Floral scent and its correlation with AFLP data in Sorbus

Comparisons between floral scent-based and DNA-molecular-based taxonomies are rare, yet such comparisons indicate that scent can provide useful taxonomic information. Here, we correlate the phytochemical differentiation in floral scent to the DNA-molecular-based differentiation in the genus Sorbus. Inflorescence scent patterns of the apomictic and endemic Sorbus latifolia microspecies Sorbus franconica, Sorbus adeana, and Sorbus cordigastensis originated by hybridization as well as their parental taxa Sorbus aria agg. and Sorbus torminalis were investigated with the dynamic headspace method. The scent data (presence/absence of compounds) were used to construct an UPGMA tree, to calculate a similarity matrix, and to correlate them with the published amplified fragment length polymorphism (AFLP) data of the same individuals, populations, and taxa. Flow cytometry was used to estimate the DNA-ploidy level of the taxa. Scent analyses showed a total of 68 substances, among them aromatic compounds, terpenoids, aliphatics, and nitrogen-containing compounds. The scent patterns were taxon-specific, and the number of scent components differed among taxa. The correlations with the published AFLP data on population and individual level are highly significant, indicating that the scent and AFLP data are highly congruent in the plants studied. Scent therefore provides useful taxonomic characters in Sorbus.     

Hybridization and polyploidy as drivers of continuing evolution and speciation in Sorbus

Interspecific hybridization and polyploidy are pivotal processes in plant evolution and speciation. The fate of new hybrid and polyploid taxa is determined by their ability to reproduce either sexually or asexually. Hybrids and allopolyploids with odd chromosome numbers are frequently sterile but some establish themselves through asexual reproduction (vegetative or apomixis). This allows novel genotypes to become established by isolating them from gene flow and leads to complex patterns of variation. The genus Sorbus is a good example of taxonomic complexity arising from the combined effects of hybridization, polyploidy and apomixis. The Avon Gorge in South‐west Britain contains the greatest diversity of Sorbus in Europe, with three endemic species and four putative endemic novel hybrids among its 15 native Sorbus taxa. We used a combination of nuclear microsatellite and chloroplast DNA markers to investigate the evolutionary relationships among these Sorbus taxa within the Avon Gorge. We confirm the genetic identity of putative novel taxa and show that hybridization involving sexual diploid species, primarily S. aria and S. torminalis and polyploid facultative apomictic species from subgenus Aria, has been responsible for generating this biodiversity. Importantly our data show that this creative evolutionary process is ongoing within the Avon Gorge. Conservation strategies for the rare endemic Sorbus taxa should therefore consider all Sorbus taxa within the Gorge and must strive to preserve this evolutionary process rather than simply the individual rare taxa that it produces.     

When sexual meets apomict: genome size,ploidy level and reproductive mode variation of Sorbus aria s.l. and S. austriaca (Rosaceae) in Bosnia and Herzegovina

Background and Aims Allopolyploidy and intraspecific heteroploid crosses are associated, in certain groups, with changes in the mating system. The genus Sorbus represents an appropriate model to study the relationships between ploidy and reproductive mode variations. Diploid S. aria and tetraploid apomictic S. austriaca were screened for ploidy and mating system variations within pure and sympatric populations in order to gain insights into their putative causalities.Methods Flow cytometry was used to assess genome size and ploidy level among 380 S. aria s.l. and S. austriaca individuals from Bosnia and Herzegovina, with 303 single-seed flow cytometric seed screenings being performed to identify their mating system. Pollen viability and seed set were also determined.Key Results Flow cytometry confirmed the presence of di-, tri- and tetraploid cytotype mixtures in mixed-ploidy populations of S. aria and S. austriaca. No ploidy variation was detected in single-species populations. Diploid S. aria mother plants always produced sexually originated seeds, whereas tetraploid S. austriaca as well as triploid S. aria were obligate apomicts. Tetraploid S. aria preserved sexuality in a low portion of plants. A tendency towards a balanced 2m : 1p parental genome contribution to the endosperm was shared by diploids and tetraploids, regardless of their sexual or asexual origin. In contrast, most triploids apparently tolerated endosperm imbalance.Conclusions Coexistence of apomictic tetraploids and sexual diploids drives the production of novel polyploid cytotypes with predominantly apomictic reproductive modes. The data suggest that processes governing cytotype diversity and mating system variation in Sorbus from Bosnia and Herzegovina are probably parallel to those in other diversity hotspots of this genus. The results represent a solid contribution to knowledge of the reproduction of Sorbus and will inform future investigations of the molecular and genetic mechanisms involved in triggering and regulating cytotype diversity and alteration of reproductive modes.     

Towards a better understanding of polyploid Sorbus (Rosaceae) from Bosnia and Herzegovina (Balkan Peninsula), including description of a novel,tetraploid apomictic species

Sorbus subgenus Soraria encompasses taxa originating from spontaneous hybridization between members of subgenera Aria and Sorbus disjunctly distributed across Europe and Asia. Using molecular data (amplified fragment length polymorphisms, plastid DNA sequences and nuclear microsatellites), flow cytometry (allowing for the determination of ploidy and mode of reproduction) and morphology, we disentangled the relationships among polyploid cytotypes and explored their relationships with their diploid ancestors. Among others, we focused on a large, geographically isolated hybrid population in Bosnia and Herzegovina. Molecular and morphological analyses confirmed the distinct position of this population in relation to its parental (S. aria and S. aucuparia) and other hybridogenous taxa originating independently from the same parents in different parts of Europe. After establishing its genetic and morphological divergence, we describe the isolated Bosnian population as a new tetraploid apomictic species, S orbus bosniaca , discuss its taxonomic status and propose conservation measures to protect the locus classicus of this new Balkan endemic. © 2015 The Linnean Society of London, Botanical Journal of the Linnean Society, 2015, 178 , 670–685.     

The origin of intermediate species of the genus Sorbus

The genus Sorbus in Europe contains five diploid species, Sorbus aria, Sorbus aucuparia, Sorbus torminalis, Sorbus chamaemespilus and Sorbus domestica, classified into five different subgenera. The subgenus Aria (or the S. aria aggregate) contains apomictic triploid and tetraploid species. Within the genus there are, in addition, a number of species, morphologically intermediate between plants of the four main subgenera, which are considered to reproduce partly, or exclusively, by apomixis. These are believed to have originated by hybridisation between species in the S. aria aggregate and a species from another subgenus (either S. aucuparia, S. torminalis or S. chamaemespilus). We have used restriction fragment length polymorphism (RFLP) analysis on a total of 178 Sorbus accessions to test this model. The genome relationships of the different groups have been assessed, and the hybrid nature of the major intermediate groups is unequivocally demonstrated. Polyploid species in the S. aria aggregate show genetic variation, indicating the possibility of multiple origins and/or facultative apomictic breeding behaviour. A major finding, confirmed by microsatellite analysis, is that the 'intermediate' species S. intermedia is shown to have genomes from S aria, S aucuparia and S torminalis. Polymorphic mitochondrial DNA markers were used to determine the direction of the crosses that gave rise to new 'hybrid species'; in the majority of cases the pollen was provided by the parent from the S aria aggregate.     

Growth, regeneration and shade tolerance of the Wild Service Tree (Sorbus torminalis (L.) Crantz) in aged oak coppice forests

Sorbus torminalis L. (Crantz) is a rare species in Central European forests with very limited quantitative information on its regeneration and growth dynamics. Since coppicing is no longer practiced in the most parts of Central Europe, it is unclear whether S. torminalis, which has usually a shorter end height than companion species, can persist in high forest systems. Here, we quantified species frequency on three 1 ha sample plots of former oak coppice forest. To determine whether S. torminalis regenerated continuously and how it might compete with oaks, the age of 80 trees was determined, and diameter and height growth were reconstructed for the 20 largest trees by stem analysis. To assess its shade tolerance, photosynthesis was measured for leaves located in high and low light conditions. Dendrochronological data demonstrated that, over the last 80 years, continuous recruitment of S. torminalis occurred. Growth patterns and photosynthesis measurements suggest that S. torminalis is a highly shade-tolerant species. We conclude that abandonment of coppicing in these forests does not threaten the status of S. torminalis, which can persist beneath the canopy of oaks.     

Origins and distribution of invasive Rubus fruticosus L. agg. (Rosaceae) clones in the Western United States

In facultatively asexual invasive species, an understanding of the origins and diversity of clones can reveal introduction and invasion pathways and inform management efforts. In this study, we use microsatellite and chloroplast DNA markers to infer clonal diversity of the Rubus fruticosus agg. invasion in the Western United States, determine the relationship of these clones to clones found in other exotic ranges, and determine the geographic and genetic origins of the invasive clones. We found two invasive clones in the Western United States, where the invasion had previously been thought to consist of a single asexual lineage. The most common clone was genetically identical to the microspecies R. armeniacus from the native range of Germany, while the second clone was identical to the microspecies R. anglocandicans in the invaded range of Australia and closely related to samples from the native ranges of England and Serbia. A third distinct clone was identified in a collection from the exotic range of Chile. Our results demonstrate that cryptic genetic diversity may be present in asexual invasions that are thought to be homogeneous. However, the asexual relationships between R. fruticosus agg. clones in the native and multiple exotic ranges indicate that preadaptation has played an important role in invasion success in this species aggregate.     

Reproductive strategy and ploidy determine the genetic variability of Sorbus aria

Sorbus aria (L.) Crantz (common whitebeam) from the Canary Islands has not been characterised genetically. We analysed the genetic variability of 184 individuals belonging to seven natural populations of S. aria from the Canarian Archipelago and the Iberian Peninsula. Our main aims were to obtain essential information to enable the exploration of the genetic relationship between populations from the Canary Islands and the Iberian Peninsula; to establish the existence of a spatial genetic structure and formulate appropriate management and conservation genetics strategies. Genetic variation was analysed using nine polymorphic microsatellite loci. The Canary Island populations (triploids) were found to have very low genetic variability and to be considerably differentiated from the populations from the peninsula (diploid and triploid), although with a connection to the Sierra Nevada population in the south of the Peninsula. This population, in turn, had many different genotypes, which is indicative of the existence of various origins. The level of genetic diversity was higher in all-diploid populations, which, in addition, presented a greater interpopulation gene flow, possibly the result of a prevalence of sexual reproduction. On the other hand, the triploid populations presented lower levels of genetic variability, with a significant degree of fixed heterozygosity, possibly due to asexual reproduction, mainly by apomixis. The reproductive biology and ploidy appear to be responsible for the levels of genetic variability in S. aria.     

Conservation of Britain's biodiversity: the case of Hieracium cyathis (Asteraceae) and its relation to other apomictic taxa

Hieracium cyathis (Ley) W. R. Linton (Asteraceae), Chalice Hawkweed, is a rare species endemic to Britain. It is a distinct member of Hieracium Section Oreadea , characterized by a short acladium and out-curved tips to the phyllaries. Historic records and current field survey indicate it has occurred in two localities in SW England and seven in South Wales. It is still present in all sites except for one Welsh locality, is significantly threatened at Cheddar Gorge, and two Welsh sites have only three plants each. Comparative morphology, cultivation experiments and amplified fragment length polymorphism (AFLP) analysis do not support the reported differences between material from SW England and Wales. AFLP analysis showed that every plant in the field was genetically similar but different, and minor genetic variation was found in one progeny array. It is a triploid (2 n  = 27) with apomictic seed production, and 26% potentially viable pollen. It qualifies as 'Near Threatened' under the 2001 IUCN threat criteria. General guidelines for conservation of microspecies of Hieracium are discussed.  © 2008 The Linnean Society of London, Botanical Journal of the Linnean Society , 2008, 156 , 669–680.     

Low genetic diversity in small peripheral populations of a rare European tree <Emphasis Type="Italic">(Sorbus torminalis)</Emphasis> dominated by clonal reproduction

Analysis of nuclear microsatellites in six small populations of Sorbus torminalis on its northern distribution limit was carried out to evaluate the dominating mode of reproduction as well as genetic diversity within and among populations. Within populations there was a low genetic diversity with heterozygote excess in five populations. Genetic differentiation was high among populations and the amount of clonally arisen recruits was 94–100%. This suggests that clonal growth, small population size and a possible self-incompability system maintain heterozygosity, although some genetic drift would also be expected. The results are discussed with respect to potential conservation strategies for S. torminalis in the study region.     

Changes in stomatal characteristics and photochemical efficiency during leaf development in six species of Sorbus

Measurements of Sorbus stomata size and density, maximal photochemical efficiency of photosystem II (F_v/F_m), variable-to-initial fluorescence ratio (F_v/F₀) and potential electron acceptor capacity (‘area’) were performed during leaf development in four parental diploid species, S. aria, S. aucuparia, S. chamaemespilus, S. torminalis, and two hybrid species, S. hazslinszkyana and S. intermedia. In fully expanded mature leaves, stomata lengths and densities were significantly larger in the shrub S. chamaemespilus than in the five tree species. The best performance of both the F_v/F_m and the F_v/F₀ ratio was recorded in S. intermedia, whereas S. chamaemespilus had the highest value of ‘area’. From a physiological point of view, the results of this study showed that the photosystem II reaction centers remained intact functionally through all phenological stages of leaf expansion for all examined species of Sorbus.     

Evolution of the Portulaca oleracea L. aggregate in Egypt on molecular and phenotypic levels revealed by morphology,inter-simple sequence repeat (ISSR) and 18S rDNA gene sequence markers

Molecular markers including inter simple sequence repeats (ISSR) and 18S rDNA gene sequence markers were combined with a detailed morphological analysis to seek characters that discriminate four taxa of Portulaca oleracea s.l. These taxa were identified as the microspecies Portulaca nitida, Portulaca oleracea, Portulaca rausii and Portulaca granulatostellulata. Morphological characters did not provide a clear distinction among the four taxa of P. oleracea s.l. It was found that mixed populations of the taxa occur in several locations and morphological similarities between the microspecies were detected. ISSR analysis indicates that gene flow among populations of different taxa was high and most of the genetic variation (61.9%) occurs within population. These results are inconsistent with the general characteristics of P. oleracea as a self- or 5% cross-pollinated species. A close relationship between P. oleracea, P. rausii and P. granulatostellulata was supported by ISSR and 18S rDNA gene sequence. The ISSR and 18s data support the specific status of P. nitida as a recently evolved taxon. We conclude that P. oleracea exists as a polymorphic species and is not divisible into microspecies based on seed surface as the primary morphological trait, and inter simple sequence repeats (ISSR) and 18S rDNA gene sequence markers. We suggest that the taxa do not merit specific rank as morphological characters and absence of a breeding barrier fail to separate the different populations when they become sympatric.     

Genetic polymorphism in Chondrilla (Asteraceae) in southern European Russia and the nature of Chondrilla juncea L

We studied genetic diversity in 54 populations of nine sexual and apomictic species of the genus Chondrilla (C. acantholepis, C. ambigua, C. brevirostris, C. canescens, C. graminea, C. juncea, C. laticoronata, C. latifolia and C. pauciflora) in SE European Russia and neighboring territories of NW Kazakhstan. We analysed the trnT–trnF region of plastid DNA and the internal transcribed spacer of ribosomal DNA (ITS1–5.8S–ITS2) using statistical parsimony, maximum likelihood and neighbor net methods. Two major evolutionary lineages, roughly corresponding to the two subgenera traditionally recognized in the region, were revealed. Within the first evolutionary lineage (subgenus Brachyrhynchus), the sexual diploid C. ambigua and its putatively hybrid apomictic derivatives C. brevirostris, C. laticoronata and C. pauciflora could be recognized. Their identity was also confirmed by analyses of ISSR markers. The second evolutionary lineage (subgenus Chondrilla) comprises C. juncea, C. acantholepis, C. canescens, C. graminea and C. latifolia in European Russia, but analyses of morphological variability and the genealogy of plastid and nuclear markers favor their treatment as the single facultatively apomictic species C. juncea. The results demonstrate that an apomictic mode of reproduction does not necessarily result in the formation of genetically separated microspecies.     

AFLP analyses demonstrate genetic divergence, hybridization, and multiple polyploidization in the evolution of Achillea (Asteraceae-Anthemideae)

Achillea, a temperate genus of herbaceous allogamous perennials, is a model for evolutionary radiation through hybridization and polyploidization. AFLP analyses were performed on 300 individuals of 66 populations and 27 taxa/cytotypes, mainly from the polyploid A. millefolium aggregate and its suspected hybrid links with other clades of the genus. The mosaic genetic structure of hybrids and polyploids is revealed by specific AFLP bands shared with their assumed parents. In E Asia, A. alpina-4x and A. wilsoniana-4x are allotetraploids between A. acuminata-2x (sect. Ptarmica) and A. asiatica-2x (sect. Achillea-A. millefolium agg.). A. virescens-4x is a hybrid species linking A. nobilis agg. and A. millefolium agg. in S Europe. The hybrid swarm A. clypeolata-2x yen A. collina-4x recently formed in Bulgaria shows no AFLP bands additive to its parents; by contrast, other more ancient allopolyploids exhibit genetic innovations. Relationships within A. millefolium agg. are complex. Five 2x-taxa, mostly well separated and regressive, are limited to Eurasia; seven 4x- and 6x-taxa are intimately linked by hybridization, are expansive, and through A. asiatica-2x/4x have formed the N American polyploids. All these results from AFLPs correspond well to other evidence, and indicate a long history of reticulate evolution in Achillea.     

Genetic relationship and genetic diversity among Typha taxa from East Asia based on AFLP markers

The genetic relationship and diversity among four Typha taxa in East Asia were evaluated using amplified fragment length polymorphism (AFLP) markers. Three AFLP selective primer combinations generated a total of 707 amplification products, of which 704 (99.6%) were polymorphic. The unweighted pair-group method with arithmetic mean (UPGMA) dendrogram and principal component analysis (PCA) plot confirmed the taxonomic status of four separate species. East Asian Typha taxa separated into two groups: the first with Typha angustifolia and the second with T. orientalis, T. laxmanni, and T. latifolia with a high bootstrap value for UPGMA (93%) and a low first score for PCA (25%). The two clusters corresponded with two sections based on the bracteoles in the female flower: section Bracteolatae and section Ebracteolatae. T. angustifolia showed the highest genetic diversity among the four Typha taxa (percentage of polymorphic loci [PPL] = 71%, H_o = 0.157), whereas T. latifolia had the lowest genetic diversity (PPL = 40%, H_o = 0.117). Genetic diversity was related to the presence of the gap between male and female inflorescences. A positive correlation between genetic distance and geographic distance was clearly found in the two species with continuous inflorescences (T. latifolia and T. orientalis). This positive correlation was not observed in the other species with discontinuous spikes (T. angustifolia and T. laxmanni).     

The Detection,Rate and Manifestation of Residual Sexuality in Apomictic Populations of Pilosella (Asteraceae,Lactuceae)

The effect of maternal, facultatively apomictic plants on population diversity was evaluated in seven hybridizing polyploid Pilosella populations, where apomictic (P. bauhini or P. aurantiaca) and sexual (P. officinarum) biotypes coexist. The ploidy level, reproductive system, morphology, clonal structure and chloroplast DNA haplotypes were used to characterize these plants and their hybrids. The reproductive origins of the progeny were assessed through either a flow cytometric seed screen and/or a comparison between the ploidy level of progeny embryos/seedlings and the maternal ploidy level. The cultivated progeny derived from residual sexuality in maternal apomicts were also identified based on their morphology and reproductive behaviour. The progeny different from their maternal parents (0.6?92.3 % of progeny embryos and 0?100 % of progeny seedlings) originated either sexually or via haploid parthenogenesis. Comparing the facultatively apomictic and sexual mothers, the progeny arrays generated in the field showed that apomictic mothers produce progeny that is more variable in ploidy level. This effect was demonstrated at both the embryonic and seedling stages of progeny development. Residual sexuality in apomicts was also effective in experimental crosses, generating progeny similar to spontaneous hybrids in the field. The 2n + n hybrids produced from an apomictic and a sexual parent displayed similar reproductive behaviour, producing polyhaploid, sexual and apomictic progeny in variable ratios. Repeated hybridizations between parental species and/or multi-step crosses can result in hybrid swarms rich in cytotypes and morphotypes. The variation recorded in these populations suggests prevailing introgressive hybridization towards the sexual species P. officinarum.     

Microsatellite primers for Sorbus torminalis and related species

This study reports the cloning and characterization of nine microsatellite primer pairs in a scattered woody species (Sorbus torminalis), and shows their potential for further use in 36 species of the Maloideae, a Rosaceae subfamily containing important fruit crop and ornamental species. These primers were designed from a microsatellite library constructed from genomic DNA of S. torminalis and enriched for CA and GA repeats. Genotyping 48 S. torminalis of a natural population with the six best markers yielded a mean of 10.7 alleles per locus, and an expectation of exclusion probability for paternity analysis greater than 0.993.     

The Limonium binervosum aggregate (Plumbaginaceae) in the British Isles

A taxometric analysis of 153 colonies of the agamospermous Limonium binervosum aggregate in the British Isles has revealed an ordered hierarchy of variation. A taxonomic treatment of the group describes five new species, redefines four others and describes other infraspecific variants as subspecies or varieties. Each species is morphologically distinct and has a defined geographic range. There are four widespread and morphologically variable species and five geographically restricted species (four monotypic). Limonium recurvum , previously described as a microspecies endemic to a small area of the Dorset Coast, has been widened to include all triploid (2 n = 27) L. binervosum agg. variants found also in W and N Ireland, SW Scotland and Cumbria. All other taxa are tetraploid, usually with 2 n = 35.     

Evaluation of AFLP markers to reveal genetic diversity in Typha

We investigated to what extent DNA-markers can assist species determination in the genus Typha. A set of AFLP markers was used to discriminate samples of the species Typha latifolia and Typha angustifolia collected in Flanders (North Belgium). The T. latifolia samples formed a compact cluster while the T. angustifolia samples were divided into smaller groups. It was not clear whether interspecific hybrids or higher levels of diversity present in the T. angustifolia dataset could account for this. As in previous surveys, using isozyme and VNTR markers, AFLP markers revealed an almost complete lack of genetic variation in Flemish T. latifolia. Despite the low degree of diversity, a significant level of genetic differentiation was found between the T. latifolia samples originating from different river basins. Whether this differentiation has any ecological relevance remains to be investigated. The methodology applied was not able to detect clonal reproduction in T. latifolia. Probably, the low levels of diversity present in this species can account for this, indicating that the usefulness of the methodology applied depends on the level of diversity present in the species studied.     

Variation in Two Sorbus Species Endemic to the Isle of Arran, Scotland

A study of the morphology of plants from four species of Sorbus,S. arraensis, S. pseudofennica, S. aucuparia and S. rupicola,two of them endemic to North Arran, suggested that the originof the two endemic groups could have been by hybridization. Variation between individuals within the hybrid groups indicatesthat the two groups, probably of hybrid origin (S. arranensisand S. pseudofennica), could each consist of a set of geneticallyisolated clones reproducing by apomixis, with the possibilityin the case of S. pseudofennica of occasional genetic interchangeby sexual reproduction. The overlap of characters between the two hybrid groups suggeststhat the inflow of genetic material by hybridization and geneticmodification in these groups is continuing. Sorbus arranensis Hedl., Sorbus pseudofennica E. F. Warb., Sorbus aucuparia L., Sorbus rupicola (Syme) Hedl., interspecific hybridization, apomixis