmark and Germany. Among flowers at different positions on the same plant, average outcrossing varied from 11% at the top to 39% at the bottom of the plant. In conclusion, environmental factors can greatly influence the outcrossing rate in rapeseed, and an investigation therefore merit further studies.     

Intraspecific hybridization and the recovery of fitness in the native legume Chamaecrista fasciculata

Genetic incompatibilities and low offspring fitness are characteristic outcomes of hybridization between species. Yet, the creative potential of recombination following hybridization continues to be debated. Here we quantify the outcome of hybridization and recombination between adaptively divergent populations of the North American legume Chamaecrista fasciculata in a large-scale field experiment. Previously, hybrids between these populations demonstrated hybrid breakdown, suggesting the expression of adaptive epistatic interactions underlying population genetic differentiation. However, the outcome of hybridization ultimately rests on the performance of even later generation recombinants. In experiments that compared the performance of recombinant F6 and F2 generations with nonrecombinant F1 and parental genotypes, we observed that increasing recombination had contrasting effects on different life-history components. Lifetime fitness, defined as the product of survivorship and reproduction, showed a strong recovery of fitness in the F6. The overall gain in fitness with increased recombination suggests that hybridization and recombination may provide the necessary genetic variation for adaptive evolution within species. We discuss the mechanisms that may account for the gain in fitness with recombination, and explore the implications for hybrid speciation and phenotypic evolution.     

Taxonomic and functional implications of stigma morphology in species ofCassia,Chamaecrista, andSenna (Leguminosae:Caesalpinioideae)

Two stigma forms occur inChamaecrista andSenna, but only one inCassia. In the common chambered form, a stigma pore is positioned on the reflexed style tip and is the entrance to a tapering chamber. The pore rim is fringed by hairs which vary in number, size, distribution and shape. In the alternative form the stigma is situated at the apex of the curved style and is crateriform. The crater rim is fringed by hairs of variable number and shape. The stigmatic hairs are predominantly unicellular and cutinized. Stigma and hair differences aid in the taxonomy of the genera. Their functions in pollination biology are discussed.     

Anther and stigma morphology in mirror-image flowers of Chamaecrista chamaecristoides (Fabaceae): implications for buzz pollination

Enantiostyly (mirror-image flowers) is usually associated with buzz pollination. In buzz-pollinated flowers, pollen is released through terminal pores after bees vibrate the stamens. Several studies have evaluated the function of 'buzzing' in pollen release, but less attention has been paid to the effect of buzzing on pollen capture and deposition on stigmas. Evaluating the mechanism of pollen dispersion in buzz-pollinated flowers is important because it may affect mating patterns and reproductive success. In this study, we analysed the morphology of sexual organs (anther and stigma) using electron microscopy, and determined the relationship between sexual organ structure and pollen capture function through experimental manipulations of buzz-pollinated flowers of Chamaecrista chamaecristoides, as well as vibration frequencies on floral visitors. Pollen release occurs through two terminal pores at the tip of the stamens. However, unlike most angiosperms that have their stigmatic surface exposed, C.?chamaecristoides presents a stigmatic surface inside a cavity covered by trichomes. Experimental manipulations showed that effective fertilisation is only achieved when the style is vibrated, suggesting that buzzing is not only important for pollen release but also for pollen capture and deposition on the stigma. This result, in addition to vibration frequency analysis, suggests that although all floral visitors buzz flowers only those that buzz at higher frequencies achieve effective fertilisation. The anatomical features of sexual organs in flowers of C.?chamaecristoides demonstrate that this species possesses a highly specialised, elaborate morphology, with both genders selected for traits that promote buzz pollination.     

The impact of habitat fragmentation on fitness‐related traits in a native prairie plant,Chamaecrista fasciculata (Fabaceae)

Loss and fragmentation of the native prairies in the Midwestern United States have resulted in isolated and smaller habitats and populations. The populations remaining in these prairies are expected to show a decline in the extent of genetic variation and an increase in genetic drift load (accumulation of deleterious recessive alleles due to genetic drift) in fitness‐related traits. Using complementary greenhouse experiments, we tested whether these expected changes have occurred in the native annual prairie plant Chamaecrista fasciculata. In the first experiment, open pollinated C. fasciculata seeds from 12 prairie fragments representing a range in area of habitat were grown in competition with Schizachyrium scoparium to determine if there are changes in plant vigour with changes in fragment size and corresponding changes in population size. Plants from smaller prairie fragments exhibited a slight but significant decline in biomass, suggesting an increase in genetic drift load. In the second experiment, a formal genetic crossing design of four prairie fragment populations was used to estimate quantitative genetic diversity and genetic drift load. We did not find extensive quantitative genetic variation, but we did find a strong effect of genetic drift load on five traits in this experiment. Our overall conclusion is that a decline in relative‐fitness traits in smaller prairie fragments is probably associated with fixation of deleterious alleles due to more isolated and smaller populations, i.e. genetic drift load. © 2012 The Linnean Society of London, Biological Journal of the Linnean Society, 2012, ?? , ??–??.     

Variation in outcrossing rate and population genetic structure of Clarkia tembloriensis (Onagraceae)

Summary Outcrossing rate estimates for eight accessions of Clarkia tembloriensis indicate that this annual plant species has a wide interpopulational range of outcrossing rate ( ). Populations' t estimates were significantly correlated with observed heterozygosity and mean number of alleles per locus. Estimated fixation indices, , for most populations were very close to their expected values, F_eq, for a given Nei's gene diversity statistics showed that the group of outcrossing populations have more total genetic variation and less differentiation among populations than does the group of selfing populations. These results indicate that the breeding system of C. tembloriensis has had a strong influence on the amount and distribution of genetic variation within and among its populations.     

Cloning and characterization of a caesalpinoid (Chamaecrista fasciculata) hemoglobin: the structural transition from a nonsymbiotic hemoglobin to a leghemoglobin

Nonsymbiotic hemoglobins (nsHbs) and leghemoglobins (Lbs) are plant proteins that can reversibly bind O(2) and other ligands. The nsHbs are hexacoordinate and appear to modulate cellular concentrations of NO and maintain energy levels under hypoxic conditions. The Lbs are pentacoordinate and facilitate the diffusion of O(2) to symbiotic bacteroids within legume root nodules. Multiple lines of evidence suggest that all plant Hbs evolved from a common ancestor and that Lbs originated from nsHbs. However, little is known about the structural intermediates that occurred during the evolution of pentacoordinate Lbs from hexacoordinate nsHbs. We have cloned and characterized a Hb (ppHb) from the root nodules of the ancient caesalpinoid legume Chamaecrista fasciculata. Protein sequence, modeling data, and spectral analysis indicated that the properties of ppHb are intermediate between that of nsHb and Lb, suggesting that ppHb resembles a putative ancestral Lb. Predicted structural changes that appear to have occurred during the nsHb to Lb transition were a compaction of the CD-loop and decreased mobility of the distal His inhibiting its ability to coordinate directly with the heme-Fe, leading to a pentacoordinate protein. Other predicted changes include shortening of the N- and C-termini, compaction of the protein into a globular structure, disappearance of positive charges outside the heme pocket and appearance of negative charges in an area located between the N- and C-termini. A major consequence for some of these changes appears to be the decrease in O(2)-affinity of ancestral nsHb, which resulted in the origin of the symbiotic function of Lbs.     

Molecular separation of genera in Cassiinae (Leguminosae), and analysis of variation in the nodulating species of Chamaecrista

Randomly Amplified Polymorphic DNA (RAPD) methods have been adapted for use as a phenetic tool on the legume tribe Cassiinae. RAPD-generated polymorphism within local populations was lower than between populations from different geographic regions, between species and genera. Examination of three Cassia species, 12 Chamaecrista species and 13 Senna species using eight primers showed the potential for separation of the nodulated/nitrogen fixing genus Chamaecrista from the previously congeneric groups Cassia and Semis. Similarly, RAPD analysis of two groups of nine Ch. rotundifolia and nine Ch. mimosoides samples using 11 primers has given separation according to both species and to geographical location. Analysis of a small sample of five Chamaecrista species from Brazil with eight primers gave separation consistent with known variations in nodule structure.     

Natural selection on extrafloral nectar production in Chamaecrista fasciculata: the costs and benefits of a mutualism trait

Cost-benefit models of the evolution of mutualism predict that the current state of mutualism results from trade-offs between fitness costs of mutualist traits and the fitness benefits of association. We test the assumptions of such models by measuring patterns of natural selection on a mutualist trait, extrafloral nectar production in Chamaecrista fasciculata. Selection was measured on plants from which ants had been excluded (removing the mutualist benefit of the trait), from which all insects had been excluded (removing costs of herbivory in addition to mutualist benefits), and unmanipulated plants (where both costs and benefits were present). Selection analysis based on half-sibling-mean regressions of fitness on the trait revealed no evidence of costs of extrafloral nectar production in the absence of all insects or in the absence of ants. However, examination of the selective surfaces for these treatments suggest that costs of nectar production may exist and are exacerbated by the presence of herbivory. In the presence of ants, natural selection favors high extrafloral nectar production, consistent with a fitness benefit to this mutualist trait in the presence of the mutualist partner. In this study, the interaction of costs and benefits did not produce an evolutionary optimum for the trait within the range of variation observed, suggesting that application of a cost-benefit framework to this trait will benefit from considering the influence of temporal and spatial variation on the quality of costs and benefits.     

Honeybees enhance reproduction without affecting the outcrossing rate in endemic Pedicularis densispica (Orobanchaceae)

There has been substantial debate in recent years surrounding the impact of introduced honeybees on native biota. This study reports on an investigation of Pedicularis densispica, a subalpine annual herb endemic to Southwest China, in an attempt to determine the impact of introduced domestic honeybees on pollen dispersal and thus on their reproductive success and mating system. Honeybees were introduced into the study site in 2004, and a sudden seasonal pollinator shift from bumblebees to honeybees was observed. Intra- and inter-plant visits by different pollinators were recorded in the field in 2003 and 2004. Fruit and seed sets prior to and after the pollinator shift were measured. Experimental pollinations were performed to characterize the breeding system. Outcrossing rates at the seed stage were estimated for both years using RAPD markers. Our results indicated that honeybees foraged between plants more frequently than bumblebees did. Our results also revealed that the introduction of honeybees significantly enhanced reproductive success. However, no significant difference was detected between the outcrossing rates due to bumblebee and honeybee pollination. P. densispica was almost completely outcrossing ( T(m) = 0.956 and 0.967, respectively in 2003 and 2004) but partially self-compatible. This study presents the first report of the outcrossing rate in the genus pedicularis and reveals a limited influence of pollination on the mating system in P. densispica. The pollinator shift did not reduce reproductive success of the plants and honeybees may be used to augment pollinator services for nectariferous P. densispica.     

Unexpectedly high outcrossing rate in both dense and sparse patches in self-compatible Pedicularis rex (Orobanchaceae)

Pedicularis rex, a self-compatible alpine herb, depends exclusively on pollinators for reproduction. Previous study has revealed that 74.7 % of genetic variability occurs among populations in Yunnan, indicating a mixed mating system for the species. The large floral display also indicates high potential geitonogamy. Therefore, plant density and display size may be important factors influencing the realized mating system of P. rex. In the study reported here we explored the effects of plant density and floral display size on pollinator movements and thus outcrossing rate in two patches of in P. rex at low and high plant densities. We examined foraging behavior of bumblebees, measured reproductive outputs under hand and natural pollination, and estimated the outcrossing rates using RAPD markers. Floral display size was significantly larger and bumblebees visited significantly more flowers in sequence on individual plants in the sparse patch than in the dense patch. We found a significant positive correlation between floral display size and successive visits in the sparse patch, but not in the dense patch. Regression analysis revealed a significant inverse correlation between fruit production and flower production per plant with a theoretical maximum of 63 fruits per plant, but no relationship between seed production and flower production per plant. Contrary to previous studies on genetic structure, however, we found an unexpectedly high outcrossing rate in both the sparse and the dense patch (t _m = 1.151 and 0.924, respectively). Resource limitation may intensify seed competition and result in selected seed abortion, which may largely explain the unexpectedly high outcrossing rate.     

Novelties in Chamaecrista section Absus (Caesalpiniaceae)

Described, figured and discussed are two new species ofChamaecrista sect.Absus from campo rupestre habitats in southeastern Brazil:Ch. axiliflora Irwin & Barneby, of seriesAdenophyllae, from central Bahia, related toCh. adenophylla (Taubert) Irwin & Barneby; andCh. gumminans Irwin & Barneby, of seriesGlutinosae, from central Minas Gerais, related toCh. stilifera Irwin & Barneby.