Dissertação (mestrado)—Universidade de Brasília, Departamento de Botânica, Mestrado em Botânica, 2015. / Submitted by Albânia Cézar de Melo (albania@bce.unb.br) on 2015-05-19T16:09:10Z No. of bitstreams: 1 2015_TamaraSilvaDantas.pdf: 2336665 bytes, checksum: c1a40f86652384265e20fef812a4a640 (MD5) / Approved for entry into archive by Guimaraes Jacqueline(jacqueline.guimaraes@bce.unb.br) on 2015-05-20T11:17:50Z (GMT) No. of bitstreams: 1 2015_TamaraSilvaDantas.pdf: 2336665 bytes, checksum: c1a40f86652384265e20fef812a4a640 (MD5) / Made available in DSpace on 2015-05-20T11:17:50Z (GMT). No. of bitstreams: 1 2015_TamaraSilvaDantas.pdf: 2336665 bytes, checksum: c1a40f86652384265e20fef812a4a640 (MD5) / Peperomia é um dos maiores gêneros das angiospermas basais, com aproximadamente 1.600 espécies, distribuídas pantropicalmente. A Ilha da Trindade é uma ilha oceânica brasileira localizada a 1.200 km da costa do Brasil que emergiu a cerca de 3 milhões de anos da zona abissal no Atlântico Sul. A Ilha possui uma área de 9,28 km² e altitude máxima de 620 metros onde ocorre uma floresta nebular, a floresta de samambaias. São registradas na Ilha da Trindade duas espécies de Peperomia: P. beckeri, endêmica e conhecida apenas pelo tipo, e P. glabella. Espécimes de P. glabella da ilha da Trindade e de P. glabella que ocorre no continente foram analisados com o objetivo de investigar se a Peperomia que ocorre em Trindadeé uma espécie nova e endêmica ou uma variação morfológica de P. glabella que ocorre no continente. Foram utilizados materiais oriundos de 22 herbários além de materiais coletados na Ilha da Trindade, na Bahia e no Rio de Janeiro. Foram amplificados o gene matk e o espaçador ITS para análises filogenéticas e moleculares, além de análises morfológicas comparativas entre P. glabella e suas variedades e a Peperomia que ocorre em Trindade. Foram encontradas diferenças genéticas e morfológicas significativas entre P. glabella e a Peperomia que ocorre em Trindade e uma nova espécie foi descrita para a Ilha da Trindade. Peperomia beckeri não foi reencontrada e foi analisado apenas o material desta espécie. Material-tipo das variedades que ocorrem no Brasil foram analisados e propõe-se novos sinônimos. / Peperomia is one of the largest genera of basal angiosperms with about 1,600 species pantropically distributed. Trindade is a Brazilian oceanic island located about 1,200 km of the coast Brazilian, that emerged about 3 million years of abyssal zone in the South Atlantic, has a maximum altitude of 620 meters and an area of 9.28 km². Are registered in the Trindade Island the occurrence of two species of Peperomia: P. beckeri, endemic and known only by the type and P. glabella. The aims of this study were know whether the Peperomia occurring in Trindade is a new species and endemic species or morphological variation of P. glabella. We used the matk gene and the spacer ITS for phylogenetic and molecular analysis, and comparative morphological analysis between P. glabella and varieties and Peperomia occurring in Trindade Island. Genetic and morphological significant differences were observed between P. glabella and Peperomia occurring in Trindade. The Peperomia from the Trindade island is a different species of P. glabella and P. beckeri, according to the description of the authors of the species. Therefore, the Peperomia occurring in the Trindade Island is a new and endemic species of this site.

As técnicas de DNA barcoding (código de barras de DNA) têm como objetivo principal a identificação taxonômica de organismos através da amplificação e análise de sequências de DNA curtas, padronizadas e previamente definidas. Apesar do sucesso relativo desta abordagem em animais usando um único locus, a aplicação deste método em plantas apresenta menor capacidade de identificar espécies usando uma única região gênica, levando a necessidade de utilização de múltiplos loci. Além disso, ainda há certo debate sobre qual região gênica seria mais apropriada para o DNA barcoding em plantas, embora as regiões plastidiais rbcL, matK e o espaçador intergênico trnH-psbA juntamente com o espaçador intergênico nuclear do RNA ribossomal (ITS) sejam as mais comumente utilizadas até então. As tribos Sisyrinchieae e Tigridieae da família Iridaceae foram testadas de acordo com diferentes métodos e marcadores indicados para o DNA barcoding em plantas. Os resultados indicaram uma alta universalidade para membros da tribo Sisyrinchieae, mas também revelaram uma capacidade de identificação de espécies considerada baixa. Apesar disto, os espaçadores ITS foram indicados como a melhor sequência para DNA barcoding em Sisyrinchieae. Em Tigridieae, problemas inerentes ao sequenciamento impediram a utilização dos ITS em nossas análises. Assim, apenas marcadores plastidiais foram utilizados na tentativa de identificar espécies, apresentando novamente resultados modestos. A região gênica que atingiu maior capacidade de identificação em Tigridieae foi o gene matK. A incapacidade de se alcançar maiores taxas de identificação provavelmente está relacionada à complexa história evolutiva apresentada pelos grupos em análise. Este trabalho forneceu o primeiro conjunto significativo de dados de DNA barcoding aplicados a dois importantes grupos de Iridaceae de considerável biodiversidade no Brasil. As tribos em análise apresentam espécies consideradas filogeneticamente próximas e são de difícil identificação devido a sua morfologia homogênea, principalmente em estado vegetativo, justificando plenamente o uso de métodos molecularespara a identificação taxonômica. / The main objective of DNA barcoding methods is the taxonomic identification of organisms by amplifying and analyzing short, standardized and previously defined DNA sequences. In spite of the relative success of this approach in animals using a single locus, the application of this method in plants has less ability to identify species using a single gene region, leading to the need of using multiple loci. Furthermore, there is still some debate concerning which gene region would be more suitable for DNA barcoding in plants, although the plastid regions rbcL, matK and the trnH-psbA intergenic spacer along with the nuclear intergenic spacer of ribossomal DNA (ITS) are the most commonly regions used thus far. The tribes Sisyrinchieae and Tigridieae of the family Iridaceae were tested according to different methods and markers used for DNA barcoding in plants. The results indicated a great universality for members of tribe Sisyrinchieae, but also showed a low ability to identify species. Nevertheless, ITS was imputed as the best sequence for DNA barcoding in Sisyrinchieae. In Tigridieae, problems inherent of ITS sequencing prevented its use in our analysis. Thus, only plastid markers were used in an attempt to identify species, showing modest results once again. The gene region that reached higher identification ability in Tigridieae was matK. The inability to achieve higher identification levels is probably related to the complex evolutionary history presented by the groups in question. This work provided the first large data set of DNA barcoding applied to two important groups of Iridaceae with significant biodiversity in Brazil. The tribes in question present species considered phylogenetically related and are difficult to identify due to their homogeneous morphology, especially in vegetative stage, fully justifying the use of molecular methods for taxonomic identification.

As técnicas de DNA barcoding (código de barras de DNA) têm como objetivo principal a identificação taxonômica de organismos através da amplificação e análise de sequências de DNA curtas, padronizadas e previamente definidas. Apesar do sucesso relativo desta abordagem em animais usando um único locus, a aplicação deste método em plantas apresenta menor capacidade de identificar espécies usando uma única região gênica, levando a necessidade de utilização de múltiplos loci. Além disso, ainda há certo debate sobre qual região gênica seria mais apropriada para o DNA barcoding em plantas, embora as regiões plastidiais rbcL, matK e o espaçador intergênico trnH-psbA juntamente com o espaçador intergênico nuclear do RNA ribossomal (ITS) sejam as mais comumente utilizadas até então. As tribos Sisyrinchieae e Tigridieae da família Iridaceae foram testadas de acordo com diferentes métodos e marcadores indicados para o DNA barcoding em plantas. Os resultados indicaram uma alta universalidade para membros da tribo Sisyrinchieae, mas também revelaram uma capacidade de identificação de espécies considerada baixa. Apesar disto, os espaçadores ITS foram indicados como a melhor sequência para DNA barcoding em Sisyrinchieae. Em Tigridieae, problemas inerentes ao sequenciamento impediram a utilização dos ITS em nossas análises. Assim, apenas marcadores plastidiais foram utilizados na tentativa de identificar espécies, apresentando novamente resultados modestos. A região gênica que atingiu maior capacidade de identificação em Tigridieae foi o gene matK. A incapacidade de se alcançar maiores taxas de identificação provavelmente está relacionada à complexa história evolutiva apresentada pelos grupos em análise. Este trabalho forneceu o primeiro conjunto significativo de dados de DNA barcoding aplicados a dois importantes grupos de Iridaceae de considerável biodiversidade no Brasil. As tribos em análise apresentam espécies consideradas filogeneticamente próximas e são de difícil identificação devido a sua morfologia homogênea, principalmente em estado vegetativo, justificando plenamente o uso de métodos molecularespara a identificação taxonômica. / The main objective of DNA barcoding methods is the taxonomic identification of organisms by amplifying and analyzing short, standardized and previously defined DNA sequences. In spite of the relative success of this approach in animals using a single locus, the application of this method in plants has less ability to identify species using a single gene region, leading to the need of using multiple loci. Furthermore, there is still some debate concerning which gene region would be more suitable for DNA barcoding in plants, although the plastid regions rbcL, matK and the trnH-psbA intergenic spacer along with the nuclear intergenic spacer of ribossomal DNA (ITS) are the most commonly regions used thus far. The tribes Sisyrinchieae and Tigridieae of the family Iridaceae were tested according to different methods and markers used for DNA barcoding in plants. The results indicated a great universality for members of tribe Sisyrinchieae, but also showed a low ability to identify species. Nevertheless, ITS was imputed as the best sequence for DNA barcoding in Sisyrinchieae. In Tigridieae, problems inherent of ITS sequencing prevented its use in our analysis. Thus, only plastid markers were used in an attempt to identify species, showing modest results once again. The gene region that reached higher identification ability in Tigridieae was matK. The inability to achieve higher identification levels is probably related to the complex evolutionary history presented by the groups in question. This work provided the first large data set of DNA barcoding applied to two important groups of Iridaceae with significant biodiversity in Brazil. The tribes in question present species considered phylogenetically related and are difficult to identify due to their homogeneous morphology, especially in vegetative stage, fully justifying the use of molecular methods for taxonomic identification.

Evolution of Piperales – matK gene and trnK intron sequence data reveal lineage specific resolution contrast. Piperales are one of the largest basal angiosperm orders with a nearly worldwide distribution. The order includes three species rich genera, Piper (ca. 1,000 species), Peperomia (ca. 1,500-1,700 species), and Aristolochia s. l. (ca. 500 species). Sequences of the matK gene and the non-coding trnK group II intron are analysed for a dense set of 105 taxa representing all families (except Hydnoraceae) and all generic segregates (except Euglypha within Aristolochiaceae) of Piperales. A large number of highly informative indels are found in the Piperales trnK/matK dataset. Within a narrow region approximately 500 nt downstream in the matK coding region (CDS), a length variable simple sequence repeat (SSR) expansion segment occurs, in which insertions and deletions have led to short frame-shifts. These are corrected shortly afterwards, resulting in a maximum of 6 amino acids being affected. Furthermore, additional non-functional matK copies were found in Zippelia begoniifolia, which can easily be discriminated from the functional open reading frame (ORF). The trnK/matK sequence data fully resolve relationships within Peperomia, whereas they are not effective within Piper. The resolution contrast is correlated with the rate heterogenity between those lineages. Parsimony, Bayesian and likelihood analyses result in virtually the same topology, and converge on the monophyly of Piperaceae and Saururaceae. Lactoris gains high support as sister to Aristolochiaceae subf. Aristolochioideae, but the different tree inference methods yield conflicting results with respect to the relationships of subfam. Asaroideae. In Piperaceae, a clade formed by the monotypic genus Zippelia and the small genus Manekia (=Sarcorhachis) is sister to the two large genera Piper and Peperomia. Systematics of pipevines – Combining morphological and fast-evolving molecular characters to investigate the relationships within subfamily Aristolochioideae (Aristolochiaceae) A combined phylogenetic analysis of the Aristolochioideae was conducted based on 72 morphological characters and molecular datasets (matK gene, trnK intron, trnL intron, trnL-trnF spacer). The analysis sampled 33 species as the ingroup, including two species of Thottea and 30 species of Aristolochia and the monotypic genus Euglypha, which represent all the infrageneric taxa formally described; Saruma henryi and Asarum caudatum were used as the outgroup. The results corroborate a sister-group relationship between Thottea and Aristolochia, and the paraphyly of Aristolochia with respect to Euglypha that consequently should be included into Aristolochia. Two of the three subgenera within Aristolochia (Isotrema and Pararistolochia) are shown to be monophyletic, whereas the signal obtained from the different datasets about the relationships within subg. Aristolochia is low and conflicting, resulting in collapsed or unsupported branches. The relationship between the New World and the Old World species of subgenus Aristolochia is conflictive because morphological data support these two groups as monophyletic, whereas molecular data show the monophyletic Old World species of Aristolochia nested within the New World species. A sister group relationship is proposed between A. lindneri and pentandrous species, which suggests that a group of five species from central and southern South America (including A. lindneri) could be monophyletic and sister to Aristolochia subsection Pentandrae, a monophyletic taxon consisting of about 35 species from southern USA, Mesoamerica, and the West Indies. Colonisation, phylogeography and evolution of endemism in Mediterranean Aristolochia (Aristolochiaceae). This study provides evidence for a multiple colonisation of the western Old World from Asian ancestors within Aristolochia section Diplolobus (subsection Aristolochia and Podanthemum). Within subsection Podanthemum it is assumed, that the colonisation of the African continent happened at least two times independently. In contrast, for subsection Aristolochia, a rapid morphological radiation in the Near East (or close to this area) with subsequent star like colonisation of the different current distribution areas, which is not paralleled on the molecular level, appears to be more likely. Phylogenetic tree reconstruction is unsupported for these clades, but most clades are highly supported as monophyletic. Interestingly the Mediterranean and temperate Eurasian species, which are morphologically distinct (A. pistolochia, A. clematitis) are not clustering within the main clades, but are independent lineages. Analogue, A. rigida a species from Somalia is well-supported sister to the subsection Aristolochia. Within subsection Podanthemum the colonisation event from an Asian ancestor is clearly traceable, whereas in subsection Aristolochia the path is not traceable, since the ancestors are extinct or not present in the connecting areas. Within the Mediterranean, Near East and Caucasian species of subsection Aristolochia two morphologically and biogeographically well supported groups can be identified: the Near East/Caucasian species and the West Mediterranean species. The previous groupings for the latter, based on morphological characters, could be substantiated only partly by our results. This study provides the first phylogeny of all West Mediterranean species. In addition an independent complex is established including some micro endemic species. The phylogenetic results are discussed with respect to biogeography, and morphology, to give a first insight into the radiation and colonisation of the genus Aristolochia in the Mediterranean region. Universal primers for a large cryptically simple cpDNA microsatellite region in Aristolochia. We provide a new and valuable marker to study species relationships and population genetics in order to trace evolutionary, ecological, and conservational aspects in the genus Aristolochia. Universal primers for amplification and subsequent sequencing of a chloroplast microsatellite locus inside the trnK intron are presented. Utility of the primers has been tested in 32 species representing all clades of Aristolochia, including population studies within the A. pallida complex, A. clusii and A. rotunda. The microsatellite region is characterized as a (AnTm)k repeat of 22–438 bp containing a combination of different repeats arranged as ‘cryptically simple’. Trapped! Pollination of Aristolochia pallida Willd. in the Mediterranean A first study of the pollination biology of a Mediterranean Aristolochia species in its natural habitat is presented. 183 flowers of Aristolochia pallida were investigated, which in total contained 73 arthropods, dominated by two groups of Diptera, Sciaridae (37%) and Phoridae (19%). However, only Phoridae are regarded as potential pollinators, since pollen has been found exclusively on the body surfaces of these insects. All Phoridae belong to the genus Megaselia and are recognised as four undescribed species. The measurements of flower and insect dimensions suggest that size is an important constrain for successful pollination: 1) the insects must have a definitive size for being able to enter the flower and 2) must be able to get in touch with the pollen. Only very few insect groups found in Aristolochia pallida fulfil these size requirements. However, size alone is not a sufficient constrain as too many fly species of the same size might be trapped but not function as pollinators. Instead, specific attraction is required as otherwise pollen is lost. Since all trapped Phoridae are males, a chemical attraction (pheromones) is proposed as an additional constrain. Since A. pallida flowers are protogynous, the record of Megaselia loaded with pollen found in a flower during its female stage proves that this insect must have been visited at least one different flower during its male stage before. Further on, this observation provides strong evidence that the flowers are cross-pollinated. All these factors indicate a highly specialised pollination of Aristolochia pallida by Megaselia species.

Herbal medicines are growing in popularity in the Western world and are becoming more stringently regulated under new EU legislation. Within the arena of herbal medicines, St. John’s Wort (SJW), Hypericum perforatum, is a top ten best seller with clinical evidence to support its use as an anti-depressant. A fundamental requirement of the new legislation is to prove the identity of the plant material in question. This is currently achieved via morphological and chemical methods, neither of which are ideal. A wide range of DNA based methods have been applied to this arena, standardisation is required to realise the potential of DNA based techniques. The DNA barcoding initiative aims to produce sequence data for all plant species, capable of species identification. The proposal is to use these data to design fast and effective DNA based methods of identification. For assay design, the putative barcode region nrITS was selected as a platform. Three assays were designed; • A PCR assay designed to hyper variable sequences within a barcode region. This assay is capable of distinguishing SJW from other closely related species. • A quantitative qPCR assay designed to measure total DNA and specific SJW DNA within a mixed sample. • A multiplex PCR incorporating fluorescently labelled primers, allowing amplicon detection by capillary electrophoresis. This assay identifies four separate Hypericum species, including SJW, with a mixed sample in one reaction. The suitability of the nrITS and three other barcode regions is assessed based on sequence data generated for 32 vouchered samples of different Hypericum species, and a Lithuanian sample set of 22 and 16 H. perforatum and H. maculatum samples respectively. The matK is currently unusable, the rbcL highly conserved, trnH-psbA problematically variable and the nrITS proved to be ideal for assay design.

La culture de saules (Salix sp.) est une pratique courante en Europe et en Amérique du Nord pour produire de la biomasse végétale. Cependant, le développement d’outils moléculaires est très récent. De plus, la phylogénie des saules est incomplète. Il y a un manque d’information pour les programmes de sélection d'espèces indigènes et pour la compréhension de l’évolution du genre. Le genre Salix inclut 500 espèces réparties principalement dans les régions tempérées et boréo-arctique de l’hémisphère nord. Nous avons obtenu l’ensemble des espèces retrouvées naturellement en Amérique (121 indigènes et introduites). Dans un premier temps, nous avons développé de nouveaux outils moléculaires et méthodes : extraction d’ADN, marqueurs microsatellites et gènes nucléaires. Puis, nous avons séquencé deux gènes chloroplastiques (matK et rbcL) et la région ITS. Les analyses phylogénétiques ont été réalisées selon trois approches : parcimonie, maximum de vraisemblance et Bayésienne. L’arbre d’espèces obtenu a un fort support et divise le genre Salix en deux sous-genres, Salix et Vetrix. Seize espèces ont une position ambiguë. La diversité génétique du sous-genre Vetrix est plus faible. Une phylogénie moléculaire complète a été établie pour les espèces américaines. D’autres analyses et marqueurs sont nécessaires pour déterminer les relations phylogénétiques entre certaines espèces. Nous affirmons que le genre Salix est divisé en deux clades. / Fast growing willows (Salix sp.) are increasingly used in Europe and North America for biomass production and other environmental applications. However, the development of molecular tools is recent. The phylogeny of willows is incomplete, which slows down the selection of suitable native species and the development of improvement programs. The genus Salix includes approximately 500 species worldwide, and these are mainly located in temperate and cold regions of the Northern Hemisphere. We gathered leaf material from all 121 willows of North America (species native and introduced). We developed three molecular tools-methods: DNA extraction, SSR markers, and nuclear genes. We sequenced two chloroplast genes matK and rbcL and the ITS region. Phylogenetic analyses were carried out using parsimony, maximum likelihood and Bayesian approaches. The species tree provides strong support for a division of the genus into two subgenera, Salix and Vetrix. Sixteen species have ambiguous positions. A complete molecular phylogeny of American willows has been established. It needs to be confirmed and further resolved using other molecular data. Nonetheless, the genus clearly has two clades.

Bauhinia s.l. est le plus vaste genre de la tribu des Cercideae (Ceasalpinioideae, Leguminoseae), avec plus de 300 espèces. Il présente une distribution pantropicale et une grande variabilité morphologique. Ces deux caractéristiques ont limité les études taxonomiques sur le genre complet, résultant en plusieurs études taxonomiques de certains groupes seulement. En 1987, Wunderlin et al. proposent une vaste révision taxonomique de la tribu des Cercideae, basée sur des données morphologiques, et divisent le genre Bauhinia en quatre sous-genres. En 2005, Lewis et Forest publient une nouvelle classification préliminaire basée sur des données moléculaires, mais sur un échantillonnage taxonomique restreint. Leurs conclusions remettent en question le monophylétisme du genre Bauhinia et suggèrent plutôt la reconnaissance de huit genres au sein du grade Bauhinia s.l. Afin de vérifier les hypothèses de Lewis et Forest, et obtenir une vision plus claire de l’histroire de Bauhinia s.l., nous avons séquencé deux régions chloroplastiques (trnL-trnF et matK-trnK) et deux régions nucléaires (Leafy et Legcyc) pour un vaste échantillonnage représentatif des Cercideae. Une première phylogénie de la tribu a tout d’abord été réalisée à partir des séquences de trnL-trnF seulement et a confirmé le non-monoplylétisme de Bauhinia s.l., avec l’inclusion du genre Brenierea, traditionnellement reconnu comme genre frère de Bauhinia s.l. Afin de ne pas limiter notre vision de l’histoire évolutive des Cercideae à un seul type de données moléculaires et à une seule région, une nouvelle série d’analyse a été effectuée, incluant toutes les séquences chloroplastiques et nucléaires. Une phylogénie individuelle a été reconstruite pour chacune des régions du génome, et un arbre d’espèce ainsi qu’un arbre de supermatrice ont été reconstruits. Bien que certaines contradictions apparaissent entre les phylogénies, les grandes lignes de l’histoire des Cercideae ont été résolues. Bauhinia s.l. est divisée en deux lignées : les groupes Phanera et Bauhinia. Le groupe Bauhinia est constitué des genres Bauhinia s.s., Piliostigma et Brenierea. Le groupe Phanera est constitué des genres Gigasiphon, Tylosema, Lysiphyllum, Barklya, Phanera et Schnella. Les genres Cercis, Adenolobus et Griffonia sont les groupes-frères du clade Bauhinia s.l. Au minimum un événement de duplication de Legcyc a été mis en évidence pour la totalité de la tribu des Cercideae, excepté Cercis, mais plusieurs évènements sont suggérés à la fois par Legcyc et Leafy. Finalement, la datation et la reconstruction des aires ancestrales de la tribu ont été effectuées. La tribu est datée de 49,7 Ma et est originaire des régions tempérées de l’hémisphère nord, probablement autour de la mer de Thétys. La tribu s’est ensuite dispersée vers les régions tropicales sèches de l’Afrique, où la séparation des groupes Bauhinia et Phanera a eu lieu. Ces deux groupes se sont ensuite dispersés en parallèle vers l’Asie du sud-est au début du Miocène. À la même période, une dispersion depuis l’Afrique de Bauhinia s.s. a permis la diversification des espèces américaines de ce genre, alors que le genre Schnella (seul genre américain du groupe Phanera) est passé par l’Australie afin de rejoindre le continent américain. Cette dispersion vers l’Australie sera également à l’origine des genres Lysiphyllum et Barklya / Bauhinia s.l. is the largest genus of the tribe Cercideae (Ceasalpinioideae, Leguminoseae), with over 300 species. It has a pantropical distribution and high morphological variability. These two features have resulted in few studies that focus on the entire genus, resulting in several regional studies or studies of certain subgroups only. In 1987, Wunderlin et al. presented a broad taxonomic revision of the tribe Cercideae, based on morphological data, and divided the genus Bauhinia into four subgenera. In 2005, Lewis and Forest published a new preliminary classification based on molecular data, but for a limited taxonomic sampling. Their findings question the monophyly of the genus Bauhinia and suggest instead the recognition of eight genera in the Bauhinia s.l. grade. To test the hypotheses of Lewis and Forest, and to obtain a clearer view of the history of Bauhinia s.l., we sequenced two chloroplast regions (trnL-trnF and matK-trnK) and two nuclear regions (Leafy and Legcyc) for a large representative sampling of the Cercideae. A primary phylogeny of the tribe was first generated based on trnL-trnF sequences only and confirmed the non-monophyly of Bauhinia s.l., with the inclusion of the genus Brenierea, traditionally recognized as sister group of Bauhinia s.l. In order to obtain a deaper view of the evolutionary history of the Cercideae, a new series of analysis was performed, including all nuclear and chloroplast sequences. Individual phylogenies were reconstructed for each region of the genome, and both a species and a supermatrix trees were reconstructed. Although certain conflicting relationships appear between phylogenies, the outline of the history of the Cercideae has been resolved. Bauhinia s.l. is divided into two lineages: Phanera and Bauhinia groups. The Bauhinia group includes Bauhinia s.s., Piliostigma and Brenierea. The Phanera group is composed of Gigasiphon, Tylosema, Lysiphyllum, Barklya, Phanera and Schnella. Cercis, Griffonia and Adenolobus are sister groups of Bauhinia s.l. At least one duplication event of Legcyc has been highlighted for the entire tribe Cercideae, excluding Cercis. Several other duplication events are also suggested by both Legcyc and Leafy . Finally, a divergence time analysis and a reconstruction of ancestral areas were conducted. The root of the tribe is evaluated to be 49.7 Mya old, and to originate from temperate regions in the northern hemisphere, mostly around the Tethys Sea. The tribe then dispersed into drier biomes in Africa, where the separation of the Bauhinia and the Phanera groups occurred. These two lineages then dispersed following parallel routes to Southeast Asia in the early Miocene. At the same time, a dispersal of the African Bauhinia s.s. to South America permitted the diversification of the American species of this genus, and Schnella (the only American genus within the Phanera group) dispersed to the American continent from Australia. This dispersal to Australia is also at the origin of Lysiphyllum and Barklya.

La culture de saules (Salix sp.) est une pratique courante en Europe et en Amérique du Nord pour produire de la biomasse végétale. Cependant, le développement d’outils moléculaires est très récent. De plus, la phylogénie des saules est incomplète. Il y a un manque d’information pour les programmes de sélection d'espèces indigènes et pour la compréhension de l’évolution du genre. Le genre Salix inclut 500 espèces réparties principalement dans les régions tempérées et boréo-arctique de l’hémisphère nord. Nous avons obtenu l’ensemble des espèces retrouvées naturellement en Amérique (121 indigènes et introduites). Dans un premier temps, nous avons développé de nouveaux outils moléculaires et méthodes : extraction d’ADN, marqueurs microsatellites et gènes nucléaires. Puis, nous avons séquencé deux gènes chloroplastiques (matK et rbcL) et la région ITS. Les analyses phylogénétiques ont été réalisées selon trois approches : parcimonie, maximum de vraisemblance et Bayésienne. L’arbre d’espèces obtenu a un fort support et divise le genre Salix en deux sous-genres, Salix et Vetrix. Seize espèces ont une position ambiguë. La diversité génétique du sous-genre Vetrix est plus faible. Une phylogénie moléculaire complète a été établie pour les espèces américaines. D’autres analyses et marqueurs sont nécessaires pour déterminer les relations phylogénétiques entre certaines espèces. Nous affirmons que le genre Salix est divisé en deux clades. / Fast growing willows (Salix sp.) are increasingly used in Europe and North America for biomass production and other environmental applications. However, the development of molecular tools is recent. The phylogeny of willows is incomplete, which slows down the selection of suitable native species and the development of improvement programs. The genus Salix includes approximately 500 species worldwide, and these are mainly located in temperate and cold regions of the Northern Hemisphere. We gathered leaf material from all 121 willows of North America (species native and introduced). We developed three molecular tools-methods: DNA extraction, SSR markers, and nuclear genes. We sequenced two chloroplast genes matK and rbcL and the ITS region. Phylogenetic analyses were carried out using parsimony, maximum likelihood and Bayesian approaches. The species tree provides strong support for a division of the genus into two subgenera, Salix and Vetrix. Sixteen species have ambiguous positions. A complete molecular phylogeny of American willows has been established. It needs to be confirmed and further resolved using other molecular data. Nonetheless, the genus clearly has two clades.

Evolutionary relationships within and among three Astragalus sections (Incani DC., Hypoglottidei DC., and Dissitiflori DC.) that were native to Turkey were inferred from variations of nucleotide sequences of both chloroplast and nuclear genome regions. In the current study, Fifty-six species included in the three Astragalus sections were utilized to figure out phylogenetic relationships and estimate evolutionary divergence time based on DNA sequence of trnL intron (trnL5&rsquo / -L3&rsquo / ) , trnL3&rsquo / -F(GAA) (trnL-F intergenic spacer), trnV intron, matK (maturase kinase) cpDNA (chloroplast) and ITS (internal transcribed spacer) nDNA (nuclear) regions. Fifty-six Astragalus species with their replicas and one Cicer species as outgroup were analyzed by polymerase chain reaction amplification and DNA sequencing methods. Eleven unknown samples were also used in the current study to understand their section and species name. The results of the study indicated that unknown A35 and A52 samples could be named as A. dasycarpus, while unknown A65 and A66 samples as A. ovatus and lastly unknown A2 sample as A. nitens or A. aucheri. Section of unknown A3, A16, A20, A108, A109 and A110 samples were determined as Incani, but the exact species identification of these samples were not possible because of their close phylogenetic associations with more than one species. Highest genetic diversity was observed when the DNA sequences of ITS nrDNA (nuclear ribosomal) region comprising three subregions as ITS1, 5.8S and ITS2 was used, while the lowest one was calculated when DNA sequence of trnL-F cpDNA region was analyzed. The genetic divergence between Incani and Dissitiflori sections was highest whereas between Hypoglottidei and Dissitiflori was lowest based on all used regions. To figure out phylogenetic relationships among Astragalus species distributed in Turkey and in other regions of the World, DNA sequences of studied regions of foreign samples were collected from the NCBI database and were evaluated with DNA sequence of Turkish species used in the curent study. The Iranian samples either scattered in the phylogenetic tree or attached to our samples externally. South and North American samples (New World Astragalus or Neo Astragalus group) were nested within a different subcluster, which was located in the main cluster produced by samples of Old World Astragalus group (Turkish samples). With these results, we can say that New World Astragalus group is monophyletic and diverged from Old World Astragalus group. Evolutionary divergence time for Astragalus genus was estimated as about 12.5 - 14.5 million years (Ma), and that of New World Astragalus group as 5.0 - 4.0 Ma when rates of nucleotide substitutions of trnL intron and matK cpDNA regions were analyzed. In addition to evolutionary divergence time estimation for Astragalus and New World Astragalus group, divergence times among used three sections of the genus were also calculated by using DNA sequences of trnL, trnV intron and matK cpDNA regions and results indicated that Hypoglottidei and Dissitiflori sections diverged about 5.0-7.0 million years later than Incani section.

Dentre as orquídeas terrícolas brasileiras, dois grupos são particularmente importantes do ponto de vista taxonômico, sistemático e biogeográfico: as orquídeas das subtribo Chloraeinae e Spiranthinae. Na sua atual delimitação, Chloraeinae compreende aproximadamente 70 espécies em três gêneros: Chloraea, Bipinnula e Gavilea. Por outro lado, Spiranthinae compreende cerca de 470 espécies exclusivamente neotropicais, distribuídas em 42 gêneros. As análises filogenéticas mais recentes suportam a existência de quatro grupos: os clados Stenorhynchos, Pelexia, Eurystyles+Lankesterella e Spiranthes. Assim, os objetivos deste estudo são: 1) realizar um estudo taxonômico e nomenclatural das orquídeas brasileiras do clado Pelexia; 2) documentar a morfologia floral e vegetativa, com ênfase nos caracteres diagnósticos; 3) realizar um estudo taxonômico e nomenclatural das espécies brasileiras de Chloraeinae; 4) construir uma filogenia molecular mais completa e representativa do clado Pelexia; 5) estabelecer se os gêneros do clado Pelexia são monofiléticos; 6) construir uma filogenia molecular mais completa e representativa de Chloraeinae, com a inclusão de táxons brasileiros; 7) estabelecer um conjunto de caracteres diagnósticos para os gêneros envolvidos neste estudo e, se necessário, propor uma nova delimitação genérica; e 8) propor cenários para a provável evolução de caracteres de importância ecológica. Os resultados de um estudo detalhado, bem como de análises morfológicas e moleculares são: 1) propostas de 48 tipificações, novas sinonímias e notas taxonômicas para as espécies de Goodyerinae e Spiranthinae descritas por José Velloso e Barbosa Rodrigues; 2) novos registros de Pteroglossa e Lyroglossa para o Rio Grande do Sul; 3) uma correção nomenclatural de Spiranthes bicolor e 4) uma sinopse taxonômica das Chloraeinae brasileiras; 5) uma análise filogenética molecular para testar a monofilia dos gêneros atualmente circunscritos no clado Pelexia; 6) uma atualização das relações filogenéticas de Chloraeinae, incluindo espécies Pampeanas de Bipinnula e a espécie tipo do gênero, B. biplumata; 7) um panorama sobre a biologia reprodutiva de duas espécies de Pachygenium e dados adicionais sobre Brachystele e Cyclopogon s.l.; e 8) um estudo detalhado da morfologia floral, biologia da polinização e sistema reprodutivo de Chloraea membranacea (Chloraeinae). / Among the Brazilian terrestrial orchids, two groups are particularly important from a taxonomic, systematic and biogeographical aspects: orchids of subtribe Chloraeinae and Spiranthinae. In its current delimitation, Chloraeinae comprises approximately 70 species distributed in three genera: Chloraea, Bipinnula and Gavilea. On the other hand, Spiranthinae comprises about 470 exclusively neotropical species distributed in ca. 42 genera. The most recent phylogenetic analyses support the existence of four clades: Stenorrhynchos, Pelexia, Eurystyles+Lankesterella and Spiranthes clade, respectively. Thus, the aims of this study are: 1) to make a taxonomic and nomenclatural study of Brazilian orchids of Pelexia clade; 2) to document the floral and vegetative morphology, with emphasis on diagnostic features; 3) to make a taxonomic and nomenclatural study of the Brazilian species of Chloraeinae; 4) to assemble a more complete and representative molecular phylogeny of the Pelexia clade; 5) to establish if the genera of the Pelexia clade are monophyletic; 6) to construct a more complete and representative molecular phylogeny of the Chloraeinae, with the inclusion of Brazilian taxa; 7) to establish sets of diagnostic characters for the genera involved in this study and, if necessary, propose a new generic delimitation; and 8) to propose scenarios for the evolution of characters of ecological importance. The main results are: 1) 48 typifications, new synonymies and taxonomic notes for Spiranthinae and Goodyerinae species described by José Velloso and Rodrigues Barbosa; 2) new records of Pteroglossa and Lyroglossa to Rio Grande do Sul; 3) a nomenclatural correction of Spiranthes bicolor and 4) a taxonomic synopsis of Brazilian Chloraeinae; 5) molecular phylogenetic analyses to test the monophyly of the currently circumscribed genera in the Pelexia clade; 6) an update of the phylogenetic relationships of the Chloraeinae, including Pampean species of Bipinnula, and the type species of the genus, B. biplumata; 7) an overview on the reproductive biology of two Pachygenium species and additional data on Brachystele and Cyclopogon s.l.; and 8) a detailed study of the floral morphology, pollination biology and reproductive system of Chloraea membranacea (Chloraeinae).