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Sex, drugs and pupusas: Disentangling relationships in Echiteae (Apocynaceae)

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Echiteae (Apocynaceae) comprises about 200 species divided among 19 genera, 16 of which are restricted to the Neotropics, the other 3 to the Paleotropics. There are two large genera, Prestonia and Parsonsia, in the Neotropics and Paleotropics, respectively, whereas three-fourths of the genera contain only one to four species. In this study DNA of 82 species was extracted and amplified for four molecular markers: three plastid (trnL intron + trnL-trnF intergenic spacer, rpl16 intron, matK + 5′/ 3′ trnK intron) and the nuclear ribosomal region ITS. The ingroup comprised 70 species from 17 genera in the tribe, and included representatives of all subtribes; the outgroup included 5 species each from the two putatively most closely related tribes, Odontadenieae and Mesechiteae, as well as 2 rooting species. Phylogenetic analyses were conducted using Bayesian inference and maximum likelihood approaches. Echiteae and four of its subtribes (Echitinae, Parsonsiinae, Peltastinae, Prestoniinae) as currently circumscribed were shown to be non-monophyletic. The fifth subtribe, Pentalinoninae, though monophyletic, does not belong to Echiteae; it was resolved as sister to a clade of Odontadenieae, where it fits better, since in both the characteristic secondary compounds are cardenolides. Prestonia and Temnadenia were resolved as polyphyletic, with P. riedelii grouped with Rhodocalyx and T. ornata nested within Prestonia. Fernaldia is nested within Echites and Peltastes is nested in Macropharynx. Fourteen genera are maintained. The new subtribe Laubertiinae is proposed, as are the combinations resulting from the merging of Fernaldia and Peltastes and the transfer of P. riedelii and T. ornata to Rhodocalyx and Prestonia, respectively. In the ancestors of Echiteae, we hypothesize that an evolutionary shift took place in which steroidal alkaloids and/or cardenolides, characteristic for apocynoids, were replaced by parsonsine-type pyrrolizidine alkaloids as the predominant defense compounds.
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... Description of the family Apocynaceae and the studied species of Rauvolfia with their medicinal importance Apocynaceae Juss. is positioned within Gentianales (asterids, lamiids) and includes about 400 genera and 5000 species located in tropical and subtropical regions around the world (Stevens 2001 onwards;Endress 2004;APG IV 2016). The family is currently divided into five subfamilies: Rauvolfioideae (cosmopolitan; 10 tribes/83 genera; 915 species), Apocynoideae (cosmopolitan; eight tribes/80 genera; 822 species), Periplocoideae (Old World; 33 genera), Secamonoideae (Old World; eight genera) and Asclepiadoideae (cosmopolitan; four tribes/172 genera) (Endress and Bruyns 2000;Middleton 2007;Endress et al. 2014;Morales et al. 2017). The Apocynaceae have been the subject of several taxonomic, evolutionary and phylogenetic analyses (Endress and Stevens 2001;Endress et al. 2007Endress et al. , 2014Ionta and Judd 2007;Simões et al. 2007;Wyatt and Lipow 2007;Rapini 2012;Morales et al. 2017;Fishbein et al. 2018), many of which have included descriptions of pollen morphological characteristics (Nilsson 1990;Nilsson et al. 1993;Verhoeven and Venter 1998;Furness 2007;Van Der Ham et al. 2001; Van Der Weide and Van der Ham 2012). ...
... The family is currently divided into five subfamilies: Rauvolfioideae (cosmopolitan; 10 tribes/83 genera; 915 species), Apocynoideae (cosmopolitan; eight tribes/80 genera; 822 species), Periplocoideae (Old World; 33 genera), Secamonoideae (Old World; eight genera) and Asclepiadoideae (cosmopolitan; four tribes/172 genera) (Endress and Bruyns 2000;Middleton 2007;Endress et al. 2014;Morales et al. 2017). The Apocynaceae have been the subject of several taxonomic, evolutionary and phylogenetic analyses (Endress and Stevens 2001;Endress et al. 2007Endress et al. , 2014Ionta and Judd 2007;Simões et al. 2007;Wyatt and Lipow 2007;Rapini 2012;Morales et al. 2017;Fishbein et al. 2018), many of which have included descriptions of pollen morphological characteristics (Nilsson 1990;Nilsson et al. 1993;Verhoeven and Venter 1998;Furness 2007;Van Der Ham et al. 2001; Van Der Weide and Van der Ham 2012). Rauvolfia (Rauvolfioideae: Apocynaceae) is a genus of evergreen trees and shrubs. ...
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Rauvolfia belongs to the family Apocynaceae and encompasses herbs or shrubs with leaves in whorls of three or four. It is an endangered plant of the tropics and subtropics. We present a preliminary study and analysis of the morphological details of the pollen of two extant species of Rauvolfia (R. serpentina (L.) Benth. ex Kurz and R. tetraphylla L.) from the Ganga Plain using light microscopy (LM), confocal laser scanning microscopy (CLSM) and field emission scanning electron microscopy (FESEM). The critical point drying method (CPD) was adopted to test the pollen size difference from the conventional acetolysis method (ACE). The pollen morphology of R. tetraphylla differs from that of R. serpentina in several specific traits. Its pollen grain is 3-colporate; it is oblate to oblate-spheroidal; sexine is punctate to obscure and mostly as thick as nexine; and a distinct thickening is present around the ectocolpi. The study shows that R. serpentina can be distinguished from R. tetraphylla by its pollen shape, size, sexine ornamentation (particularly the presence and absence of punctae/perforations in the mesocolpial region) and length of the ectocolpi thickening. The pollen shape and aperture number are more or less common features in Rauvolfia spp., but the presence and absence of a punctate pattern at the mesocolpial position marks the primary difference between the two species. The t-test was applied to determine the statistical significance of pollen morphological data of both species. This study provides a source of information for systematic and conservation purposes and provides a baseline to facilitate palynological studies of past vegetation and palaeoenvironments.
... In some subfamilies of the Apocynaceae, including those in which the study species occur (Echiteae [Echites P. Browne] and Odontadenieae [Angadenia Miers and Pentalinon Voigt]; Morales et al., 2017), the terminal portion of the style head is non-receptive, functioning only to bear pollen in what can be called the pollen chamber; the middle area of the style head is a secretory area; and the receptive stigmatic area is at the base of the style head (Fig. 1). Pollen is deposited by the mouthparts of visitors searching for nectar in the flower. ...
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Premise: Flowers of the Apocynaceae (milkweed family) have complex structures and pollination mechanisms. Pollen removal and deposition in Angadenia, Pentalinon, and Echites are similar, with anthers releasing pollen onto the sterile style head. The mid-style head excretes a glue that coats the mouthparts of vistors to aid in the transfer of pollen. Subsequent probes may deposit pollen on the receptive stigmatic surface on the lowest part of the style head, with fertilization resulting after pollination by compatible pollen. Methods: By employing fishing line of different diameters, which reflected the diameters of the mouthparts of the different insect visitors, we determined the widths best able to remove and deposit pollen, thereby revealing which of the visitors could be effective pollinators, and which may be only nectar robbers. Results: We previously found that mouthpart (proboscis) width is correlated with pollen transfer effectiveness in Angadenia berteroi and confirmed this here in two other species, Pentalinon luteum and Echites umbellatus. Our data allowed the prediction of the most effective pollinators of these two other species. Discussion: The simulation of flower visitor mouthparts using fishing line can provide useful data for evaluating the potential for effective pollen removal and deposition by different visitors.
... The five major taxonomic divisions of Apocynaceae recognized here follow the most recent classifications; former subfamilies Rauvolfioideae and Apocynoideae have repeatedly been shown to be paraphyletic (Livshultz et al., 2007;Straub et al., 2014;Fishbein et al., 2018) and are here recognized informally as Rauvolfioids and Apocynoids, respectively, following Simões et al. (2016), Morales et al. (2017) andFishbein et al. (2018). Apocynoids + Periplocoideae + Secamonoideae + Asclepiadoideae (known as the APSA clade - Livshultz et al., 2007) is monophyletic, and apart from a few exceptions, shares a number of reproductive morphological features that demarcates the group from Rauvolfioids. ...
... The five major taxonomic divisions of Apocynaceae recognized here follow the most recent classifications; former subfamilies Rauvolfioideae and Apocynoideae have repeatedly been shown to be paraphyletic (Livshultz et al., 2007;Straub et al., 2014;Fishbein et al., 2018) and are here recognized informally as Rauvolfioids and Apocynoids, respectively, following Simões et al. (2016), Morales et al. (2017) andFishbein et al. (2018). Apocynoids + Periplocoideae + Secamonoideae + Asclepiadoideae (known as the APSA clade - Livshultz et al., 2007) is monophyletic, and apart from a few exceptions, shares a number of reproductive morphological features that demarcates the group from Rauvolfioids. ...
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Background and Aims Large clades of angiosperms are often characterized by diverse interactions with pol-linators, but how these pollination systems are structured phylogenetically and biogeographically is still uncertain for most families. Apocynaceae is a clade of >5300 species with a worldwide distribution. A database representing >10 % of species in the family was used to explore the diversity of pollinators and evolutionary shifts in pollination systems across major clades and regions. • Methods The database was compiled from published and unpublished reports. Plants were categorized into broad pollination systems and then subdivided to include bimodal systems. These were mapped against the five major divisions of the family, and against the smaller clades. Finally, pollination systems were mapped onto a phylogenetic reconstruction that included those species for which sequence data are available, and transition rates between pollination systems were calculated. • Key Results Most Apocynaceae are insect pollinated with few records of bird pollination. Almost three-quarters of species are pollinated by a single higher taxon (e.g. flies or moths); 7 % have bimodal pollination systems, whilst the remaining approx. 20 % are insect generalists. The less phenotypically specialized flowers of the Rauvolfioids are pollinated by a more restricted set of pollinators than are more complex flowers within the Apocynoids + Periplocoideae + Secamonoideae + Asclepiadoideae (APSA) clade. Certain combinations of bimodal pollination systems are more common than others. Some pollination systems are missing from particular regions, whilst others are over-represented. • Conclusions Within Apocynaceae, interactions with pollinators are highly structured both phylogenetically and biogeographically. Variation in transition rates between pollination systems suggest constraints on their evolution, whereas regional differences point to environmental effects such as filtering of certain pollinators from habitats. This is the most extensive analysis of its type so far attempted and gives important insights into the diversity and evolution of pollination systems in large clades.
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Vincetoxicum emeiense (Apocynaceae), a new species from Emeishan City, Sichuan Province, China, is described and illustrated. It is morphologically similar to V. hui and V. koi but can be distinguished by the glabrous stem, marginally finely ciliate with adaxially puberulent leaves and orange to purple corolla. Color photographs, conservation assessment and other relevant notes are also provided.
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Apocynaceae are well known for diverse specialized metabolites that are distributed in a phylogenetically informative manner. Pyrrolizidine alkaloids (PAs) have been reported sporadically in one lineage in the family, the APSA clade, but few species have been studied to date. We conducted the first systematic survey of Apocynaceae for retronecine-type PAs, sampling leaves from 231 species from 13 of 16 major lineages within the APSA clade using HPLC-MS/MS. We also followed up preliminary evidence for infra-specific variation of PA detectability in Echites umbellatus Jacq. Four precursor ion scans (PREC) were developed for a high-throughput survey for chemicals containing a structural moiety common to many PAs, the retronecine core. We identified with high confidence PAs in 7 of 8 sampled genera of tribe Echiteae, but not in samples from the closely related Odontadenieae and Mesechiteae, confirming the utility of PAs as a taxonomic character in tribal delimitation. Occurrence of PAs in Malouetieae is reported with moderate confidence in Galactophora schomburgkiana Woodson and Eucorymbia alba Stapf, but currently we have low confidence of their presence in Holarrena pubescens Wall. ex G. Don (the one Malouetieae species where they were previously reported), as well as in Holarrena curtisii King & Gamble and in Kibatalia macrophylla (Pierre ex Hua) Woodson. Candidate PAs in some species of Wrightia R. Br. (Wrightieae) and Marsdenia R. Br. (Marsdenieae) are proposed with moderate confidence, but a subset of the compounds in these taxa presenting with a PA-like fragmentation pattern are more likely to be aminobenzoyl glycosides. Candidate PAs are reported in species with predicted (VXXXD) and unexpected (IXXXN) amino acid motifs in their homospermidine synthase-like genes. Detectability of PAs varies among samples of Echites umbellatus and intra-individual plasticity contributes to this variation. Of toxicological importance, novel potential sources of human exposure to pro-toxic PAs were identified in the medicinal plant, Wrightia tinctoria R.Br., and the food plants, Marsdenia glabra Cost. and Echites panduratus A. DC., warranting immediate further research to elucidate the structures of the candidate PAs identified. Method development and limitations are discussed.
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Apocynaceae are well-known for diverse specialized metabolites that are distributed in a phylogenetically informative manner. Pyrrolizidine alkaloids (PAs) have been reported sporadically in one lineage in the family, the APSA clade, but few species had been studied to date. We conduct the first systematic survey of Apocynaceae for retronecine-type PAs, sampling leaves from 231 species from 13 of 16 major lineages within the APSA clade using HPLC-MS/MS. We also follow up on preliminary evidence for infra-specific variation of PA detectability in Echites umbellatus Jacq. Four precursor ion scans (PREC) were developed for a high-throughput survey for chemicals containing a structural moiety common to many PAs, the retronecine core. We identified with high confidence PAs in 7 of 8 sampled genera of tribe Echiteae, but not in samples from the closely related Odontadenieae and Mesechiteae, confirming the utility of PAs as a taxonomic character in tribal delimitation. The presence of PAs in Malouetieae was confirmed, as we report with high confidence their presence in Galactophora schomburgkiana Woodson and Eucorymbia alba Stapf, but currently we have low confidence of their presence in Holarrena pubescens Wall. ex G. Don (the one Malouetieae species where they were previously reported), as well as in Kibatalia macrophylla (Pierre ex Hua) Woodson and in Holarrena curtisii King & Gamble. For the first time the presence of PAs in species of Wrightia R. Br. (Wrightieae) and Marsdenia R. Br. (Marsdenieae) was confirmed. Detectability of PAs was found to vary among samples of Echites umbellatus and intra-individual plasticity contributes to this variation. Of toxicological importance, novel potential sources of human exposure to pro-toxic PAs were identified in the medicinal plants, Wrightia tinctoria R.Br. and Marsdenia tinctoria R.Br., and the food plant, Echites panduratus A. DC., warranting immediate further research to elucidate the structures of the candidate PAs identified. Method development and limitations are discussed.
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Antecedentes: La familia Apocynaceae está entre las 13 más diversas de México y su conocimiento taxonómico ha tenido una atención creciente con la implementación de filogenias y otras herramientas para resolver su sistemática. Con más de 10 años del último listado enfocado en la familia, resulta necesario actualizar el conocimiento sistemático del grupo. Pregunta: ¿Cómo se ha consolidado hasta nuestros días el conocimiento sistemático del grupo en México? ¿Cómo ha cambiado el número de géneros y especies de Apocynaceae en el país y su distribución geográfica por estado? Especie de estudio: Apocynaceae Sitio de estudio: México Método: Revisión de herbarios nacionales e internacionales, bases de datos y una búsqueda intensiva de literatura. Así como colectas en diferentes partes del país. Resultados: Apocynaceae presenta 52 géneros y 418 especies nativas, 204 de ellas endémicas. La subfamilia Asclepiadoideae destaca con 27 géneros y 315 especies. Los estados con más especies y endemismos son Oaxaca, Chiapas, Veracruz y Guerrero. Los estados con más microendemismos son Oaxaca (17 spp.), Chiapas y Jalisco (7 spp.), Baja California Sur y Veracruz (5 spp.). Se proporciona una clave de géneros y un listado actualizado. Conclusiones: Apocynaceae se ubica en el lugar número 12 de las familias más diversas del país. Esto reitera al país como un centro de diversidad del grupo. La integración de más estudios relacionados a la aplicación de herramientas filogenéticas, modelos de distribución y redes de interacción biológica permitirá entender mejor a la familia y proporcionar una historia natural más completa.
Chapter
Woody climbers, vines, perennial herbs, trees or shrubs, more rarely annuals, sometimes with large water-storing tubers or a xylopod, sometimes succulent, with large grappling hooks and/or tendrils in several lianoid genera of Willughbeieae; latex in non-articulated laticifers present, most commonly white, but in some genera usually translucent and in others yellowish or reddish. Leaves simple and usually entire, very rarely dentate or repand, usually isophyllous, but often anisophyllous in Tabernaemontaneae-Tabernaemontiinae, sometimes with distinctly different juvenile and adult foliage, normally petiolate, sometimes sessile, usually opposite, less frequently alternate or whorled (whorled phyllotaxis characteristic for a number of Rauvolfioid genera); stipules usually absent or small and caducous, sometimes enlarged and fused into dentate interpetiolar collars (a few Periplocoid genera), commonly with interpetiolar lines or ridges, sometimes the petioles of a leaf pair connate at the node, forming a short ocrea, which may be expanded into small intrapetiolar flaps clasping the stem (Tabernaemontaneae), almost always with colleters in the axil of the leaf, sometimes on the petiole, in a cluster adaxially at the juncture of petiole and lamina or along the midrib above, occasionally with abaxial domatia in the axils of the secondary veins (mainly in Apocynoids). Flowers perfect, rarely functionally dioecious, often scented, sessile or more commonly pedicellate, in solitary or more commonly in axillary, extra-axillary or terminal multi-flowered cymes, panicles or thyrses, sometimes appearing as an axillary fascicle. Perianth almost always actinomorphic, very rarely slightly zygomorphic; calyx almost always 5- (rarely 4- or 6–7-)merous, lobes normally quincuncially arranged, synsepalous or aposepalous, commonly with colleters, in Periplocoideae, Secamonoideae and Asclepiadoideae these are usually in the sinuses, but in some Rauvolfioids and several Apocynoids colleters in a continuous ring, in multiple rows in some Tabernaemontaneae and Hunterieae, or a single antesepalous colleter (especially in Echiteae), and in several genera of Rauvolfioids and Apocynoids colleters are absent; corolla sympetalous, rarely apopetalous (a few Ceropegieae), salverform, infundibuliform, tubular, urceolate or rotate, lobes almost always 5 (very rarely 4), usually contorted in bud, either dextrorse or sinistrorse, more rarely valvate; corolline or gynostegial coronas often present; stamens 5 (rarely 4), filaments mostly straight, sometimes geniculate, sometimes connate around the style (some species of Forsteronia, Thoreauea), sometimes coiled around the style (Dewevrella, some species of Parsonsia and Thenardia), inserted on the corolla tube, on prominent staminal feet (broadened filament base fused with corolla tube) or forming a staminal tube, included to exserted; anthers introrse, rarely latrorse, in almost all Apocynoids, Secamonoideae and Asclepiadoideae with highly elaborated and lignified guide rails (lignified guide rails absent in most Rauvolfioids and in Periplocoideae) and often with an apical connective appendage, thecae 4, unequal in most Apocynoids, with dorsal ones smaller through presence of guide rails, reduced to 2 in Asclepiadoideae, dehiscence longitudinal, attached to the style-head forming a gynostegium (gynostegium absent in Rauvolfioids); nectaries in alternistaminal pockets on the staminal tube, on sides of staminal feet or 5 (rarely 2) lobes encircling the base of the ovary, these often fused to varying degrees into an (often deeply lobed) ring (in some Rauvolfioids and early-branching Apocynoids nectaries are adnate to the outer wall of the ovary at the base or are sometimes nonfunctional or absent); gynoecium normally of two carpels (very rarely up to five); ovary mostly apocarpous, sometimes congenitally (Rauvolfioids only) or postgenitally syncarpous (several Apocynoids), in some genera only one carpel developing, superior to subinferior; placentation marginal when the ovary is apocarpous, parietal or axile when syncarpous, when apocarpous upper part of the carpels fusing postgenitally to form a complex style-head that produces adhesive for pollen transport, with a pollen-trapping basal collar and/or pollen-presenting upper crest present in many Rauvolfioids and Apocynoids; stigma mostly on the underside of the style-head, often restricted to five chambers behind the guide rails, but style-head scarcely morphologically differentiated and nearly uniformly receptive in some Rauvolfioids; adhesive a sticky foam or mucilage, or differentiated into five translators with a scoop-like pollen receptacle and sticky base, or as five hard clips (corpuscles) usually accompanied by five pairs of flexible arms (caudicles) forming a pollinarium. Fruit in Rauvolfioids diverse: drupes, berries, follicles or capsules; seeds usually without a coma, naked, arillate, or winged or fimbriate at the margin very rarely with a coma (Haplophyton); in the remainder of the family, fruit almost always a pair of ventrally dehiscent follicles (often only one due to abortion or due to postgenital fusion; rarely a septicidally dehiscent capsule) with small seeds with a micropylar coma, rarely with a chalazal coma, coma at both ends (only in early-branching Apocynoids), or fringed with long trichomes circumferentially (a few Periplocoid and Hoya species), or without a coma.
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Vinceae comprises 9 genera and about 153 species. The genera differ greatly in a number of morphological traits, especially those relating to fruit and seeds, which resulted in their placement in different tribes and/or subtribes by earlier taxonomists. Here, 378 new chloroplast DNA sequences from 98 species from 5 tribes of the rauvolfioid grade were analyzed phylogenetically to test the monophyly of Vinceae, its 6 subtribes, and constituent genera, as well as the sectional treatments of Rauvolfia, and to find morphological synapomorphies for major clades. Vinceae, as currently circumscribed, was found to be monophyletic as were four of the six subtribes. Kopsia and Ochrosia were confirmed as successive sister groups to the rest. All nine genera currently recognized in the tribe were resolved as monophyletic, Laxoplumeria and Tonduzia for the first time. The broad circumscription of Petchia and Ochrosia proposed by previous authors was supported. The pantropical genus Rauvolfia is divided into two major lineages: one comprised entirely of paleotropical species, and the other the neotropical species, a recurring geographic pattern in the family. Surprisingly, the Hawaiian species of Vinceae have probably arisen by two different trans-oceanic routes, either by long-distance dispersal from Australasian (Ochrosia spp.) or Caribbean (Rauvolfia sandwicensis) ancestors. Within Rauvolfia, most of the sections, series and subseries recognized in previous classifications are paraphyletic. Only 2 of the 14 sections of Rauvolfia proposed by Pichon and 1 of the 11 sections proposed by Rao that are not monospecific were found to be monophyletic. Mapping of 30 morphological characters onto the molecular tree identified an unlobed upper wreath on the style head as an unambigous synapomorphy characterizing the tribe as well as synapomorphies for each of its genera. The two earliest-branching genera, Kopsia and Ochrosia, have dextrorse corolla lobe aestivation, in contrast to the sinistrorse corollas of the other Vinceae and the great majority of genera throughout the rauvolfioid grade. Drupes have arisen in parallel at least three times within Vinceae. The adaptations of the endocarp for water dispersal are possibly ancestral in the tribe, and the observed morphological differences between the drupaceous fruits of Kopsia/Ochrosia versus Petchia and Rauvolfia may be due to their abiotic versus biotic dispersal mode, respectively.
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The nuclear gene phytochrome A (PHYA) from 71 species of crown clade Apocynaceae (subfamilies Asclepiadoideae, Secamonoideae, Periplocoideae, and four lineages of Apocynoideae) and outgroups is used to (1) test the chloroplast phylogeny that places the African tribe Baisseeae, with solitary pollen grains, as sister to the pollinia‐bearing milkweeds (Secamonoideae plus Asclepiadoideae); (2) resolve the position of tetrad‐bearing Periplocoideae, the proposed milkweed sister group based on morphology; (3) place the enigmatic Dewevrella; and (4) clarify relationships of the three other primary crown clade lineages: Rhabdadenia, New World clade, and Asian clade. Separate analyses of PHYA and chloroplast sequences agree in placing an African monad‐bearing clade (Baisseeae plus Dewevrella) as the sister group of the milkweeds. Combined PHYA and chloroplast datasets under parsimony and maximum likelihood reject Periplocoideae as the milkweed sister group with statistical significance. Rhabdadenia is placed as sister to the rest of the crown clade, consistent with aspects of its floral morphology and wood anatomy, but without statistical support. Other relationships among the primary crown clade Apocynaceae lineages are also not supported statistically, but power analysis indicates that four to eight times as many characters (26,376–52,752 aligned positions) will be sufficient for a robust estimate.
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Introduction: Within the Apocynoideae (Apocynaceae) pro-toxic dehydropyrrolizidine alkaloids have been reported only in Echiteae. However, attraction of pyrrolizidine alkaloid-pharmacophagous insects suggested their presence in Alafia cf. caudata Stapf (Nerieae: Alafiinae) and Amphineurion marginatum (Roxb.) D.J. Middleton (Apocyneae: Amphineuriinae), both used as medicinal plants. Objective: To confirm the presence of dehydropyrrolizidine alkaloids in Alafia cf. caudata and Amphineurion marginatum and identify their structures. Methods: Methanol extracts of air-dried roots, stems and leaves of non-flowering plants were analysed using HPLC-ESI(+)MS and MS/MS or collision-induced dissociation MS in low and/or high resolution modes. Pyrrolizidine alkaloids were tentatively identified based on the mass spectrometry data. Solid phase extraction combined with semi-preparative HPLC were used to isolate major alkaloids. Structures were elucidated using NMR spectroscopy. Results: Monoesters of retronecine with senecioic, hydroxysenecioic or syringic acids were identified in roots of Alafia cf. caudata. Two unprecedented 10-membered macrocyclic dehydropyrrolizidine alkaloid diesters were isolated from roots of Amphineurion marginatum. Pyrrolizidine alkaloids were detected in root and leaf material of Alafia cf. caudata at 0.34 and 0.01% dry weight (DW), and 0.13, 0.02 and 0.09% DW in root, leaf and stem material of Amphineurion marginatum. Conclusions: The presence of pro-toxic dehydropyrrolizidine alkaloids suggests that medical preparations of these plants pose potential health risks to consumers. Dehydropyrrolizidine alkaloids are evidently more widespread in Apocynoideae than previously assumed, and it would seem rewarding to study other members of this family for the presence of pyrrolizidines, dehydropyrrolizidines and dihydropyrrolizines. Copyright © 2016 John Wiley & Sons, Ltd.
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A synopsis of the three species of the genus Laubertia (Apocynaceae, Apocynoideae, Echiteae) is presented here. Keys, descriptions, distributional data, and taxonomic index are provided.