Visualisation of calcium oxalate crystal macropatterns in plant leaves using an improved fast preparation method.

Leaves of the majority of plants contain calcium oxalate (CaOx) crystals or druses which often occur in spectacular distribution patterns. Numerous studies on CaOx in plant tissues across many different plant groups have been published, since it can be visualised readily under a light microscope (LM). However, there is surprisingly limited knowledge on the actual, precise distribution of CaOx in the leaves of quite ordinary plants such as common native and exotic trees. Traditional sample preparation for the documentation of the distribution of CaOx crystals in a given sample - including overall distribution - requires time-consuming clearing procedures. Here we present a refined fast preparation method to visualise the overall CaOx complement in a sample: The plant material is ashed and the ash viewed under the polarising microscope. This is a rapid method which overcomes many shortcomings of other methods and permits the visualisation of the entire CaOx content in most leaf samples. Pros and cons in comparison with the conventional clearing technique are discussed. Further aspects for CaOx investigations by micro-CT and scanning electron microscopy are discussed.

Plant migration under long-lasting hyperaridity - phylogenomics unravels recent biogeographic history in one of the oldest deserts on Earth.

The post-Miocene climatic histories of arid environments have been identified as key drivers of dispersal and diversification. Here, we investigate how climatic history correlates with the historical biogeography of the Atacama Desert genus Cristaria (Malvaceae). We analyze phylogenetic relationships and historical biogeography by using next-generation sequencing (NGS), molecular clock dating, Dispersal Extinction Cladogenesis and Bayesian sampling approaches. We employ a novel way to identify biogeographically meaningful regions as well as a rarely utilized program permitting the use of dozens of ancestral areas. Partial incongruence between the established taxonomy and our phylogenetic data argue for a complex historical biogeography with repeated introgression and incomplete lineage sorting. Cristaria originated in the central southern part of the Atacama Desert, from there the genus colonized other areas from the late Miocene onwards. The more recently diverged lineages appear to have colonized different habitats in the Atacama Desert during pluvial phases of the Pliocene and early Pleistocene. We show that NGS combined with near-comprehensive sampling can provide an unprecedented degree of phylogenetic resolution and help to correlate the historical biogeography of plant communities with cycles of arid and pluvial phases.

Asterid Phylogenomics/Phylotranscriptomics Uncover Morphological Evolutionary Histories and Support Phylogenetic Placement for Numerous Whole-Genome Duplications.

Asterids are one of the most successful angiosperm lineages, exhibiting extensive morphological diversity and including a number of important crops. Despite their biological prominence and value to humans, the deep asterid phylogeny has not been fully resolved, and the evolutionary landscape underlying their radiation remains unknown. To resolve the asterid phylogeny, we sequenced 213 transcriptomes/genomes and combined them with other data sets, representing all accepted orders and nearly all families of asterids. We show fully supported monophyly of asterids, Berberidopsidales as sister to asterids, monophyly of all orders except Icacinales, Aquifoliales, and Bruniales, and monophyly of all families except Icacinaceae and Ehretiaceae. Novel taxon placements benefited from the expanded sampling with living collections from botanical gardens, resolving hitherto uncertain relationships. The remaining ambiguous placements here are likely due to limited sampling and could be addressed in the future with relevant additional taxa. Using our well-resolved phylogeny as reference, divergence time estimates support an Aptian (Early Cretaceous) origin of asterids and the origin of all orders before the Cretaceous-Paleogene boundary. Ancestral state reconstruction at the family level suggests that the asterid ancestor was a woody terrestrial plant with simple leaves, bisexual, and actinomorphic flowers with free petals and free anthers, a superior ovary with a style, and drupaceous fruits. Whole-genome duplication (WGD) analyses provide strong evidence for 33 WGDs in asterids and one in Berberidopsidales, including four suprafamilial and seven familial/subfamilial WGDs. Our results advance the understanding of asterid phylogeny and provide numerous novel evolutionary insights into their diversification and morphological evolution.

Fruit evolution in Hydrophyllaceae.

PREMISE: Fruit type and morphology are tightly connected with angiosperm diversification. In Boraginales, the first-branching families, including Hydrophyllaceae, have one- to many-seeded capsules, whereas most of the remaining families have four-seeded indehiscent fruits. This fact argues for many-seeded capsules as the ancestral condition. However, little is known about the evolution of fruit dehiscence and seed number. The present study investigated the gynoecium and fruit development and morphology and the evolution of seed-numbers in Hydrophyllaceae. METHODS: Gynoecium and fruit development and morphology were studied using scanning electron microscopy and x-ray microcomputed tomography. Ancestral character state reconstruction of seed number was performed using a broadly sampled phylogeny of Boraginales (ndhF and ITS) with an emphasis on Hydrophyllaceae. RESULTS: Our ontogenetic studies not only demonstrate parallel developmental trajectories across Hydrophyllaceae, but also a striking diversity regarding the internal organization of the gynoecium. Ovule number appears to determine ovary structure. Many-seeded capsules are retrieved as the ancestral state of Hydrophyllaceae. At least seven transitions to fruits with (one to) four seeds and four reversals (i.e., from four- to many-seeded fruits) were reconstructed in Hydrophyllaceae. CONCLUSIONS: Several shifts in seed number from "many" to "four" and back to "many" have taken place in capsular-fruited Hydrophyllaceae, a strikingly high number considering that seed number is virtually conserved across the rest of the order. The groups with a conserved seed number of four are characterized by indehiscent schizocarps or drupes and by seeds that are integrated into mericarps. This functional integration probably acts as an evolutionary constraint to shifts in seed number.

Quaternary diversification of a columnar cactus in the driest place on earth.

PREMISE: The cactus family (Cactaceae) is a speciose lineage with an almost entirely New World distribution. The genus Eulychnia with eight currently recognized species is endemic to the Atacama and Peruvian Deserts. Here we investigated the phylogeny of this group based on a complete taxon sampling to elucidate species delimitation and biogeographic history of the genus. METHODS: A family-wide Bayesian molecular clock dating based on plastid sequence data was conducted to estimate the age of Eulychnia and its divergence from its sister genus Austrocactus. A second data set obtained from genotyping by sequencing (GBS) was analyzed, using the family-wide age estimate as a secondary calibration to date the GBS phylogeny using a penalized likelihood approach. Ancestral ranges were inferred employing the dispersal extinction cladogenesis approach. RESULTS: Our GBS phylogeny of Eulychnia was fully resolved with high support values nearly throughout the phylogeny. The split from Austrocactus occurred in the late Miocene, and Eulychnia diversified during the early Quaternary. Three lineages were retrieved: Eulychnia ritteri from Peru is sister to all Chilean species, which in turn fall into two sister clades of three and four species, respectively. Diversification in the Chilean clades started in the early Pleistocene. Eulychnia likely originated at the coastal range of its distribution and colonized inland locations several times. CONCLUSIONS: Diversification of Eulychnia during the Pleistocene coincides with long periods of hyperaridity alternated with pluvial phases. Hyperaridity caused habitat fragmentation, ultimately leading to speciation and resulting in the current allopatric distribution of taxa.

Major lineages of Loasaceae subfam. Loasoideae diversified during the Andean uplift.

The Loasoideae is the largest clade in the Loasaceae. This subfamily is widespread throughout the Neotropics and centered in the Andes, presenting an excellent opportunity to study diversification across much of temperate and mid to high-elevation areas of South America. Despite that, no studies have addressed the historical biogeography of the Loasoideae to date, leaving an important knowledge gap in this plant group. Here, we used four plastid markers (i.e., trnL-trnF, matK, trnS-trnG, and rps16) and sequenced 170 accessions (134 ingroup taxa) to infer the phylogeny of Loasoideae. We then used this phylogeny as basis to estimate divergence times using an uncorrelated relaxed molecular clock approach and seven fossils as primary calibration points. We employed the Dispersal-Extinction-Cladogenesis (DEC) approach to reconstruct the ancestral ranges of the subfamily. Our results indicate that stem Loasoideae diverged from its sister group in the Late Cretaceous to Early Paleocene (ca. 83-62 Ma). The crown node of the whole clade goes back to the Middle Paleocene to Middle Eocene (ca. 60-45 Ma), corresponding to the earliest diversification events of the extant groups, prior to most of the Andean orogeny and roughly coinciding with the Paleocene-Eocene Thermal Maximum. On the other hand, the crown nodes of most genera appear to have originated in the Oligocene and Miocene (median ages: 28-10 Ma). The diversification of some extant lineages appears to have happened in parallel to Andean uplift pulses that seem to have had an effect on the orogeny and concomitant establishment of new habitats and latitudinal corridors. The most likely ancestral areas retrieved for crown Loasoideae, are the tropical Andes and Pacific arid coast. Most of the extant clades have remained restricted to their ancestral areas. Transoceanic Long Distance Dispersal appears to have been involved in the arrival of Loasoid ancestors to South America, and in the distribution of the small clades Kissenia in Africa and Plakothira on the Marquesas Archipelago. The results presented here suggest that the historical biogeography of the continental scale radiation of Loasoideae, follows the sequence and timing of the development of temperate and mid to high-elevation habitats across South America during the Tertiary.

Phylogeny and historical biogeography of Lithospermeae (Boraginaceae): Disentangling the possible causes of Miocene diversifications.

Studies about the drivers of angiosperm clade diversifications have revealed how the environment continuously alters the species chances to adapt or to go extinct. This process depends on complex interactions between abiotic and biotic factors, conditioned to the geological and tectonic settings, the genetic variability of species and the rate at which speciation occurs. In this study, we aim to elucidate the timing of diversification of the Lithospermeae, the second largest tribe within Boraginaceae, and to identify the possible morphological and ecological characters associated with shifts in diversification rates of the most species-rich clades. Lithospermeae includes ca. 470 species and 26 genera, among which are some of the largest genera of the family such as Onosma (150 spp.), Echium (60 spp.), and Lithospermum (80 spp.). An exhaustive study of the whole clade is not available to date and its evolutionary history and diversification rates are incompletely known. In the present study, we provide the most comprehensive phylogeny of the group so far, sampling 242 species and all 26 genera. We found that crown-groups and diversification rates of Lithospermeae largely date back to the Mid-Miocene, with high diversification rates in the largest genera, though only significantly high in Onosma. Our analysis fails to associate any of the functional or morphological traits considered with significant shifts in diversification rates. The timing of the diversification of the species-rich clades corresponds with Miocene tectonic events and global climate changes increasing aridity across Eurasia and western North America. These results suggest a causal link between known ecological features of Lithospermeae (i.e., pre-adaptation to arid, open habitats, and mineral soils) and their diversification. Future studies should expand the sampling of individual subclades and detailed functional analyses to identify the contribution of adaptations to arid conditions and pollinator shifts.

Calcium phosphate in plant trichomes: the overlooked biomineral.

MAIN CONCLUSION: Calcium phosphate was unknown as a plant biomineral until recently reported in Neotropical Loasaceae. Here, we demonstrate its widespread occurrence in the trichomes of several plant families, including Brassicaceae. Calcium phosphate is the primary biomineral in, e.g., the bones and teeth of higher animals; in plants, it was only recently discovered in the stinging hairs and scabrid-glochidiate trichomes of South American Loasaceae (Ensikat et al. in Sci Rep UK 6:26073, 2016), where it appears to be deposited highly specifically, often replacing the common plant biomineral silica. We initiated a broader survey in a range of different plant orders to investigate a possibly wider distribution of calcium phosphate biomineralization in plants. Scanning electron microscopy with EDX element analysis and mapping was used for the detection of the biominerals: calcium phosphate, calcium carbonate, and silica in the trichomes of several common plant species of different orders. Results were authenticated with Raman spectroscopy. Calcium phosphate was found in the trichomes of several species in the orders Malpighiales, Rosales, Boraginales, and Brassicales. It occurred in trichome tips, replacing the more common silica, or together with silica and calcium carbonate at specific locations in the trichome cell walls. Most surprisingly, it was found in the trichomes of Arabidopsis thaliana, one of the most studied plant species-where it had been overlooked so far. The wide distribution of calcium phosphate as plant biomineral here demonstrated and the striking mineralization patterns with three different biominerals in the walls of single-celled trichomes underscore an unexpected complexity in plant biomineralization.

Mineralized trichomes in Boraginales: complex microscale heterogeneity and simple phylogenetic patterns.

Background and Aims: Boraginales are often characterized by a dense cover of stiff, mineralized trichomes, which may act as a first line of defence against herbivores. Recent studies have demonstrated that the widely reported silica and calcium carbonate in plant trichomes may be replaced by calcium phosphate. The present study investigates mineralization patterns in 42 species from nine families of the order Boraginales to investigate detailed patterns of mineralization and the possible presence of a phylogenetic signal in different mineralization patterns. Methods: The distribution of biominerals was analysed by scanning electron microscopy (SEM) including cryo-SEM and energy-dispersive X-ray analyses with element mapping. The observed distribution of biominerals was plotted onto a published phylogeny of the Boraginales. Three colours were selected to represent the principal elements: Si (red), Ca (green) and P (blue). Key Results: Calcium carbonate was present in the mineralized trichomes of all 42 species investigated, silica in 30 and calcium phosphate in 25; multiple mineralization with calcium carbonate and silica or calcium phosphate was found in all species, and 13 of the species were mineralized with all three biominerals. Trichome tips featured the most regular pattern - nearly all were exclusively mineralized with either silica or calcium phosphate. Biomineralization of the trichome shafts and bases was found to be more variable between species. However, the trichome bases were also frequently mineralized with calcium phosphate or silica, indicating that not only the tip is under functional constraints requiring specific patterns of chemical heterogeneity. The complete absence of either silica or phosphate may be an additional feature with systematic relevance. Conclusions: This study demonstrates that complex, site-specific and differential biomineralization is widespread across the order Boraginales. Calcium phosphate, only recently first reported as a structural plant biomineral, is common and appears to be functionally analogous to silica. A comparison with the phylogeny of Boraginales additionally reveals striking phylogenetic patterns. Most families show characteristic patterns of biomineralization, such as the virtual absence of calcium phosphate in Cordiaceae and Boraginaceae, the triple biomineralization of Heliotropiaceae and Ehretiaceae, or the absence of silica in Namaceae and Codonaceae. The complex chemical and phylogenetic patterns indicate that trichome evolution and functionalities are anything but simple and follow complex functional and phylogenetic constraints.

The geometry of gender: hyper-diversification of sexual systems in Urtica L. (Urticaceae).

Urtica L. (Urticaceae) is generally reported as a genus of monoecious and dioecious taxa. However, the gender information found in the literature does not at all reflect the actual diversity of gender patterns in Urtica. Dioecy appears to be truly absent from Urtica, but otherwise there has been a major diversification in the geometry of gender and no comparable patterns exist in other plant groups. Thus, we here define technical terms for all unique architectural types of monoecy found in Urtica and closely related genera and reconstruct the ancestral gender states in a Bayesian framework. Our studies are based on a near-comprehensive sampling, including 61 of the 63 Urtica species recognized. We report polygamy, two types of gynodioecy and five different architectural types of monoecy. A total of 15 switches appear to have taken place within the genus. Although gender characteristics have diversified strongly, they are relatively conserved within clades. Monoecy is the predominant sexual system within Urtica and specifically basiandrous monoecy (i.e. basal inflorescence branches of each individual male only, apical branches female) is the most widespread type, reported for 11 different clades. In particular, it characterizes the basally branching pilulifera-clade and the sister genus Zhengyia, and may thus represent the plesiomorphic condition for Urtica. Gender distribution and gross morphology appear to evolve largely independently from each other and gender distribution is largely independent of growth habit. However, polygamous taxa are most common amongst rhizomatous perennials (one-third of the taxa).

Stinging hair morphology and wall biomineralization across five plant families: Conserved morphology versus divergent cell wall composition.

PREMISE OF THE STUDY: Stinging hairs are striking examples of plant microengineering-the plant equivalent of the hypodermic syringe. The requisite mechanical properties are mostly achieved by cell wall mineralization. Stinging hairs of Urtica dioica (Urticaceae) are known to be mineralized with silica and calcium carbonate and those of Loasaceae also with calcium phosphate, but no comparative study has been provided across different taxa with stinging hairs. METHODS: Light microscopy and scanning electron microscopy (SEM) with cryo-SEM and energy-dispersive x-ray spectroscopy were used to analyze morphology and biomineralization of stinging hairs of 43 species from the families Caricaceae, Euphorbiaceae, Loasaceae, Namaceae, and Urticaceae. KEY RESULTS: Stinging hair morphology is similar across the taxa studied, in striking contrast to the divergent patterns of biomineralization. Trichome bases are mostly calcified, sometimes silicified, the shafts are mostly calcified, and the apices silicified (Urticaceae), and contain calcium phosphate (Caricaceae, Namaceae), both silica and calcium phosphate (some Loasaceae), or no minerals (Cnidoscolus, Euphorbiaceae). Some stinging hairs are superficially thinly coated with silica over a cell wall otherwise mineralized with calcium carbonate or calcium phosphate. CONCLUSIONS: Mineralization patterns are surprisingly diverse and involve three different biominerals deposited in different parts of individual trichomes with calcium phosphate a common component. The physical properties of different wall regions of the stinging trichomes are thus fine-tuned to optimize their function via modulation of wall thickness and differential element deposition. Similar function is apparently achieved through divergent wall compositions.

Geraniales flowers revisited: evolutionary trends in floral nectaries.

BACKGROUND AND AIMS: The detailed relationships in Geraniales in their current circumscription have only recently been clarified. The disparate floral morphologies and especially the nectaries of the corresponding group have consequently not previously been studied in a phylogenetic context. METHODS: The present study investigates floral and especially nectary morphology and structure for representatives of 12 of the 13 currently accepted genera in the five families of the Geraniales. Flowers were studied using light microscopy and scanning electron microscopy. KEY RESULTS: The data demonstrate the derivation of even the most disparate floral morphologies from a basic pentamerous and pentacyclic organization, with an obdiplostemonous androecium and receptacular nectaries associated with the antesepalous stamens. Divergent morphologies are explained by modifications of merosity (tetramerous flowers), symmetry (several transitions to zygomorphic flowers) and elaboration of the nectaries into variously shaped outgrowths and appendages, especially in Francoaceae. The divergent development of nectar glands ultimately leads to either a reduction in their number (to one in some Geraniaceae and Melianthaceae) or their total loss (some Vivianiaceae). CONCLUSIONS: Floral morphology of the Geraniales shows a high degree of similarity, despite the variation in overall floral appearance and nectary morphology. A hypothesis on the transformation of the nectaries within the Geraniales is presented.

The relationship between nectaries and floral architecture: a case study in Geraniaceae and Hypseocharitaceae.

Background and Aims: Flowers of Geraniaceae and Hypseocharitaceae are generally considered as morphologically simple. However, previous studies indicated complex diversity in floral architecture including tendencies towards synorganization. Most of the species have nectar-rewarding flowers which makes the nectaries a key component of floral organization and architecture. Here, the development of the floral nectaries is studied and placed into the context of floral architecture. Methods: Seven species from Geraniaceae and one from Hypseocharitaceae were investigated using scanning electron microscopy and light microscopy. Samples were prepared and processed using standard protocols. Key Results: The development of the nectary glands follows the same trajectory in all species studied. Minor differences occur in the onset of nectarostomata development. The most striking finding is the discovery that a short anthophore develops via intercalary growth at the level of the nectary glands. This anthophore lifts up the entire flower apart from the nectary gland itself and thus plays an important role in floral architecture, especially in the flowers of Pelargonium. Here, the zygomorphic flowers show a particularly extensive receptacular growth, resulting in the formation of a spur-like receptacular cavity ('inner spur'). The nectary gland is hidden at the base of the cavity. Various forms of compartmentalization, culminating in the 'revolver flower' of Geranium maderense, are described. Conclusions: Despite the superficial similarity of the flowers in Geraniaceae and Hypseocharitaceae, there is broad diversity in floral organization and floral architecture. While the receptacular origin of the spur-like cavity in Pelargonium had already been described, anthophore formation via intercalary growth of the receptacle in the other genera had not been previously documented. In the context of the most recent phylogenies of the families, an evolutionary series for the floral architecture is proposed, underscoring the importance of synorganization in these seemingly simple flowers.

Complex patterns of multiple biomineralization in single-celled plant trichomes of the Loasaceae.

PREMISE OF THE STUDY: Plants of the family Loasaceae are characterized by a usually dense indument of various trichome types, including two basically different types of mineralized, unicellular trichomes (stinging hairs or setae and scabrid-glochidiate trichomes). Mineralized trichomes have long been known to have silicified or calcified walls, but recent studies demonstrated that trichomes of Loasaceae may also contain calcium phosphate. The current study investigates the distribution of different biominerals in the mineralized trichomes across several different taxa. METHODS: Plants from cultivation were studied with scanning electron microscopy including energy dispersive x-ray analyses and element mapping. KEY RESULTS: The vast majority of the 31 species investigated had at least two different biominerals in their trichomes, and 22 had three different biominerals in their trichomes. Thirty of the species had calcium phosphate in their trichomes. Loasa was mostly free of silica, but contained calcium phosphate in trichome tips and barbs, whereas calcium phosphate and silica were found in representatives of other genera of the family (Blumenbachia, Caiophora, Nasa). CONCLUSIONS: Biomineralization is remarkably diversified between species, different trichome types and parts of the same trichome. Individual genera largely had different patterns of biomineralization. The presence of three biominerals in the trichomes of the basally branching Eucnide urens indicates either an early evolution and subsequent loss or several independent origins of multiple biomineralization. Differential biomineralization of the parts of individual, unicellular trichomes clearly indicates an extraordinary degree of physiological control over this process.

Ontogeny and the process of biomineralization in the trichomes of Loasaceae.

PREMISE OF THE STUDY: South American Loasaceae have a morphologically complex trichome cover, which is characterized by multiple biomineralization. The current study investigates the ontogeny of these complex trichomes and the process of their biomineralization, since both are very poorly understood. METHODS: The development of stinging trichomes on various parts of the plants and the process of mineralization were studied using scanning electron microscopy (SEM) and energy-dispersive x-ray spectroscopy (EDX). KEY RESULTS: Trichomes are initiated very early in organ development and the different trichome types begin developing their distinctive morphology at a very early developmental stage. Biomineralization in the stinging trichomes starts with the deposition of silica or calcium phosphate in the apex and then proceeds basipetally, with a more irregular, subsimultaneous mineralization of the base and the shaft. Mineralization of the scabrid-glochidiate trichomes starts on the surface processes and in the apex (silica, calcium phosphate), with a subsequent mineralization of the shaft with calcium carbonate. CONCLUSION: Mineralized trichomes in Loasaceae provide an excellent model for the study of biomineralization. The overall sequence of mineralization is typically from distal and peripheral to proximal and central. Typically, three biominerals-silica, calcium carbonate, and calcium phosphate-are differentially and sequentially deposited in different parts of each unicellular stinging trichome.

Pulling the sting out of nettle systematics - A comprehensive phylogeny of the genus Urtica L. (Urticaceae).

The genus Urtica L. is subcosmopolitan, found on all continents (except Antarctica) and most extratropical islands and ranges from Alaska to Patagonia, Spitzbergen to the Cape and Camtschatka to the subantarctic islands. However, throughout its geographical range morphologically nearly indistinguishable species are found alongside morphologically quite disparate species, with the overall diversity of morphological characters extremely limited. The systematics of Urtica have puzzled scientists for the past 200years and no single comprehensive attempt at understanding infrageneric relationships has been published in the past, nor are species delimitations unequivocally established. We here provide the first comprehensive phylogeny of the genus including 61 of the 63 species recognized, represented by 144 ingroup accessions and 14 outgroup taxa. The markers ITS1-5.8S-ITS2, psbA-trnH intergenic spacer, trnL-trnF and trnS-trnG are used. The phylogeny is well resolved. The eastern Asian Zhengyia shennongensis T. Deng, D.G. Zhang & H. Sun is retrieved as sister to Urtica. Within Urtica, a clade comprising the western Eurasian species U. pilulifera L. and U. neubaueri Chrtek is sister to all other species of the genus. The phylogenetic analyses retrieve numerous well-supported clades, suggesting previously unsuspected relationships and implying that classically used taxonomic characters such as leaf morphology and growth habit are highly homoplasious. Species delimitation is problematical, and several accessions assigned to Urtica dioica L. (as subspecies) are retrieved in widely different places in the phylogeny. The genus seems to have undergone numerous dispersal-establishment events both between continents and onto different islands. Three recent species radiations are inferred, one in America centered in the Andes, one in New Zealand, and one in northern Eurasia which includes Urtica dioica s.str. sensu Henning et al. (2014). The present study provides the basis of a critical re-examination of species limits and taxonomy, but also of the dispersal ecology of this widespread plant group and an in-depth study of the three clades with recent radiations.

Fossil nutlets of Boraginaceae from the continental Eocene of Hamada of Méridja (southwestern Algeria): The first fossil of the Borage family in Africa.

PREMISE OF THE STUDY: The Paleogene deposits of the Hamada of Méridja, southwestern Algeria, are currently dated as lower-to-middle Eocene in age based on fossil gastropods and charophytes. Here we report the presence of fruits that can be assigned to the Boraginaceae s.str., apparently representing the first fossil record for this family in Africa, shedding new light on the historical biogeography of this group. METHODS: Microscopic studies of the fossil nutlets were carried out and compared to extant Boraginaceae nutlets, and to types reported in the literature for this family. KEY RESULTS: The fossils are strikingly similar in general size and morphology, particularly in the finer details of the attachment scar and ornamentation, to nutlets of extant representatives of the Boraginaceae tribe Echiochileae, and especially the genus Ogastemma. We believe that these nutlets represent an extinct member of this lineage. CONCLUSIONS: The Ogastemma-like fossils indicate that the Echiochileae, which are most diverse in northern Africa and southwestern Asia, have a long history in this region, dating back to the Eocene. This tribe corresponds to the basal-most clade in Boraginaceae s.str., and the fossils described here agree well with an assumed African origin of the family and the Boraginales I, providing an important additional calibration point for dating the phylogenies of this clade.

Nectar sugars and bird visitation define a floral niche for basidiomycetous yeast on the Canary Islands.

BACKGROUND: Studies on the diversity of yeasts in floral nectar were first carried out in the late 19th century. A narrow group of fermenting, osmophilous ascomycetes were regarded as exclusive specialists able to populate this unique and species poor environment. More recently, it became apparent that microorganisms might play an important role in the process of plant pollination. Despite the importance of these nectar dwelling yeasts, knowledge of the factors that drive their diversity and species composition is scarce. RESULTS: In this study, we linked the frequencies of yeast species in floral nectars from various host plants on the Canary Islands to nectar traits and flower visitors. We estimated the structuring impact of pollination syndromes (nectar volume, sugar concentration and sugar composition) on yeast diversity.The observed total yeast diversity was consistent with former studies, however, the present survey yielded additional basidiomycetous yeasts in unexpectedly high numbers. Our results show these basidiomycetes are significantly associated with ornithophilous flowers. Specialized ascomycetes inhabit sucrose-dominant nectars, but are surprisingly rare in nectar dominated by monosaccharides. CONCLUSIONS: There are two conclusions from this study: (i) a shift of floral visitors towards ornithophily alters the likelihood of yeast inoculation in flowers, and (ii) low concentrated hexose-dominant nectar promotes colonization of flowers by basidiomycetes. In the studied floral system, basidiomycete yeasts are acknowledged as regular members of nectar. This challenges the current understanding that nectar is an ecological niche solely occupied by ascomycetous yeasts.