A Bird's Eye View of the Systematics of Convolvulaceae: Novel Insights From Nuclear Genomic Data.

Convolvulaceae is a family of c. 2,000 species, distributed across 60 currently recognized genera. It includes species of high economic importance, such as the crop sweet potato (Ipomoea batatas L.), the ornamental morning glories (Ipomoea L.), bindweeds (Convolvulus L.), and dodders, the parasitic vines (Cuscuta L.). Earlier phylogenetic studies, based predominantly on chloroplast markers or a single nuclear region, have provided a framework for systematic studies of the family, but uncertainty remains at the level of the relationships among subfamilies, tribes, and genera, hindering evolutionary inferences and taxonomic advances. One of the enduring enigmas has been the relationship of Cuscuta to the rest of Convolvulaceae. Other examples of unresolved issues include the monophyly and relationships within Merremieae, the "bifid-style" clade (Dicranostyloideae), as well as the relative positions of Erycibe Roxb. and Cardiochlamyeae. In this study, we explore a large dataset of nuclear genes generated using Angiosperms353 kit, as a contribution to resolving some of these remaining phylogenetic uncertainties within Convolvulaceae. For the first time, a strongly supported backbone of the family is provided. Cuscuta is confirmed to belong within family Convolvulaceae. "Merremieae," in their former tribal circumscription, are recovered as non-monophyletic, with the unexpected placement of Distimake Raf. as sister to the clade that contains Ipomoeeae and Decalobanthus Ooststr., and Convolvuleae nested within the remaining "Merremieae." The monophyly of Dicranostyloideae, including Jacquemontia Choisy, is strongly supported, albeit novel relationships between genera are hypothesized, challenging the current tribal delimitation. The exact placements of Erycibe and Cuscuta remain uncertain, requiring further investigation. Our study explores the benefits and limitations of increasing sequence data in resolving higher-level relationships within Convolvulaceae, and highlights the need for expanded taxonomic sampling, to facilitate a much-needed revised classification of the family.

The evolutionary history of the Caribbean magnolias (Magnoliaceae): Testing species delimitations and biogeographical hypotheses using molecular data.

The Caribbean islands provide an ideal setting for studying biodiversity, given their complex geological and environmental history, and their historical and current geographical proximity to the American mainland. Magnolia, a flagship tree genus that has 15 endemic and threatened taxa (12 species and 3 subspecies) on the Caribbean islands, offers an excellent case study to empirically test Caribbean biogeographical hypotheses. We constructed phylogenetic hypotheses to: (1) reveal their evolutionary history, (2) test the current largely morphology-based classification and assess species limits, and (3) investigate major biogeographic hypotheses proposed for the region. Nuclear and chloroplast DNA sequence data of all 15 Caribbean Magnolia taxa are included, supplemented by a selection of American mainland species, and species representing most major clades of the Magnoliaceae family. We constructed phylogenetic hypotheses in a time-calibrated Bayesian framework, supplemented with haplotype network analyses and ancestral range estimations. Genetic synapomorphies in the studied markers confirm the species limits of 14 out of 15 morphologically recognizable Caribbean Magnolia taxa. There is evidence for four colonization events of Magnolia into the Caribbean from the American mainland, which most likely occurred by overwater dispersal, given age estimates of maximum 16 mya for their presence on the Caribbean islands.

Chromosome-level reference genome of the soursop (Annona muricata): A new resource for Magnoliid research and tropical pomology.

The flowering plant family Annonaceae includes important commercially grown tropical crops, but development of promising species is hindered by a lack of genomic resources to build breeding programs. Annonaceae are part of the magnoliids, an ancient lineage of angiosperms for which evolutionary relationships with other major clades remain unclear. To provide resources to breeders and evolutionary researchers, we report a chromosome-level genome assembly of the soursop (Annona muricata). We assembled the genome using 444.32 Gb of DNA sequences (676× sequencing depth) from PacBio and Illumina short-reads, in combination with 10× Genomics and Bionano data (v1). A total of 949 scaffolds were assembled to a final size of 656.77 Mb, with a scaffold N50 of 3.43 Mb (v1), and then further improved to seven pseudo-chromosomes using Hi-C sequencing data (v2; scaffold N50: 93.2 Mb, total size in chromosomes: 639.6 Mb). Heterozygosity was very low (0.06%), while repeat sequences accounted for 54.87% of the genome, and 23,375 protein-coding genes with an average of 4.79 exons per gene were annotated using de novo, RNA-seq and homology-based approaches. Reconstruction of the historical population size showed a slow continuous contraction, probably related to Cenozoic climate changes. The soursop is the first genome assembled in Annonaceae, supporting further studies of floral evolution in magnoliids, providing an essential resource for delineating relationships of ancient angiosperm lineages. Both genome-assisted improvement and conservation efforts will be strengthened by the availability of the soursop genome. As a community resource, this assembly will further strengthen the role of Annonaceae as model species for research on the ecology, evolution and domestication potential of tropical species in pomology and agroforestry.

Twelve new and exciting Annonaceae from the Neotropics.

As a result of concerted efforts of the community of Annonaceae taxonomists, increasingly detailed knowledge of the diversity of the Neotropical genera has been documented. With the exception of just two large genera, Annona and Xylopia, all Neotropical Annonaceae have been revised within the last 25 years. Subsequent to these publications, many new specimens have been collected and sent to us in Leiden for identification. These included a number that, despite the advanced state of taxonomic knowledge, proved to represent rarely collected, undescribed species. Here we describe 12 new species of Annona, Guatteria, Klarobelia, Tetrameranthus, and Xylopia. These species serve to illustrate the still underestimated diversity of the Neotropical flora, even in well studied plant groups like Annonaceae.

A taxonomic revision of the Neotropical genus Cremastosperma (Annonaceae), including five new species.

We present a taxonomic revision of Cremastosperma, a genus of Neotropical Annonaceae occurring in lowland to premontane wet forest, mostly in areas surrounding the Andean mountain chain. We recognise 34 species, describing five as new here: from east of the Andes, C.brachypodum Pirie & Chatrou, sp. nov. and C.dolichopodum Pirie & Maas, sp. nov., endemic to Peru; C.confusum Pirie, sp. nov., from southern Peru and adjacent Bolivia and Brazil; and C.alticola Pirie & Chatrou, sp. nov., at higher elevations in northern Peru and Ecuador; and from west of the Andes, C.osicola Pirie & Chatrou, sp. nov. endemic to Costa Rica, the most northerly distributed species of the genus. We provide an identification key, document diagnostic characters and distributions and provide illustrations and extensive lists of specimens, also presenting the latter in the form of mapping data with embedded links to images available online. Of the 34 species, 22 are regional endemics. On the basis of the extent of occurrence and area of occupancy of species estimated from the distribution data, we designate IUCN threat categries for all species. Fourteen species proved to be endangered (EN) and a further one critically endangered (CR), reflecting their rarity and narrow known distributions.

Floral evolution by simplification in Monanthotaxis (Annonaceae) and hypotheses for pollination system shifts.

Simplification by reduction has occurred many times independently in the floral evolution of angiosperms. These reductions have often been attributed to changes in reproductive biology. In the angiosperm plant family Annonaceae, most species have flowers with six petals, and many stamens and carpels. In the genus Monanthotaxis several deviations from this pattern have been observed, including flowers that contain three petals and three stamens only. New DNA sequences were generated for 42 specimens of Monanthotaxis. Five chloroplast markers and two nuclear markers for 72 out of 94 species of Monanthotaxis were used to reconstruct a phylogeny of the genus, which revealed several well-supported, morphologically distinct clades. The evolution of four quantitative and two qualitative floral characters was mapped onto this phylogeny, demonstrating a reduction in flower size and number of flower parts in Monanthotaxis. A large variation in stamen forms and numbers, strong correlations between petal size, stamen and carpel number, combined with a non-gradual mode of evolution and the sympatric co-occurrence of Monanthotaxis species from different clades suggest that the high diversity in the African rainforest of this genus is caused by switches in pollination systems.

Parallel diversifications of Cremastosperma and Mosannona (Annonaceae), tropical rainforest trees tracking Neogene upheaval of South America.

Much of the immense present day biological diversity of Neotropical rainforests originated from the Miocene onwards, a period of geological and ecological upheaval in South America. We assess the impact of the Andean orogeny, drainage of Lake Pebas and closure of the Panama isthmus on two clades of tropical trees (Cremastosperma, ca 31 spp.; and Mosannona, ca 14 spp.; both Annonaceae). Phylogenetic inference revealed similar patterns of geographically restricted clades and molecular dating showed diversifications in the different areas occurred in parallel, with timing consistent with Andean vicariance and Central American geodispersal. Ecological niche modelling approaches show phylogenetically conserved niche differentiation, particularly within Cremastosperma. Niche similarity and recent common ancestry of Amazon and Guianan Mosannona species contrast with dissimilar niches and more distant ancestry of Amazon, Venezuelan and Guianan species of Cremastosperma, suggesting that this element of the similar patterns of disjunct distributions in the two genera is instead a biogeographic parallelism, with differing origins. The results provide further independent evidence for the importance of the Andean orogeny, the drainage of Lake Pebas, and the formation of links between South and Central America in the evolutionary history of Neotropical lowland rainforest trees.

A linear sequence to facilitate curation of herbarium specimens of Annonaceae.

This paper provides a linear sequence of four subfamilies, 15 tribes and 106 genera of the magnoliid family Annonaceae, based on state-of-the-art and stable phylogenetic relationships. The linear sequence facilitates the organisation of Annonaceae herbarium specimens.

Anatolian origins and diversification of Aethionema, the sister lineage of the core Brassicaceae.

PREMISE OF THE STUDY: The Irano-Turanian region harbors three biodiversity hotspots and ∼25% of Brassicaceae species are endemic to the region. Aethionema (∼61 species) is the sister lineage to the core Brassicaceae and occurs mainly in the Irano-Turanian region. The evolutionary important position of Aethionema makes it an ideal reference for broader comparative genetics and genomics. To understand the evolution of Aethionema, and for a broader understanding of crucifer evolution, a time-calibrated phylogenetic tree and biogeographical history of the genus is needed. METHODS: Seventy-six plastome coding regions and nuclear rDNA genes, mainly from herbarium material, covering 75% of all Aethionema species, were used to resolve a time-calibrated Aethionema phylogeny. The different clades were characterized based on four morphological characters. The ancestral area of Aethionema was estimated with historical biogeographical analyses. KEY RESULTS: Three well-supported major clades within Aethionema were resolved. The ancestral area reconstruction and divergence-time estimates are consistent with major dispersal events during the Pliocene from the Anatolian Diagonal. CONCLUSIONS: We find that most Aethionema lineages originated along the Anatolian Diagonal, a floristic bridge connecting the east to the west, during the Pliocene. The dispersal of Aethionema correlates with the local geological events, such as the uplift of the Anatolian and Iranian plateaus and the formation of the major mountain ranges of the Irano-Turanian region. Knowing the paleo-ecological context for the evolution of Aethionema, in addition to the other lineages of Brassicaceae, facilitates our broader understanding for trait evolution and species diversification across the Brassicaceae.

Correlated evolutionary rates across genomic compartments in Annonaceae.

The molecular clock hypothesis is an important concept in biology. Deviations from a constant rate of nucleotide substitution have been found widely among lineages, genomes, genes and individual sites. Phylogenetic research can accommodate for these differences in applying specific models of evolution. Lineage-specific rate heterogeneity however can generate bi- or multimodal distributions of substitution rates across the branches of a tree and this may mislead phylogenetic inferences with currently available models. The plant family Annonaceae is an excellent case to study lineage-specific rate heterogeneity. The two major sister subfamilies, Annonoideae and Malmeoideae, have shown great discrepancies in branch lengths. We used high-throughput sequencing data of 72 genes, 99 spacers and 16 introns from 24 chloroplast genomes and nuclear ribosomal DNA of 23 species to study the molecular rate of evolution in Annonaceae. In all analyses, longer branch lengths and/or higher substitution rates were found for the Annonoideae compared to the Malmeoideae. The Annonaceae had wide variability in chloroplast length, ranging from minimal 175,684bp to 201,723 for Annonoideae and minimal 152,357 to 170,985bp in Malmeoideae, mostly reflecting variation in inverted-repeat length. The Annonoideae showed a higher GC-content in the conserved parts of the chloroplast genome and higher omega (dN/dS)-ratios than the Malmeoideae, which could indicate less stringent purifying selection, a pattern that has been found in groups with small population sizes. This study generates new insights into the processes causing lineage-specific rate heterogeneity, which could lead to improved phylogenetic methods.

Extended molecular phylogenetics and revised systematics of Malagasy scincine lizards.

Among the endemic biota of Madagascar, skinks are a diverse radiation of lizards that exhibit a striking ecomorphological variation, and could provide an interesting system to study body-form evolution in squamate reptiles. We provide a new phylogenetic hypothesis for Malagasy skinks of the subfamily Scincinae based on an extended molecular dataset comprising 8060bp from three mitochondrial and nine nuclear loci. Our analysis also increases taxon sampling of the genus Amphiglossus by including 16 out of 25 nominal species. Additionally, we examined whether the molecular phylogenetic patterns coincide with morphological differentiation in the species currently assigned to this genus. Various methods of inference recover a mostly strongly supported phylogeny with three main clades of Amphiglossus. However, relationships among these three clades and the limb-reduced genera Grandidierina, Voeltzkowia and Pygomeles remain uncertain. Supported by a variety of morphological differences (predominantly related to the degree of body elongation), but considering the remaining phylogenetic uncertainty, we propose a redefinition of Amphiglossus into three different genera (Amphiglossus sensu stricto, Flexiseps new genus, and Brachyseps new genus) to remove the non-monophyly of Amphiglossus sensu lato and to facilitate future studies on this fascinating group of lizards.

A nonet of novel species of Monanthotaxis (Annonaceae) from around Africa.

As part of an ongoing revision of the genus Monanthotaxis Baill. (Annonaceae), nine new species are described and one variety is reinstated to species rank. Two new species from West Africa (Monanthotaxis aquila P.H.Hoekstra, sp. nov. and Monanthotaxis atewensis P.H.Hoekstra, sp. nov.), four new species from Central Africa (Monanthotaxis couvreurii P.H.Hoekstra, sp. nov., Monanthotaxis latistamina P.H.Hoekstra, sp. nov., Monanthotaxis tripetala P.H.Hoekstra, sp. nov. and Monanthotaxis zenkeri P.H.Hoekstra, sp. nov.), one new species from Tanzania (Monanthotaxis filipes P.H.Hoekstra, sp. nov.), one new species from the area around Maputo (Monanthotaxis maputensis P.H.Hoekstra, sp. nov.), one new species from the Comoro Islands (Monanthotaxis komorensis P.H.Hoekstra, sp. nov.) and Monanthotaxis klainei (Engl.) Verdc. var. angustifolia (Boutique) Verdc. is raised to species level leading to the replacement name Monanthotaxis atopostema P.H.Hoekstra, nom. nov. (not Monanthotaxis angustifolia (Exell) Verdc.). Complete descriptions, comparisons with related species, ecological information and IUCN conservation assessments are given for the new species. Five species were classified as critical endangered, two species as endangered, one as vulnerable and one as least concern, warranting the need of further collecting and studying those species.

Recently evolved diversity and convergent radiations of rainforest mahoganies (Meliaceae) shed new light on the origins of rainforest hyperdiversity.

Tropical rainforest hyperdiversity is often suggested to have evolved over a long time-span (the 'museum' model), but there is also evidence for recent rainforest radiations. The mahoganies (Meliaceae) are a prominent plant group in lowland tropical rainforests world-wide but also occur in all other tropical ecosystems. We investigated whether rainforest diversity in Meliaceae has accumulated over a long time or has more recently evolved. We inferred the largest time-calibrated phylogeny for the family to date, reconstructed ancestral states for habitat and deciduousness, estimated diversification rates and modeled potential shifts in macro-evolutionary processes using a recently developed Bayesian method. The ancestral Meliaceae is reconstructed as a deciduous species that inhabited seasonal habitats. Rainforest clades have diversified from the Late Oligocene or Early Miocene onwards. Two contemporaneous Amazonian clades have converged on similar ecologies and high speciation rates. Most species-level diversity of Meliaceae in rainforest is recent. Other studies have found steady accumulation of lineages, but the large majority of plant species diversity in rainforests is recent, suggesting (episodic) species turnover. Rainforest hyperdiversity may best be explained by recent radiations from a large stock of higher level taxa.

Phylogenetics, ancestral state reconstruction, and a new infrafamilial classification of the pantropical Ochnaceae (Medusagynaceae, Ochnaceae s.str., Quiinaceae) based on five DNA regions.

Ochnaceae s.str. (Malpighiales) are a pantropical family of about 500 species and 27 genera of almost exclusively woody plants. Infrafamilial classification and relationships have been controversial partially due to the lack of a robust phylogenetic framework. Including all genera except Indosinia and Perissocarpa and DNA sequence data for five DNA regions (ITS, matK, ndhF, rbcL, trnL-F), we provide for the first time a nearly complete molecular phylogenetic analysis of Ochnaceae s.l. resolving most of the phylogenetic backbone of the family. Based on this, we present a new classification of Ochnaceae s.l., with Medusagynoideae and Quiinoideae included as subfamilies and the former subfamilies Ochnoideae and Sauvagesioideae recognized at the rank of tribe. Our data support a monophyletic Ochneae, but Sauvagesieae in the traditional circumscription is paraphyletic because Testulea emerges as sister to the rest of Ochnoideae, and the next clade shows Luxemburgia+Philacra as sister group to the remaining Ochnoideae. To avoid paraphyly, we classify Luxemburgieae and Testuleeae as new tribes. The African genus Lophira, which has switched between subfamilies (here tribes) in past classifications, emerges as sister to all other Ochneae. Thus, endosperm-free seeds and ovules with partly to completely united integuments (resulting in an apparently single integument) are characters that unite all members of that tribe. The relationships within its largest clade, Ochnineae (former Ochneae), are poorly resolved, but former Ochninae (Brackenridgea, Ochna) are polyphyletic. Within Sauvagesieae, the genus Sauvagesia in its broad circumscription is polyphyletic as Sauvagesia serrata is sister to a clade of Adenarake, Sauvagesia spp., and three other genera. Within Quiinoideae, in contrast to former phylogenetic hypotheses, Lacunaria and Touroulia form a clade that is sister to Quiina. Bayesian ancestral state reconstructions showed that zygomorphic flowers with adaptations to buzz-pollination (poricidal anthers), a syncarpous gynoecium (a near-apocarpous gynoecium evolved independently in Quiinoideae and Ochninae), numerous ovules, septicidal capsules, and winged seeds with endosperm are the ancestral condition in Ochnoideae. Although in some lineages poricidal anthers were lost secondarily, the evolution of poricidal superstructures secured the maintenance of buzz-pollination in some of these genera, indicating a strong selective pressure on keeping that specialized pollination system.

A plastid DNA phylogeny of tribe Miliuseae: insights into relationships and character evolution in one of the most recalcitrant major clades of Annonaceae.

PREMISE OF THE STUDY: Tribe Miliuseae (∼25 genera and ∼510 species) includes a substantial part of the species and generic diversity in the pantropical flowering-plant family Annonaceae (∼108 genera and ∼2400 species). Previous molecular phylogenetic analyses have failed to resolve the backbone phylogeny of the tribe, impeding biogeographical and evolutionary studies. We use a dense generic taxon sample (∼89% of generic diversity in Miliuseae) and plastid DNA sequence data (∼7 kb) to clarify the phylogenetic relationships of and within the tribe. METHODS: Parsimony and Bayesian phylogenetic reconstructions and ancestral character-state reconstructions of several reproductive characters were performed. KEY RESULTS: Dendrokingstoniae, Monocarpieae, and Miliuseae are recovered in a strongly supported clade, and each tribe is strongly supported as monophyletic. Miliuseae are characterized by a synapomorphic cryptoaperturate/disulculate pollen apertural system. Stenanona is shown to be nested within the paraphyletic genus Desmopsis. The only Neotropical clade (Sapranthus, Tridimeris, Desmopsis, and Stenanona) in the predominantly Asian Miliuseae is shown to be closely related to an undescribed genus from continental Southeast Asia and the Indo-Malayan and Austral-Pacific genus Meiogyne. Ancestral character-state reconstructions of several reproductive characters that are diagnostically important at the generic level indicate a considerable degree of homoplasy. CONCLUSIONS: The results improve our understanding of the relationships of and within Miliuseae, but parts of the backbone of the phylogeny remain poorly supported. Additional data from variable nuclear markers or reduced-genome-representation approaches seem to be required to further resolve relationships within this recalcitrant clade.

Little ecological divergence associated with speciation in two African rain forest tree genera.

BACKGROUND: The tropical rain forests (TRF) of Africa are the second largest block of this biome after the Amazon and exhibit high levels of plant endemism and diversity. Two main hypotheses have been advanced to explain speciation processes that have led to this high level of biodiversity: allopatric speciation linked to geographic isolation and ecological speciation linked to ecological gradients. Both these hypotheses rely on ecology: in the former conservation of ecological niches through time is implied, while in the latter adaptation via selection to alternative ecological niches would be a prerequisite. Here, we investigate the role of ecology in explaining present day species diversity in African TRF using a species level phylogeny and ecological niche modeling of two predominantly restricted TRF tree genera, Isolona and Monodora (Annonaceae). Both these genera, with 20 and 14 species, respectively, are widely distributed in African TRFs, with a few species occurring in slightly less humid regions such as in East Africa. RESULTS: A total of 11 sister species pairs were identified most of them occurring in allopatry or with little geographical overlap. Our results provide a mixed answer on the role of ecology in speciation. Although no sister species have identical niches, just under half of the tests suggest that sister species do have more similar niches than expected by chance. PCA analyses also support little ecological differences between sister species. Most speciation events within both genera predate the Pleistocene, occurring during the Late Miocene and Pliocene periods. CONCLUSIONS: Ecology is almost always involved in speciation, however, it would seem to have had a little role in species generation within Isolona and Monodora at the scale analyzed here. This is consistent with the geographical speciation model for TRF diversification. These results contrast to other studies for non-TRF plant species where ecological speciation was found to be an important factor of diversification. The Pliocene period appears to be a vital time in the generation of African TRF diversity, whereas Pleistocene climatic fluctuations have had a smaller role on speciation than previously thought.Ecological niche modeling, species level phylogeny, ecological speciation, African tropics, Isolona, Monodora, Annonaceae.

Insights into the influence of priors in posterior mapping of discrete morphological characters: a case study in Annonaceae.

BACKGROUND: Posterior mapping is an increasingly popular hierarchical Bayesian based method used to infer character histories and reconstruct ancestral states at nodes of molecular phylogenies, notably of morphological characters. As for all Bayesian analyses specification of prior values is an integrative and important part of the analysis. He we provide an example of how alternative prior choices can seriously influence results and mislead interpretations. METHODS/PRINCIPAL FINDINGS: For two contrasting discrete morphological characters, namely a slow and a fast evolving character found in the plant family Annonaceae, we specified a total of eight different prior distributions per character. We investigated how these prior settings affected important summary statistics. Our analyses showed that the different prior distributions had marked effects on the results in terms of average number of character state changes. These differences arise because priors play a crucial role in determining which areas of parameter space the values of the simulation will be drawn from, independent of the data at hand. However, priors seemed to fit the data better if they would result in a more even sampling of parameter space (normal posterior distribution), in which case alternative standard deviation values had little effect on the results. The most probable character history for each character was affected differently by the prior. For the slower evolving character, the same character history always had the highest posterior probability independent of the priors used. In contrast, the faster evolving character showed different most probable character histories depending on the prior. These differences could be related to the level of homoplasy exhibited by each character. CONCLUSIONS: Although our analyses were restricted to two morphological characters within a single family, our results underline the importance of carefully choosing prior values for posterior mapping. Prior specification will be of crucial importance when interpreting the results in a meaningful way. It is hard to suggest a statistically sound method for prior specification without more detailed studies. Meanwhile, we propose that the data could be used to estimate the prior value of the gamma distribution placed on the transformation rate in posterior mapping.

Flanking regions of monomorphic microsatellite loci provide a new source of data for plant species-level phylogenetics.

Well-resolved phylogenetic trees are essential for us to understand evolutionary processes at the level of species. The degree of species-level resolution in the plant phylogenetic literature is poor, however, largely due to the dearth of sufficiently variable molecular markers. Unlike the common genic approach to marker development, we generated DNA sequences of monomorphic nuclear microsatellite flanking regions in a phylogenetic study of Annona species (Annonaceae). The resulting data showed no evidence of paralogy or allelic diversity that would confound attempts to reconstruct the species tree. Microsatellite flanking regions are short, making them practical to use, yet have astounding proportions of variable characters. They have 3.5- to 10-fold higher substitution rates compared to two commonly used chloroplast markers, have no rate heterogeneity among nucleotide positions, evolve in a clock-like fashion, and show no evidence of saturation. These advantages are offset by the short length of the flanking regions, resulting in similar numbers of parsimony informative characters to the chloroplast markers. The neutral evolution and high variability of flanking regions, together with the wide availability of monomorphic microsatellite loci in angiosperms, are useful qualities for species-level phylogenetics. The general methodology we present here facilitates to find phylogenetic markers in groups where microsatellites have been developed.

Molecular phylogenetics reveal multiple tertiary vicariance origins of the African rain forest trees.

BACKGROUND: Tropical rain forests are the most diverse terrestrial ecosystems on the planet. How this diversity evolved remains largely unexplained. In Africa, rain forests are situated in two geographically isolated regions: the West-Central Guineo-Congolian region and the coastal and montane regions of East Africa. These regions have strong floristic affinities with each other, suggesting a former connection via an Eocene pan-African rain forest. High levels of endemism observed in both regions have been hypothesized to be the result of either 1) a single break-up followed by a long isolation or 2) multiple fragmentation and reconnection since the Oligocene. To test these hypotheses the evolutionary history of endemic taxa within a rain forest restricted African lineage of the plant family Annonaceae was studied. Molecular phylogenies and divergence dates were estimated using a Bayesian relaxed uncorrelated molecular clock assumption accounting for both calibration and phylogenetic uncertainties. RESULTS: Our results provide strong evidence that East African endemic lineages of Annonaceae have multiple origins dated to significantly different times spanning the Oligocene and Miocene epochs. Moreover, these successive origins (c. 33, 16 and 8 million years--Myr) coincide with known periods of aridification and geological activity in Africa that would have recurrently isolated the Guineo-Congolian rain forest from the East African one. All East African taxa were found to have diversified prior to Pleistocene times. CONCLUSION: Molecular phylogenetic dating analyses of this large pan-African clade of Annonaceae unravels an interesting pattern of diversification for rain forest restricted trees co-occurring in West/Central and East African rain forests. Our results suggest that repeated reconnections between the West/Central and East African rain forest blocks allowed for biotic exchange while the break-ups induced speciation via vicariance, enhancing the levels of endemicity. These results provide an explanation for present day distribution patterns and origins of endemicity for African rain forest trees. Moreover, given the pre-Pleistocene origins of all the studied endemic East African genera and species, these results also offer important insights for setting conservation priorities in these highly diversified but threatened ecosystems.

Diversification of myco-heterotrophic angiosperms: evidence from Burmanniaceae.

BACKGROUND: Myco-heterotrophy evolved independently several times during angiosperm evolution. Although many species of myco-heterotrophic plants are highly endemic and long-distance dispersal seems unlikely, some genera are widely dispersed and have pantropical distributions, often with large disjunctions. Traditionally this has been interpreted as evidence for an old age of these taxa. However, due to their scarcity and highly reduced plastid genomes our understanding about the evolutionary histories of the angiosperm myco-heterotrophic groups is poor. RESULTS: We provide a hypothesis for the diversification of the myco-heterotrophic family Burmanniaceae. Phylogenetic inference, combined with biogeographical analyses, molecular divergence time estimates, and diversification analyses suggest that Burmanniaceae originated in West Gondwana and started to diversify during the Late Cretaceous. Diversification and migration of the species-rich pantropical genera Burmannia and Gymnosiphon display congruent patterns. Diversification began during the Eocene, when global temperatures peaked and tropical forests occurred at low latitudes. Simultaneous migration from the New to the Old World in Burmannia and Gymnosiphon occurred via boreotropical migration routes. Subsequent Oligocene cooling and breakup of boreotropical flora ended New-Old World migration and caused a gradual decrease in diversification rate in Burmanniaceae. CONCLUSION: Our results indicate that extant diversity and pantropical distribution of myco-heterotrophic Burmanniaceae is the result of diversification and boreotropical migration during the Eocene when tropical rain forest expanded dramatically.