How Does Selfing Affect the Pace and Process of Speciation?

Surprisingly little attention has been given to the impact of selfing on speciation, even though selfing reduces gene flow between populations and affects other key population genetics parameters. Here we review recent theoretical work and compile empirical data from crossing experiments and genomic and phylogenetic studies to assess the effect of mating systems on the speciation process. In accordance with theoretical predictions, we find that accumulation of hybrid incompatibilities seems to be accelerated in selfers, but there is so far limited empirical support for a predicted bias toward underdominant loci. Phylogenetic evidence is scarce and contradictory, including studies suggesting that selfing either promotes or hampers speciation rate. Further studies are therefore required, which in addition to measures of reproductive barrier strength and selfing rate should routinely include estimates of demographic history and genetic divergence as a proxy for divergence time.

Genetic and environmental manipulation of Arabidopsis hybridization barriers uncovers antagonistic functions in endosperm cellularization.

Speciation involves reproductive isolation, which can occur by hybridization barriers acting in the endosperm of the developing seed. The nuclear endosperm is a nutrient sink, accumulating sugars from surrounding tissues, and undergoes coordinated cellularization, switching to serve as a nutrient source for the developing embryo. Tight regulation of cellularization is therefore vital for seed and embryonic development. Here we show that hybrid seeds from crosses between Arabidopsis thaliana as maternal contributor and A. arenosa or A. lyrata as pollen donors result in an endosperm based post-zygotic hybridization barrier that gives rise to a reduced seed germination rate. Hybrid seeds display opposite endosperm cellularization phenotypes, with late cellularization in crosses with A. arenosa and early cellularization in crosses with A. lyrata. Stage specific endosperm reporters display temporally ectopic expression in developing hybrid endosperm, in accordance with the early and late cellularization phenotypes, confirming a disturbance of the source-sink endosperm phase change. We demonstrate that the hybrid barrier is under the influence of abiotic factors, and show that a temperature gradient leads to diametrically opposed cellularization phenotype responses in hybrid endosperm with A. arenosa or A. lyrata as pollen donors. Furthermore, different A. thaliana accession genotypes also enhance or diminish seed viability in the two hybrid cross-types, emphasizing that both genetic and environmental cues control the hybridization barrier. We have identified an A. thaliana MADS-BOX type I family single locus that is required for diametrically opposed cellularization phenotype responses in hybrid endosperm. Loss of AGAMOUS-LIKE 35 significantly affects the germination rate of hybrid seeds in opposite directions when transmitted through the A. thaliana endosperm, and is suggested to be a locus that promotes cellularization as part of an endosperm based mechanism involved in post-zygotic hybrid barriers. The role of temperature in hybrid speciation and the identification of distinct loci in control of hybrid failure have great potential to aid the introduction of advantageous traits in breeding research and to support models to predict hybrid admixture in a changing global climate.

Structural evidence for MADS-box type I family expansion seen in new assemblies of Arabidopsis arenosa and A. lyrata.

Arabidopsis thaliana diverged from A. arenosa and A. lyrata at least 6 million years ago. The three species differ by genome-wide polymorphisms and morphological traits. The species are to a high degree reproductively isolated, but hybridization barriers are incomplete. A special type of hybridization barrier is based on the triploid endosperm of the seed, where embryo lethality is caused by endosperm failure to support the developing embryo. The MADS-box type I family of transcription factors is specifically expressed in the endosperm and has been proposed to play a role in endosperm-based hybridization barriers. The gene family is well known for its high evolutionary duplication rate, as well as being regulated by genomic imprinting. Here we address MADS-box type I gene family evolution and the role of type I genes in the context of hybridization. Using two de-novo assembled and annotated chromosome-level genomes of A. arenosa and A. lyrata ssp. petraea we analyzed the MADS-box type I gene family in Arabidopsis to predict orthologs, copy number, and structural genomic variation related to the type I loci. Our findings were compared to gene expression profiles sampled before and after the transition to endosperm cellularization in order to investigate the involvement of MADS-box type I loci in endosperm-based hybridization barriers. We observed substantial differences in type-I expression in the endosperm of A. arenosa and A. lyrata ssp. petraea, suggesting a genetic cause for the endosperm-based hybridization barrier between A. arenosa and A. lyrata ssp. petraea.

Sedimentary ancient DNA metabarcoding as a tool for assessing prehistoric plant use at the Upper Paleolithic cave site Aghitu-3, Armenia.

Current knowledge about Paleolithic human plant use is limited by the rare survival of identifiable plant remains as well as the availability of methods for plant detection and identification. By analyzing DNA preserved in cave sediments, we can identify organisms in the absence of any visible remains, opening up new ways to study details of past human behavior, including plant use. Aghitu-3 Cave contains a 15,000-yearlong record (from ∼39,000 to 24,000 cal BP) of Upper Paleolithic human settlement and environmental variability in the Armenian Highlands. Finds from this cave include stone artifacts, faunal remains, bone tools, shell beads, charcoal, and pollen, among others. We applied sedimentary ancient DNA (sedaDNA) metabarcoding to the Aghitu-3 sedimentary sequence and combined this with pollen data to obtain a temporal reconstruction of plant assemblages. Our results reveal a stratification of plant abundance and diversity where sedaDNA reflects periods of human occupation, showing higher diversity in layers with increased human activity. Low pollen concentrations combined with high sedaDNA abundance indicate plant remains may have been brought into the cave by animals or humans during the deposition of the lower two archaeological horizons. Most of the recovered plants are reported to be useful for food, flavor, medicine, and/or technical purposes, demonstrating the potential of the environment around Aghitu-3 Cave to support humans during the Upper Paleolithic. Moreover, we identified several specific plant taxa that strengthen previous findings about Upper Paleolithic plant use in this region (i.e., for medicine and the manufacturing and dyeing of textiles). This study represents the first application of plant sedaDNA analysis of cave sediments for the investigation of potential plant use by prehistoric humans.

The genome of Draba nivalis shows signatures of adaptation to the extreme environmental stresses of the Arctic.

The Arctic is one of the most extreme terrestrial environments on the planet. Here, we present the first chromosome-scale genome assembly of a plant adapted to the high Arctic, Draba nivalis (Brassicaceae), an attractive model species for studying plant adaptation to the stresses imposed by this harsh environment. We used an iterative scaffolding strategy with data from short-reads, single-molecule long reads, proximity ligation data, and a genetic map to produce a 302 Mb assembly that is highly contiguous with 91.6% assembled into eight chromosomes (the base chromosome number). To identify candidate genes and gene families that may have facilitated adaptation to Arctic environmental stresses, we performed comparative genomic analyses with nine non-Arctic Brassicaceae species. We show that the D. nivalis genome contains expanded suites of genes associated with drought and cold stress (e.g., related to the maintenance of oxidation-reduction homeostasis, meiosis, and signaling pathways). The expansions of gene families associated with these functions appear to be driven in part by the activity of transposable elements. Tests of positive selection identify suites of candidate genes associated with meiosis and photoperiodism, as well as cold, drought, and oxidative stress responses. Our results reveal a multifaceted landscape of stress adaptation in the D. nivalis genome, offering avenues for the continued development of this species as an Arctic model plant.

Multiple Genetic Trajectories to Extreme Abiotic Stress Adaptation in Arctic Brassicaceae.

Extreme environments offer powerful opportunities to study how different organisms have adapted to similar selection pressures at the molecular level. Arctic plants have adapted to some of the coldest and driest biomes on Earth and typically possess suites of similar morphological and physiological adaptations to extremes in light and temperature. Here, we compare patterns of molecular evolution in three Brassicaceae species that have independently colonized the Arctic and present some of the first genetic evidence for plant adaptations to the Arctic environment. By testing for positive selection and identifying convergent substitutions in orthologous gene alignments for a total of 15 Brassicaceae species, we find that positive selection has been acting on different genes, but similar functional pathways in the three Arctic lineages. The positively selected gene sets identified in the three Arctic species showed convergent functional profiles associated with extreme abiotic stress characteristic of the Arctic. However, there was little evidence for independently fixed mutations at the same sites and for positive selection acting on the same genes. The three species appear to have evolved similar suites of adaptations by modifying different components in similar stress response pathways, implying that there could be many genetic trajectories for adaptation to the Arctic environment. By identifying candidate genes and functional pathways potentially involved in Arctic adaptation, our results provide a framework for future studies aimed at testing for the existence of a functional syndrome of Arctic adaptation in the Brassicaceae and perhaps flowering plants in general.

Genetic variation and temperature affects hybrid barriers during interspecific hybridization.

Genomic imprinting regulates parent-specific transcript dosage during seed development and is mainly confined to the endosperm. Elucidation of the function of many imprinted genes has been hampered by the lack of corresponding mutant phenotypes, and the role of imprinting is mainly associated with genome dosage regulation or allocation of resources. Disruption of imprinted genes has also been suggested to mediate endosperm-based post-zygotic hybrid barriers depending on genetic variation and gene dosage. Here, we have analyzed the conservation of a clade from the MADS-box type I class transcription factors in the closely related species Arabidopsis arenosa, A. lyrata, and A. thaliana, and show that AGL36-like genes are imprinted and maternally expressed in seeds of Arabidopsis species and in hybrid seeds between outbreeding species. In hybridizations between outbreeding and inbreeding species the paternally silenced allele of the AGL36-like gene is reactivated in the hybrid, demonstrating that also maternally expressed imprinted genes are perturbed during hybridization and that such effects on imprinted genes are specific to the species combination. Furthermore, we also demonstrate a quantitative effect of genetic diversity and temperature on the strength of the post-zygotic hybridization barrier. Markedly, a small decrease in temperature during seed development increases the survival of hybrid F1 seeds, suggesting that abiotic and genetic parameters play important roles in post-zygotic species barriers, pointing at evolutionary scenarios favoring such effects. OPEN RESEARCH BADGES: This article has earned an Open Data Badge for making publicly available the digitally-shareable data necessary to reproduce the reported results. The data is available at https://www.ncbi.nlm.nih.gov/bioproject/?term=PRJNA562212. All sequences generated in this study have been deposited in the National Center for Biotechnology Information Sequence Read Archive (https://www.ncbi.nlm.nih.gov/sra/) with project number PRJNA562212.

A single ectomycorrhizal plant root system includes a diverse and spatially structured fungal community.

Although only a relatively small proportion of plant species form ectomycorrhizae with fungi, it is crucial for growth and survival for a number of widespread woody plant species. Few studies have attempted to investigate the fine scale spatial structure of entire root systems of adult ectomycorrhizal (EcM) plants. Here, we use the herbaceous perennial Bistorta vivipara to map the entire root system of an adult EcM plant and investigate the spatial structure of its root-associated fungi. All EcM root tips were sampled, mapped and identified using a direct PCR approach and Sanger sequencing of the internal transcribed spacer region. A total of 32.1% of all sampled root tips (739 of 2302) were successfully sequenced and clustered into 41 operational taxonomic units (OTUs). We observed a clear spatial structuring of the root-associated fungi within the root system. Clusters of individual OTUs were observed in the younger parts of the root system, consistent with observations of priority effects in previous studies, but were absent from the older parts of the root system. This may suggest a succession and fragmentation of the root-associated fungi even at a very fine scale, where competition likely comes into play at different successional stages within the root system.

RADseq provides evidence for parallel ecotypic divergence in the autotetraploid Cochlearia officinalis in Northern Norway.

Speciation encompasses a continuum over time from freely interbreeding populations to reproductively isolated species. Along this process, ecotypes - the result of local adaptation - may be on the road to new species. We investigated whether three autotetraploid Cochlearia officinalis ecotypes, adapted to different habitats (beach, estuary, spring), are genetically differentiated and result from parallel ecotypic divergence in two distinct geographical regions. We obtained genetic data from thousands of single nucleotide polymorphisms (SNPs) from restriction-site associated DNA sequencing (RADseq) and from six microsatellite markers for 12 populations to assess genetic divergence at ecotypic, geographic and population level. The genetic patterns support differentiation among ecotypes as suggested by morphology and ecology. The data fit a scenario where the ancestral beach ecotype has recurrently and polytopically given rise to the estuary and spring ecotypes. Several ecologically-relevant loci with consistent non-random segregating patterns are identified across the recurrent origins, in particular around genes related to salt stress. Despite being ecologically distinct, the Cochlearia ecotypes still represent an early stage in the process of speciation, as reproductive isolation has not (yet) developed. A sequenced annotated genome is needed to specifically target candidate genes underlying local adaptation.

Veronica officinalis Product Authentication Using DNA Metabarcoding and HPLC-MS Reveals Widespread Adulteration with Veronica chamaedrys.

Studying herbal products derived from local and traditional knowledge and their value chains is one of the main challenges in ethnopharmacology. The majority of these products have a long history of use, but non-harmonized trade and differences in regulatory policies between countries impact their value chains and lead to concerns over product efficacy, safety and quality. Veronica officinalis L. (common speedwell), a member of Plantaginaceae family, has a long history of use in European traditional medicine, mainly in central eastern Europe and the Balkans. However, no specified control tests are available either to establish the quality of derived herbal products or for the discrimination of its most common substitute, V. chamaedrys L. (germander speedwell). In this study, we use DNA metabarcoding and high performance liquid chromatography coupled with mass spectrometry (HPLC-MS) to authenticate sixteen V. officinalis herbal products and compare the potential of the two approaches to detect substitution, adulteration and the use of unreported constituents. HPLC-MS showed high resolution in detecting phytochemical target compounds, but did not enable detection of specific plant species in the products. DNA metabarcoding detected V. officinalis in only 15% of the products, whereas it detected V. chamaedrys in 62% of the products. The results confirm that DNA metabarcoding can be used to test for the presence of Veronica species, and detect substitution and/or admixture of other Veronica species, as well as simultaneously detect all other species present. Our results confirm that none of the herbal products contained exactly the species listed on the label, and all included substitutes, contaminants or fillers. This study highlights the need for authentication of raw herbals along the value chain of these products. An integrative methodology can assess both the quality of herbal products in terms of target compound concentrations and species composition, as well as admixture and substitution with other chemical compounds and plants.

Correction: Hybrid Origins of Carex rostrata var. borealis and C. stenolepis, Two Problematic Taxa in Carex Section Vesicariae (Cyperaceae).

[This corrects the article DOI: 10.1371/journal.pone.0165430.].

Endosperm-based hybridization barriers explain the pattern of gene flow between Arabidopsis lyrata and Arabidopsis arenosa in Central Europe.

Based on the biological species concept, two species are considered distinct if reproductive barriers prevent gene flow between them. In Central Europe, the diploid species Arabidopsis lyrata and Arabidopsis arenosa are genetically isolated, thus fitting this concept as "good species." Nonetheless, interspecific gene flow involving their tetraploid forms has been described. The reasons for this ploidy-dependent reproductive isolation remain unknown. Here, we show that hybridization between diploid A. lyrata and A. arenosa causes mainly inviable seed formation, revealing a strong postzygotic reproductive barrier separating these two species. Although viability of hybrid seeds was impaired in both directions of hybridization, the cause for seed arrest differed. Hybridization of A. lyrata seed parents with A. arenosa pollen donors resulted in failure of endosperm cellularization, whereas the endosperm of reciprocal hybrids cellularized precociously. Endosperm cellularization failure in both hybridization directions is likely causal for the embryo arrest. Importantly, natural tetraploid A. lyrata was able to form viable hybrid seeds with diploid and tetraploid A. arenosa, associated with the reestablishment of normal endosperm cellularization. Conversely, the defects of hybrid seeds between tetraploid A. arenosa and diploid A. lyrata were aggravated. According to these results, we hypothesize that a tetraploidization event in A. lyrata allowed the production of viable hybrid seeds with A. arenosa, enabling gene flow between the two species.

Hybrid Origins of Carex rostrata var. borealis and C. stenolepis, Two Problematic Taxa in Carex Section Vesicariae (Cyperaceae).

Hybridization is frequent in the large and ecologically significant genus Carex (Cyperaceae). In four important sections of the northern regions (Ceratocystis, Glareosae, Phacocystis and Vesicariae), the frequent occurrence of hybrids often renders the identification of "pure" species and hybrids difficult. In this study we address the origins and taxonomic rank of two taxa of section Vesicariae: Carex rostrata var. borealis and C. stenolepis. The origin and taxonomic status of C. stenolepis has been the subject of substantial debate over the years, whereas C. rostrata var. borealis has received very little attention in the years since its first description in the 19th century. By performing an extensive sampling of relevant taxa from a broad distribution range, and analyzing data from fifteen microsatellite loci developed specifically for our study together with pollen stainability measures, we resolve the hybrid origins of C. rostrata var. borealis and C. stenolepis and provide new insights into this taxonomically challenging group of sedges. Our results are in accordance with previous findings suggesting that C. stenolepis is a hybrid between C. vesicaria and C. saxatilis. They are also in accordance with a previous proposition that C. rostrata var. borealis is a hybrid between C. rostrata and C. rotundata, and furthermore suggest that both hybrids are the result of multiple, recent (i.e., postglacial) hybridization events. We found little evidence for successful sexual reproduction within C. rostrata var. borealis and C. stenolepis, but conclude that the common and recurrent, largely predictable occurrence of these taxa justifies accepting both hybrids as hybrid species with binomial names. There are, however, complications as to types and priority names, and we therefore choose to address these problems in a separate paper.

From gene trees to a dated allopolyploid network: insights from the angiosperm genus Viola (Violaceae).

Allopolyploidization accounts for a significant fraction of speciation events in many eukaryotic lineages. However, existing phylogenetic and dating methods require tree-like topologies and are unable to handle the network-like phylogenetic relationships of lineages containing allopolyploids. No explicit framework has so far been established for evaluating competing network topologies, and few attempts have been made to date phylogenetic networks. We used a four-step approach to generate a dated polyploid species network for the cosmopolitan angiosperm genus Viola L. (Violaceae Batch.). The genus contains ca 600 species and both recent (neo-) and more ancient (meso-) polyploid lineages distributed over 16 sections. First, we obtained DNA sequences of three low-copy nuclear genes and one chloroplast region, from 42 species representing all 16 sections. Second, we obtained fossil-calibrated chronograms for each nuclear gene marker. Third, we determined the most parsimonious multilabeled genome tree and its corresponding network, resolved at the section (not the species) level. Reconstructing the "correct" network for a set of polyploids depends on recovering all homoeologs, i.e., all subgenomes, in these polyploids. Assuming the presence of Viola subgenome lineages that were not detected by the nuclear gene phylogenies ("ghost subgenome lineages") significantly reduced the number of inferred polyploidization events. We identified the most parsimonious network topology from a set of five competing scenarios differing in the interpretation of homoeolog extinctions and lineage sorting, based on (i) fewest possible ghost subgenome lineages, (ii) fewest possible polyploidization events, and (iii) least possible deviation from expected ploidy as inferred from available chromosome counts of the involved polyploid taxa. Finally, we estimated the homoploid and polyploid speciation times of the most parsimonious network. Homoploid speciation times were estimated by coalescent analysis of gene tree node ages. Polyploid speciation times were estimated by comparing branch lengths and speciation rates of lineages with and without ploidy shifts. Our analyses recognize Viola as an old genus (crown age 31 Ma) whose evolutionary history has been profoundly affected by allopolyploidy. Between 16 and 21 allopolyploidizations are necessary to explain the diversification of the 16 major lineages (sections) of Viola, suggesting that allopolyploidy has accounted for a high percentage-between 67% and 88%-of the speciation events at this level. The theoretical and methodological approaches presented here for (i) constructing networks and (ii) dating speciation events within a network, have general applicability for phylogenetic studies of groups where allopolyploidization has occurred. They make explicit use of a hitherto underexplored source of ploidy information from chromosome counts to help resolve phylogenetic cases where incomplete sequence data hampers network inference. Importantly, the coalescent-based method used herein circumvents the assumption of tree-like evolution required by most techniques for dating speciation events.

Ethnolinguistic structuring of sorghum genetic diversity in Africa and the role of local seed systems.

Sorghum is a drought-tolerant crop with a vital role in the livelihoods of millions of people in marginal areas. We examined genetic structure in this diverse crop in Africa. On the continent-wide scale, we identified three major sorghum populations (Central, Southern, and Northern) that are associated with the distribution of ethnolinguistic groups on the continent. The codistribution of the Central sorghum population and the Nilo-Saharan language family supports a proposed hypothesis about a close and causal relationship between the distribution of sorghum and languages in the region between the Chari and the Nile rivers. The Southern sorghum population is associated with the Bantu languages of the Niger-Congo language family, in agreement with the farming-language codispersal hypothesis as it has been related to the Bantu expansion. The Northern sorghum population is distributed across early Niger-Congo and Afro-Asiatic language family areas with dry agroclimatic conditions. At a finer geographic scale, the genetic substructure within the Central sorghum population is associated with language-group expansions within the Nilo-Saharan language family. A case study of the seed system of the Pari people, a Western-Nilotic ethnolinguistic group, provides a window into the social and cultural factors involved in generating and maintaining the continent-wide diversity patterns. The age-grade system, a cultural institution important for the expansive success of this ethnolinguistic group in the past, plays a central role in the management of sorghum landraces and continues to underpin the resilience of their traditional seed system.

Modern maize varieties going local in the semi-arid zone in Tanzania.

BACKGROUND: Maize is the most produced crop in Sub-Saharan Africa, but yields are low and climate change is projected to further constrain smallholder production. The current efforts to breed and disseminate new high yielding and climate ready maize varieties are implemented through the formal seed system; the chain of public and private sector activities and institutions that produce and release certified seeds. These efforts are taking place in contexts currently dominated by informal seed systems; local and informal seed management and exchange channels with a long history of adapting crops to local conditions. We here present a case study of the genetic effects of both formal and informal seed management from the semi-arid zone in Tanzania. RESULTS: Two open pollinated varieties (OPVs), Staha and TMV1, first released by the formal seed system in the 1980s are cultivated on two-thirds of the maize fields among the surveyed households. Farmer-recycling of improved varieties and seed selection are common on-farm seed management practices. Drought tolerance and high yield are the most important characteristics reported as reason for cultivating the current varieties as well as the most important criteria for farmers' seed selection. Bayesian cluster analysis, PCA and FST analyses based on 131 SNPs clearly distinguish between the two OPVs, and despite considerable heterogeneity between and within seed lots, there is insignificant differentiation between breeder's seeds and commercial seeds in both OPVs. Genetic separation increases as the formal system varieties enter the informal system and both hybridization with unrelated varieties and directional selection probably play a role in the differentiation. Using a Bayesian association approach we identify three loci putatively under selection in the informal seed system. CONCLUSIONS: Our results suggest that the formal seed system in the study area distributes seed lots that are true to type. We suggest that hybridization and directional selection differentiate farmer recycled seed lots from the original varieties and potentially lead to beneficial creolization. Access to drought tolerant OPVs in combination with farmer seed selection is likely to enhance seed system security and farmers' adaptive capacity in the face of climate change.

Substantial compositional turnover of fungal communities in an alpine ridge-to-snowbed gradient.

The main gradient in vascular plant, bryophyte and lichen species composition in alpine areas, structured by the topographic gradient from wind-exposed ridges to snowbeds, has been extensively studied. Tolerance to environmental stress, resulting from wind abrasion and desiccation towards windswept ridges or reduced growing season due to prolonged snow cover towards snowbeds, is an important ecological mechanism in this gradient. The extent to which belowground fungal communities are structured by the same topographic gradient and the eventual mechanisms involved are less well known. In this study, we analysed variation in fungal diversity and community composition associated with roots of the ectomycorrhizal plant Bistorta vivipara along the ridge-to-snowbed gradient. We collected root samples from fifty B. vivipara plants in ten plots in an alpine area in central Norway. The fungal communities were analysed using 454 pyrosequencing analyses of tag-encoded ITS1 amplicons. A distinct gradient in the fungal community composition was found that coincided with variation from ridge to snowbeds. This gradient was paralleled by change in soil content of carbon, nitrogen and phosphorus. A large proportion (66%) of the detected 801 nonsingleton operational taxonomic units (OTUs) were ascomycetes, while basidiomycetes dominated quantitatively (i.e. with respect to number of reads). Numerous fungal OTUs, many with taxonomic affinity to Sebacinales, Cortinarius and Meliniomyces, showed distinct affinities either to ridge or to snowbed plots, indicating habitat specialization. The compositional turnover of fungal communities along the gradient was not paralleled by a gradient in species richness.

Spatial structure and climatic adaptation in African maize revealed by surveying SNP diversity in relation to global breeding and landrace panels.

BACKGROUND: Climate change threatens maize productivity in sub-Saharan Africa. To ensure food security, access to locally adapted genetic resources and varieties is an important adaptation measure. Most of the maize grown in Africa is a genetic mix of varieties introduced at different historic times following the birth of the trans-Atlantic economy, and knowledge about geographic structure and local adaptations is limited. METHODOLOGY: A panel of 48 accessions of maize representing various introduction routes and sources of historic and recent germplasm introductions in Africa was genotyped with the MaizeSNP50 array. Spatial genetic structure and genetic relationships in the African panel were analysed separately and in the context of a panel of 265 inbred lines representing global breeding material (based on 26,900 SNPs) and a panel of 1127 landraces from the Americas (270 SNPs). Environmental association analysis was used to detect SNPs associated with three climatic variables based on the full 43,963 SNP dataset. CONCLUSIONS: The genetic structure is consistent between subsets of the data and the markers are well suited for resolving relationships and admixture among the accessions. The African accessions are structured in three clusters reflecting historical and current patterns of gene flow from the New World and within Africa. The Sahelian cluster reflects original introductions of Meso-American landraces via Europe and a modern introduction of temperate breeding material. The Western cluster reflects introduction of Coastal Brazilian landraces, as well as a Northeast-West spread of maize through Arabic trade routes across the continent. The Eastern cluster most strongly reflects gene flow from modern introduced tropical varieties. Controlling for population history in a linear model, we identify 79 SNPs associated with maximum temperature during the growing season. The associations located in genes of known importance for abiotic stress tolerance are interesting candidates for local adaptations.

High consistency between replicate 454 pyrosequencing analyses of ectomycorrhizal plant root samples.

In this methodological study, we compare 454 sequencing and a conventional cloning and Sanger sequencing approach in their ability to characterize fungal communities PCR amplified from four root systems of the ectomycorrhizal plant Bistorta vivipara. To examine variation introduced by stochastic processes during the laboratory work, we replicated all analyses using two independently obtained DNA extractions from the same root systems. The ITS1 region was used as DNA barcode and the sequences were clustered into OTUs as proxies for species using single linkage clustering (BLASTC: lust) and 97% sequence similarity cut-off. A relatively low overlap in fungal OTUs was observed between the 454 and the clone library datasets - even among the most abundant OTUs. In a non-metric multidimensional scaling analysis, the samples grouped more according to methodology compared to plant. Some OTUs frequently detected by 454, most notably those OTUs with taxonomic affinity to Glomales, were not detected in the Sanger dataset. Likewise, a few OTUs, including Cenococcum sp., only appeared in the clone libraries. Surprisingly, we observed a significant relationship between GC/AT content of the OTUs and their proportional abundances in the 454 versus the clone library datasets. Reassuringly, a very good consistency in OTU recovery was observed between replicate runs of both sequencing methods. This indicates that stochastic processes had little impact when applying the same sequencing technique on replicate samples.

Inferring species networks from gene trees in high-polyploid North American and Hawaiian violets (Viola, Violaceae).

The phylogenies of allopolyploids take the shape of networks and cannot be adequately represented as bifurcating trees. Especially for high polyploids (i.e., organisms with more than six sets of nuclear chromosomes), the signatures of gene homoeolog loss, deep coalescence, and polyploidy may become confounded, with the result that gene trees may be congruent with more than one species network. Herein, we obtained the most parsimonious species network by objective comparison of competing scenarios involving polyploidization and homoeolog loss in a high-polyploid lineage of violets (Viola, Violaceae) mostly or entirely restricted to North America, Central America, or Hawaii. We amplified homoeologs of the low-copy nuclear gene, glucose-6-phosphate isomerase (GPI), by single-molecule polymerase chain reaction (PCR) and the chloroplast trnL-F region by conventional PCR for 51 species and subspecies. Topological incongruence among GPI homoeolog subclades, owing to deep coalescence and two instances of putative loss (or lack of detection) of homoeologs, were reconciled by applying the maximum tree topology for each subclade. The most parsimonious species network and the fossil-based calibration of the homoeolog tree favored monophyly of the high polyploids, which has resulted from allodecaploidization 9-14 Ma, involving sympatric ancestors from the extant Viola sections Chamaemelanium (diploid), Plagiostigma (paleotetraploid), and Viola (paleotetraploid). Although two of the high-polyploid lineages (Boreali-Americanae, Pedatae) remained decaploid, recurrent polyploidization with tetraploids of section Plagiostigma within the last 5 Ma has resulted in two 14-ploid lineages (Mexicanae, Nosphinium) and one 18-ploid lineage (Langsdorffianae). This implies a more complex phylogenetic and biogeographic origin of the Hawaiian violets (Nosphinium) than that previously inferred from rDNA data and illustrates the necessity of considering polyploidy in phylogenetic and biogeographic reconstruction.