Breeding systems,hybridization and continuing evolution in Avon Gorge Sorbus

Background and Aims

Interspecific hybridization and polyploidy are key processes in plant evolution and are responsible for ongoing genetic diversification in the genus Sorbus (Rosaceae). The Avon Gorge, Bristol, UK, is a world ‘hotspot’ for Sorbus diversity and home to diploid sexual species and polyploid apomictic species. This research investigated how mating system variation, hybridization and polyploidy interact to generate this biological diversity.

Methods

Mating systems of diploid, triploid and tetraploid Sorbus taxa were analysed using pollen tube growth and seed set assays from controlled pollinations, and parent–offspring genotyping of progeny from open and manual pollinations.

Key Results

Diploid Sorbus are outcrossing and self-incompatible (SI). Triploid taxa are pseudogamous apomicts and genetically invariable, but because they also display self-incompatibility, apomictic seed set requires pollen from other Sorbus taxa – a phenomenon which offers direct opportunities for hybridization. In contrast tetraploid taxa are pseudogamous but self-compatible, so do not have the same obligate requirement for intertaxon pollination.

Conclusions

The mating inter-relationships among Avon Gorge Sorbus taxa are complex and are the driving force for hybridization and ongoing genetic diversification. In particular, the presence of self-incompatibility in triploid pseudogamous apomicts imposes a requirement for interspecific cross-pollination, thereby facilitating continuing diversification and evolution through rare sexual hybridization events. This is the first report of naturally occurring pseudogamous apomictic SI plant populations, and we suggest that interspecific pollination, in combination with a relaxed endosperm balance requirement, is the most likely route to the persistence of these populations. We propose that Avon Gorge Sorbus represents a model system for studying the establishment and persistence of SI apomicts in natural populations.     

A world-wide perspective on crucifer speciation and evolution: phylogenetics,biogeography and trait evolution in tribe Arabideae

Background and Aims

Tribe Arabideae are the most species-rich monophyletic lineage in Brassicaceae. More than 500 species are distributed in the majority of mountain and alpine regions worldwide. This study provides the first comprehensive phylogenetic analysis for the species assemblage and tests for association of trait and characters, providing the first explanations for the enormous species radiation since the mid Miocene.

Methods

Phylogenetic analyses of DNA sequence variation of nuclear encoded loci and plastid DNA are used to unravel a reliable phylogenetic tree. Trait and ancestral area reconstructions were performed and lineage-specific diversification rates were calculated to explain various radiations in the last 15 Myr in space and time.

Key Results

A well-resolved phylogenetic tree demonstrates the paraphyly of the genus Arabis and a new systematic concept is established. Initially, multiple radiations involved a split between lowland annuals and mountain/alpine perennial sister species. Subsequently, increased speciation rates occur in the perennial lineages. The centre of origin of tribe Arabideae is most likely the Irano-Turanian region from which the various clades colonized the temperate mountain and alpine regions of the world.

Conclusions

Mid Miocene early diversification started with increased speciation rates due to the emergence of various annual lineages. Subsequent radiations were mostly driven by diversification within perennial species during the Pliocene, but increased speciation rates also occurred during that epoch. Taxonomic concepts in Arabis are still in need of a major taxonomic revision to define monophyletic groups.     

Phylogeny of the cycads based on multiple single-copy nuclear genes: congruence of concatenated parsimony,likelihood and species tree inference methods

Background and aims

Despite a recent new classification, a stable phylogeny for the cycads has been elusive, particularly regarding resolution of Bowenia, Stangeria and Dioon. In this study, five single-copy nuclear genes (SCNGs) are applied to the phylogeny of the order Cycadales. The specific aim is to evaluate several gene tree–species tree reconciliation approaches for developing an accurate phylogeny of the order, to contrast them with concatenated parsimony analysis and to resolve the erstwhile problematic phylogenetic position of these three genera.

Methods

DNA sequences of five SCNGs were obtained for 20 cycad species representing all ten genera of Cycadales. These were analysed with parsimony, maximum likelihood (ML) and three Bayesian methods of gene tree–species tree reconciliation, using Cycas as the outgroup. A calibrated date estimation was developed with Bayesian methods, and biogeographic analysis was also conducted.

Key Results

Concatenated parsimony, ML and three species tree inference methods resolve exactly the same tree topology with high support at most nodes. Dioon and Bowenia are the first and second branches of Cycadales after Cycas, respectively, followed by an encephalartoid clade (Macrozamia–Lepidozamia–Encephalartos), which is sister to a zamioid clade, of which Ceratozamia is the first branch, and in which Stangeria is sister to Microcycas and Zamia.

Conclusions

A single, well-supported phylogenetic hypothesis of the generic relationships of the Cycadales is presented. However, massive extinction events inferred from the fossil record that eliminated broader ancestral distributions within Zamiaceae compromise accurate optimization of ancestral biogeographical areas for that hypothesis. While major lineages of Cycadales are ancient, crown ages of all modern genera are no older than 12 million years, supporting a recent hypothesis of mostly Miocene radiations. This phylogeny can contribute to an accurate infrafamilial classification of Zamiaceae.     

Phylogeographic patterns, genetic affinities and morphological differentiation between Epipactis helleborine and related lineages in a Mediterranean glacial refugium

Background and Aims

In the Mediterranean basin, the Italian peninsula has been suggested to be one of the most important glacial refugia for temperate tree species. The orchid genus Epipactis is widely represented in the Italian peninsula by widespread species and several endemic, localized taxa, including selfing and outcrossing taxa. Here the phylogenetic and phylogeographic relationships in a group of closely related taxa in Epipactis are investigated with the aim of understanding the role of this refugial area for cladogenesis and speciation in herbaceous species, such as terrestrial orchids.

Methods

Ribosomal DNA (rDNA) was employed to assess phylogenetic relationships, and plastid sequence variation in the rbcL–accD spacer was used to reveal phylogeographic patterns among plastid haplotypes using a parsimony network.

Key Results

Low genetic variation and shared ribotypes were detected in rDNA, whereas high levels of sequence variation and a strong phylogeographic structure were found in the examined plastid region. The parsimony plastid haplotype network identified two main haplotype groups, one including E. atrorubens/microphylla/muelleri/leptochila and the other including all accessions of E. helleborine and several localized and endemic taxa, with a combination of widespread and rare haplotypes detected across the Italian peninsula. A greater genetic divergence separated the Italian and other European accessions of E. helleborine.

Conclusions

Phylogenetic and phylogeographic patterns support a working hypothesis in which the Italian peninsula has only recently been colonized by Epipactis, probably during the most recent phase of the Quaternary age and, nevertheless, it acted as a remarkable centre of diversification for this orchid lineage. Changes in pollination strategy and recurrent shifts in mating system (from allogamy to autogamy) could have represented the mechanism promoting this rapid diversification and the observed high taxonomic complexity detected in the E. helleborine species complex.     

Light and competition gradients fail to explain the coexistence of shade-tolerant Fagus sylvatica and shade-intermediate Quercus petraea seedlings

Background and Aims

The coexistence of forest tree species has often been linked to differences among species in terms of their response to light availability during the regeneration stage. From this perspective, species coexistence results from growth–growth or mortality–growth trade-offs along spatial light gradients. Experimental evidence of growth–growth trade-offs in natural conditions is sparse due to various confounding factors that potentially hinder the relationship. This study examined growth hierarchies along light gradients between two tree species with contrasting shade tolerance by controlling potential confounding factors such as seedling size, seedling status, seedling density and species composition.

Methods

Natural regenerated shade-tolerant Fagus sylvatica and shade-intermediate Quercus petraea seedlings were used, and growth rankings over a 4-year period were compared in 8- to 10-year-old tree seedlings.

Key results

No rank reversal occurs between the two species along the light gradient, or along the density, mixture or seedling size gradients. The shade-tolerant species was always the more competitive of the two. Pronounced effects of initial size on seedling growth were observed, whereas the effects of light and competition by neighbours were of secondary importance. The paramount effect of size, which results from the asymmetric nature of interseedling competition, gives a strong advantage to tall seedlings over the long term.

Conclusions

This study extends previous efforts to identify potential drivers of rank reversals in young tree mixtures. It does not support the classical assumption that spatial heterogeneity in canopy opening explains the coexistence of the two species studied. It suggests that spatial variation in local size hierarchies among seedlings that may be caused by seedling emergence time or seedling initial performance is the main driver of the dynamics of these mixed stands.     

Extreme divergence in floral scent among woodland star species (Lithophragma spp.) pollinated by floral parasites

Backgrounds and Aims

A current challenge in coevolutionary biology is to understand how suites of traits vary as coevolving lineages diverge. Floral scent is often a complex, variable trait that attracts a suite of generalized pollinators, but may be highly specific in plants specialized on attracting coevolved pollinating floral parasites. In this study, floral scent variation was investigated in four species of woodland stars (Lithophragma spp.) that share the same major pollinator (the moth Greya politella, a floral parasite). Three specific hypotheses were tested: (1) sharing the same specific major pollinator favours conservation of floral scent among close relatives; (2) selection favours ‘private channels’ of rare compounds particularly aimed at the specialist pollinator; or (3) selection from rare, less-specialized co-pollinators mitigates the conservation of floral scent and occurrence of private channels.

Methods

Dynamic headspace sampling and solid-phase microextraction were applied to greenhouse-grown plants from a common garden as well as to field samples from natural populations in a series of experiments aiming to disentangle the genetic and environmental basis of floral scent variation.

Key Results

Striking floral scent divergence was discovered among species. Only one of 69 compounds was shared among all four species. Scent variation was largely genetically based, because it was consistent across field and greenhouse treatments, and was not affected by visits from the pollinating floral parasite.

Conclusions

The strong divergence in floral scents among Lithophragma species contrasts with the pattern of conserved floral scent composition found in other plant genera involved in mutualisms with pollinating floral parasites. Unlike some of these other obligate pollination mutualisms, Lithophragma plants in some populations are occasionally visited by generalist pollinators from other insect taxa. This additional complexity may contribute to the diversification in floral scent found among the Lithophragma species pollinated by Greya moths.     

Root–shoot allometry of tropical forest trees determined in a large-scale aeroponic system

Background and Aims

This study is a first step in a multi-stage project aimed at determining allometric relationships among the tropical tree organs, and carbon fluxes between the various tree parts and their environment. Information on canopy–root interrelationships is needed to improve understanding of above- and below-ground processes and for modelling of the regional and global carbon cycle. Allometric relationships between the sizes of different plant parts will be determined.

Methods

Two tropical forest species were used in this study: Ceiba pentandra (kapok), a fast-growing tree native to South and Central America and to Western Africa, and Khaya anthotheca (African mahogany), a slower-growing tree native to Central and Eastern Africa. Growth and allometric parameters of 12-month-old saplings grown in a large-scale aeroponic system and in 50-L soil containers were compared. The main advantage of growing plants in aeroponics is that their root systems are fully accessible throughout the plant life, and can be fully recovered for harvesting.

Key Results

The expected differences in shoot and root size between the fast-growing C. pentandra and the slower-growing K. anthotheca were evident in both growth systems. Roots were recovered from the aeroponically grown saplings only, and their distribution among various diameter classes followed the patterns expected from the literature. Stem, branch and leaf allometric parameters were similar for saplings of each species grown in the two systems.

Conclusions

The aeroponic tree growth system can be utilized for determining the basic allometric relationships between root and shoot components of these trees, and hence can be used to study carbon allocation and fluxes of whole above- and below-ground tree parts.     

Divergent selection in trailing- versus leading-edge populations of Biscutella laevigata

Background and Aims

Knowledge on how climate-induced range shifts might affect natural selection is crucial to understand the evolution of species ranges.

Methods

Using historical demographic perspectives gathered from regional-scale phylogeography on the alpine herb Biscutella laevigata, indirect inferences on gene flow and signature of selection based on AFLP genotyping were compared between local populations persisting at the trailing edge and expanding at the leading edge.

Key Results

Spatial autocorrelation revealed that gene flow was two times more restricted at the trailing edge and genome scans indicated divergent selection in this persisting population. In contrast, no pattern of selection emerged in the expanding population at the leading edge.

Conclusions

Historical effects may determine different architecture of genetic variation and selective patterns within local populations, what is arguably important to understand evolutionary processes acting across the species ranges.     

Paternity analysis-based inference of pollen dispersal patterns, male fecundity variation, and influence of flowering tree density and general flowering magnitude in two dipterocarp species

Background and Aims

Knowledge of pollen dispersal patterns and variation of fecundity is essential to understanding plant evolutionary processes and to formulating strategies to conserve forest genetic resources. Nevertheless, the pollen dispersal pattern of dipterocarp, main canopy tree species in palaeo-tropical forest remains unclear, and flowering intensity variation in the field suggests heterogeneity of fecundity.

Methods

Pollen dispersal patterns and male fecundity variation of Shorea leprosula and Shorea parvifolia ssp. parvifolia on Peninsular Malaysian were investigated during two general flowering seasons (2001 and 2002), using a neighbourhood model modified by including terms accounting for variation in male fecundity among individual trees to express heterogeneity in flowering.

Key Results

The pollen dispersal patterns of the two dipterocarp species were affected by differences in conspecific tree flowering density, and reductions in conspecific tree flowering density led to an increased selfing rate. Active pollen dispersal and a larger number of effective paternal parents were observed for both species in the season of greater magnitude of general flowering (2002).

Conclusions

The magnitude of general flowering, male fecundity variation, and distance between pollen donors and mother trees should be taken into account when attempting to predict the effects of management practices on the self-fertilization and genetic structure of key tree species in tropical forest, and also the sustainability of possible management strategies, especially selective logging regimes.     

Temporal changes in population genetic diversity and structure in red and white clover grown in three contrasting environments in northern Europe

Backgound and Aims

Extending the cultivation of forage legume species into regions where they are close to the margin of their natural distribution requires knowledge of population responses to environmental stresses. This study was conducted at three north European sites (Iceland, Sweden and the UK) using AFLP markers to analyse changes in genetic structure over time in two population types of red and white clover (Trifolium pratense and T. repens, respectively): (1) standard commercial varieties; (2) wide genetic base (WGB) composite populations constructed from many commercial varieties plus unselected material obtained from germplasm collections.

Methods

At each site populations were grown in field plots, then randomly sampled after 3–5 years to obtain survivor populations. AFLP markers were used to calculate genetic differentiation within and between original and survivor populations.

Key Results

No consistent changes in average genetic diversity were observed between original and survivor populations. In both species the original varieties were always genetically distinct from each other. Significant genetic shift was observed in the white clover ‘Ramona’ grown in Sweden. The WGB original populations were more genetically similar. However, genetic differentiation occurred between original and survivor WGB germplasm in both species, particularly in Sweden. Regression of climatic data with genetic differentiation showed that low autumn temperature was the best predictor. Within the set of cold sites the highest level of genetic shift in populations was observed in Sweden.

Conclusions

The results suggest that changes in population structure can occur within a short time span in forage legumes, resulting in the rapid formation of distinct survivor populations in environmentally challenging sites.     

New chloroplast microsatellite markers suitable for assessing genetic diversity of Lolium perenne and other related grass species

Background and Aims

Lolium perenne (perennial ryegrass) is the most important forage grass species of temperate regions. We have previously released the chloroplast genome sequence of L. perenne ‘Cashel’. Here nine chloroplast microsatellite markers are published, which were designed based on knowledge about genetically variable regions within the L. perenne chloroplast genome. These markers were successfully used for characterizing the genetic diversity in Lolium and different grass species.

Methods

Chloroplast genomes of 14 Poaceae taxa were screened for mononucleotide microsatellite repeat regions and primers designed for their amplification from nine loci. The potential of these markers to assess genetic diversity was evaluated on a set of 16 Irish and 15 European L. perenne ecotypes, nine L. perenne cultivars, other Lolium taxa and other grass species.

Key Results

All analysed Poaceae chloroplast genomes contained more than 200 mononucleotide repeats (chloroplast simple sequence repeats, cpSSRs) of at least 7 bp in length, concentrated mainly in the large single copy region of the genome. Nucleotide composition varied considerably among subfamilies (with Pooideae biased towards poly A repeats). The nine new markers distinguish L. perenne from all non-Lolium taxa. TeaCpSSR28 was able to distinguish between all Lolium species and Lolium multiflorum due to an elongation of an A₈ mononucleotide repeat in L. multiflorum. TeaCpSSR31 detected a considerable degree of microsatellite length variation and single nucleotide polymorphism. TeaCpSSR27 revealed variation within some L. perenne accessions due to a 44-bp indel and was hence readily detected by simple agarose gel electrophoresis. Smaller insertion/deletion events or single nucleotide polymorphisms detected by these new markers could be visualized by polyacrylamide gel electrophoresis or DNA sequencing, respectively.

Conclusions

The new markers are a valuable tool for plant breeding companies, seed testing agencies and the wider scientific community due to their ability to monitor genetic diversity within breeding pools, to trace maternal inheritance and to distinguish closely related species.     

Evolutionary history of the Afro-Madagascan Ixora species (Rubiaceae): species diversification and distribution of key morphological traits inferred from dated molecular phylogenetic trees

Background and Aims

Previous work on the pantropical genus Ixora has revealed an Afro-Madagascan clade, but as yet no study has focused in detail on the evolutionary history and morphological trends in this group. Here the evolutionary history of Afro-Madagascan Ixora spp. (a clade of approx. 80 taxa) is investigated and the phylogenetic trees compared with several key morphological traits in taxa occurring in Madagascar.

Methods

Phylogenetic relationships of Afro-Madagascan Ixora are assessed using sequence data from four plastid regions (petD, rps16, rpoB-trnC and trnL-trnF) and nuclear ribosomal external transcribed spacer (ETS) and internal transcribed spacer (ITS) regions. The phylogenetic distribution of key morphological characters is assessed. Bayesian inference (implemented in BEAST) is used to estimate the temporal origin of Ixora based on fossil evidence.

Key Results

Two separate lineages of Madagascan taxa are recovered, one of which is nested in a group of East African taxa. Divergence in Ixora is estimated to have commenced during the mid Miocene, with extensive cladogenesis occurring in the Afro-Madagascan clade during the Pliocene onwards.

Conclusions

Both lineages of Madagascan Ixora exhibit morphological innovations that are rare throughout the rest of the genus, including a trend towards pauciflorous inflorescences and a trend towards extreme corolla tube length, suggesting that the same ecological and selective pressures are acting upon taxa from both Madagascan lineages. Novel ecological opportunities resulting from climate-induced habitat fragmentation and corolla tube length diversification are likely to have facilitated species radiation on Madagascar.     

The evolutionary history of sea barley (Hordeum marinum) revealed by comparative physical mapping of repetitive DNA

Background and Aims Hordeum marinum

is a species complex that includes the diploid subspecies marinum and both diploid and tetraploid forms of gussoneanum. Their relationships, the rank of the taxa and the origin of the polyploid forms remain points of debate. The present work reports a comparative karyotype analysis of six H. marinum accessions representing all taxa and cytotypes.

Methods

Karyotypes were determined by analysing the chromosomal distribution of several tandemly repeated sequences, including the Triticeae cloned probes pTa71, pTa794, pAs1 and pSc119·2 and the simple sequence repeats (SSRs) (AG)₁₀, (AAC)₅, (AAG)₅, (ACT)₅ and (ATC)₅.

Key Results

The identification of each chromosome pair in all subspecies and cytotypes is reported for the first time. Homologous relationships are also established. Wide karyotypic differences were detected within marinum accessions. Specific chromosomal markers characterized and differentiated the genomes of marinum and diploid gussoneanum. Two subgenomes were detected in the tetraploids. One of these had the same chromosome complement as diploid gussoneanum; the second subgenome, although similar to the chromosome complement of diploid H. marinum sensu lato, appeared to have no counterpart in the marinum accessions analysed here.

Conclusions

The tetraploid forms of gussoneanum appear to have come about through a cross between a diploid gussoneanum progenitor and a second, related—but unidentified—diploid ancestor. The results reveal the genome structure of the different H. marinum taxa and demonstrate the allopolyploid origin of the tetraploid forms of gussoneanum.     

Cushions of Thylacospermum caespitosum (Caryophyllaceae) do not facilitate other plants under extreme altitude and dry conditions in the north-west Himalayas

Background

Cushion plants are commonly considered as keystone nurse species that ameliorate the harsh conditions they inhabit in alpine ecosystems, thus facilitating other species and increasing alpine plant biodiversity. A literature search resulted in 25 key studies showing overwhelming facilitative effects of different cushion plants and hypothesizing greater facilitation with increased environmental severity (i.e. higher altitude and/or lower rainfall). At the same time, emerging ecological theory alongside the cushion-specific literature suggests that facilitation might not always occur under extreme environmental conditions, and especially under high altitude and dryness.

Methods

To assess these hypotheses, possible nursing effects of Thylacospermum caespitosum (Caryophyllaceae) were examined at extremely high altitude (5900 m a.s.l.) and in dry conditions (precipitation <100 mm year⁻¹) in Eastern Ladakh, Trans-Himalaya. This is, by far, the highest site, and the second driest, at which the effects of cushions have been studied so far.

Key Results

In accordance with the theoretical predictions, no nursing effects of T. caespitosum on other alpine plants were detected. The number and abundance of species were greater outside cushions than within and on the edge of cushions. None of the 13 species detected was positively associated with cushions, while nine of them were negatively associated. Plant diversity increased with the size of the area sampled outside cushions, but no species–area relationship was found within cushions.

Conclusions

The results support the emerging theoretical prediction of restricted facilitative effects under extreme combinations of cold and dryness, integrating these ideas in the context of the ecology of cushion plants. This evidence suggests that cases of missing strong facilitation are likely to be found in other extreme alpine conditions.     

Haemophilus influenzae: using comparative genomics to accurately identify a highly recombinogenic human pathogen

Background

Haemophilus influenzae is an opportunistic bacterial pathogen that exclusively colonises humans and is associated with both acute and chronic disease. Despite its clinical significance, accurate identification of H. influenzae is a non-trivial endeavour. H. haemolyticus can be misidentified as H. influenzae from clinical specimens using selective culturing methods, reflecting both the shared environmental niche and phenotypic similarities of these species. On the molecular level, frequent genetic exchange amongst Haemophilus spp. has confounded accurate identification of H. influenzae, leading to both false-positive and false-negative results with existing speciation assays.

Results

Whole-genome single-nucleotide polymorphism data from 246 closely related global Haemophilus isolates, including 107 Australian isolate genomes generated in this study, were used to construct a whole-genome phylogeny. Based on this phylogeny, H. influenzae could be differentiated from closely related species. Next, a H. influenzae-specific locus, fucP, was identified, and a novel TaqMan real-time PCR assay targeting fucP was designed. PCR specificity screening across a panel of clinically relevant species, coupled with in silico analysis of all species within the order Pasteurellales, demonstrated that the fucP assay was 100 % specific for H. influenzae; all other examined species failed to amplify.

Conclusions

This study is the first of its kind to use large-scale comparative genomic analysis of Haemophilus spp. to accurately delineate H. influenzae and to identify a species-specific molecular signature for this species. The fucP assay outperforms existing H. influenzae targets, most of which were identified prior to the next-generation genomics era and thus lack validation across a large number of Haemophilus spp. We recommend use of the fucP assay in clinical and research laboratories for the most accurate detection and diagnosis of H. influenzae infection and colonisation.

Electronic supplementary material

The online version of this article (doi:10.1186/s12864-015-1857-x) contains supplementary material, which is available to authorized users.     

Extensive contemporary pollen-mediated gene flow in two herb species,Ranunculus bulbosus and Trifolium montanum,along an altitudinal gradient in a meadow landscape

Background and Aims

Genetic connectivity between plant populations allows for exchange and dispersal of adaptive genes, which can facilitate plant population persistence particularly in rapidly changing environments.

Methods

Patterns of historic gene flow, flowering phenology and contemporary pollen flow were investigated in two common herbs, Ranunculus bulbosus and Trifolium montanum, along an altitudinal gradient of 1200–1800 m a.s.l. over a distance of 1 km among five alpine meadows in Switzerland.

Key Results

Historic gene flow was extensive, as revealed by F_st values of 0·01 and 0·007 in R. bulbosus and T. montanum, respectively, by similar levels of allelic richness among meadows and by the grouping of all individuals into one genetic cluster. Our data suggest contemporary pollen flow is not limited across altitudes in either species but is more pronounced in T. montanum, as indicated by the differential decay of among-sibships correlated paternity with increasing spatial distance. Flowering phenology among meadows was not a barrier to pollen flow in T. montanum, as the large overlap between meadow pairs was consistent with the extensive pollen flow. The smaller flowering overlap among R. bulbosus meadows might explain the slightly more limited pollen flow detected.

Conclusions

High levels of pollen flow among altitudes in both R. bulbosus and T. montanum should facilitate exchange of genes which may enhance adaptive responses to rapid climate change.