Orientador: Marlies Sazima / Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-16T08:56:09Z (GMT). No. of bitstreams: 1 Brito_ViniciusLourencoGarcia_M.pdf: 8743517 bytes, checksum: efdc732f7b02ccea2c628acf24677361 (MD5) Previous issue date: 2010 / Resumo: As montanhas apresentam alta diversidade e diferentes condições ambientais ao longo de curtas distâncias. Assim, as montanhas são ideais para estudos ecológicos e evolutivos que podem somar valores e aprimorar projetos de conservação. Em altitudes elevadas as condições ambientais podem reduzir a quantidade de polinizadores, principalmente de abelhas. Desta forma, em espécies estritamente melitófilas, características como a fenologia, o sistema reprodutivo, o fluxo de pólen e a estrutura genética das populações pode variar ao longo do gradiente, uma vez que a transferência de grãos de pólen aos estigmas co-específicos também varia ao longo do gradiente. No caso de áreas de elevada altitude, a transferência de pólen é limitada (limitação de pólen), reduzindo as possibilidades de polinização cruzada. O presente estudo tem por objetivo obter informações sobre a biologia da polinização, reprodução e genética de duas populações de Tibouchina pulchra (Melastomataceae) ocorrentes em duas áreas de gradiente altitudinal: Núcleo Santa Virgínia (NSV) e Núcleo de Desenvolvimento Picinguaba (NDP) do Parque Estadual da Serra do Mar. Foram feitas observações mensais para definir padrões e estratégias de floração, registrar dados sobre a biologia floral e reprodutiva, além de verificar a riqueza e abundância dos polinizadores e caracterizar as interações dessa espécie com as abelhas visitantes. Material genético de 44 indivíduos do NSV e 45 indivíduos do NDP foram coletados para o desenvolvimento e caracterização de 12 loco microssatélites polimórficos e estes foram utilizados para fazer análises de agrupamento, ordenação bayesiana e medidas de diversidade genética nas duas populações. Os aspectos da biologia reprodutiva são diferentes entre as duas áreas: na área elevada a florada é mais intensa, a produção de pólen é menor, há limitação na transferência de pólen, mas a fertilização de sementes provindas de polinização cruzada manual é maior. Na outra área são produzidos mais frutos e há maior riqueza e abundância de polinizadores. Ocorre diferenciação genética entre as populações, mas com uma interface de contato entre elas, além de menor diversidade genética na população da área elevada. Estes resultados indicam que a ausência de polinizadores na região de altitude elevada está associada a diferentes estratégias na biologia floral e reprodutiva para balancear a limitação de pólen. Além disso, diferentes dinâmicas de fluxo gênico mediado pelo pólen nas duas populações e as características de distribuição e reprodução podem influenciar a estrutura e a diversidade genética de Tibouchina pulchra ao longo da Serra do Mar / Abstract: Mountains have high diversity and many environmental conditions at short distances. Hence, they are an ideal place to develop ecological and evolutionary studies that can improve conservation projects. At high altitudes, the environmental conditions reduce pollinator abundance, mainly bees. Therefore, traits such as phenology, breeding system and genetic structure of plant populations pollinated by bees could vary in an altitudinal gradient, because pollen grain transference to co-specific flowers varies also. At high altitudes pollen transference is limited (pollen limitation) reducing cross-pollination. The main goal of the present study was to obtain information about pollination biology, breeding system and genetic structure of two populations of Tibouchina pulchra (Melastomataceae) that occur in two different areas of an altitudinal gradient: Núcleo Santa Virgínia (NSV) and Núcleo de Desenvolvimento Picinguaba (NDP) of Parque Estadual da Serra do Mar. Field work was done monthly to describe the flowering patterns and strategies, record floral and reproductive biology, verify pollinator diversity and characterize the interactions among this plant species and bee visitors. Genetic material was collected from 44 Tibouchina pulchra individuals of NSV and 45 individuals of NDP to developed and characterize 12 microsatellite polymorphic loci, which were used to analyze population's genetics by different methods. At the higher area the plants have greater flowering intensity, the flowers produce less pollen grains and the stigmas receive less pollen, but set more seeds after manual cross pollination than individuals at the lower area, where pollinator diversity is higher and plants produce more fruits. There is genetic differentiation, but also an interface contact between the two populations. The population at higher altitude has less genetic diversity than the one at lower altitude. The lack of pollinators at the higher area is associated with different floral and reproductive strategies to compensate pollen limitation. Moreover the genetic structure and diversity respond to different dynamics of pollen flow and the patterns of distribution and reproduction of Tibouchina pulchra at the altitudinal gradient of Serra do Mar / Mestrado / Biologia Vegetal / Mestre em Biologia Vegetal

Part 1: Visitation of potential pollinators to Cornus florida and C. kousa flowers was assessed in East Tennessee in 2008 and 2009. Data regarding insect visitation rates to multiple trees per location were gathered throughout the flowering period. Diurnal and seasonal variations in visitation were recorded. Pollen coverage was assessed on portions of captured insect exoskeletons that were most likely to contact the stigma. Eleven families in four insect orders were collected from C. florida and 26 families in five orders from C. kousa. The most important pollinators in eastern Tennessee were bees in the Andrenidae and Halictidae. The most common visitors to C. kousa flowers were scarab, cerambycid and cantharid beetles. Halictid bees were also frequent visitors to C. kousa. Part 2: Dispersion of flowering dogwood pollen in an orchard was evaluated by performing parentage analysis on open pollinated seedlings collected from a single maternal tree. Pollen source for 45 seedling trees were established using three polymorphic simple sequence repeat (SSR) loci. The expected leptokurtic distribution was not seen. Although the majority of paternal trees were within the 12 m radius of the study area, the trees most likely to donate pollen were some of the furthest away. Unusual pollen movement may be a product of lack of synchronization of flowering times between the mother tree and potential pollen donors in the area. Additionally relative proportions of certain pollinating insects (andrenid and halictid bees) may have moved pollen further than expected. Appendix 1: Pollen from 6 species in the genus Cornus was analyzed with Fourier Transform Infrared (FTIR) Spectroscopy to determine the utility of this tool to identify pollen grains to species. Although there were differences visible in the spectra, principal components analysis coupled with cluster analysis could not consistently identify pollen grains. Appendix 2: Floral volatile emissions from 6 species of Cornus were collected in a headspace chamber and analyzed with gas chromatography-mass spectrometry to determine if differences between emitted volatiles of C. florida and C. kousa explain differences in floral insect visitation. The relationship between floral volatile emission and the phylogeny of Cornus was also examined.

Part 1: Visitation of potential pollinators to Cornus florida and C. kousa flowers was assessed in East Tennessee in 2008 and 2009. Data regarding insect visitation rates to multiple trees per location were gathered throughout the flowering period. Diurnal and seasonal variations in visitation were recorded. Pollen coverage was assessed on portions of captured insect exoskeletons that were most likely to contact the stigma. Eleven families in four insect orders were collected from C. florida and 26 families in five orders from C. kousa. The most important pollinators in eastern Tennessee were bees in the Andrenidae and Halictidae. The most common visitors to C. kousa flowers were scarab, cerambycid and cantharid beetles. Halictid bees were also frequent visitors to C. kousa.Part 2: Dispersion of flowering dogwood pollen in an orchard was evaluated by performing parentage analysis on open pollinated seedlings collected from a single maternal tree. Pollen source for 45 seedling trees were established using three polymorphic simple sequence repeat (SSR) loci. The expected leptokurtic distribution was not seen. Although the majority of paternal trees were within the 12 m radius of the study area, the trees most likely to donate pollen were some of the furthest away. Unusual pollen movement may be a product of lack of synchronization of flowering times between the mother tree and potential pollen donors in the area. Additionally relative proportions of certain pollinating insects (andrenid and halictid bees) may have moved pollen further than expected.Appendix 1: Pollen from 6 species in the genus Cornus was analyzed with Fourier Transform Infrared (FTIR) Spectroscopy to determine the utility of this tool to identify pollen grains to species. Although there were differences visible in the spectra, principal components analysis coupled with cluster analysis could not consistently identify pollen grains.Appendix 2: Floral volatile emissions from 6 species of Cornus were collected in a headspace chamber and analyzed with gas chromatography-mass spectrometry to determine if differences between emitted volatiles of C. florida and C. kousa explain differences in floral insect visitation. The relationship between floral volatile emission and the phylogeny of Cornus was also examined.

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No. of bitstreams: 2 Dissertação - Camila Fernanda Costa - 2013.pdf: 2572672 bytes, checksum: 54a6e93e72c746e4888a6f3583d4f3ac (MD5) license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) Previous issue date: 2013-03-23 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / Fundação de Amparo à Pesquisa do Estado de Goiás - FAPEG / Hancornia speciosa (Apocynaceae) is a fruit species which has a wide distribution in areas of Cerrado vegetation type restricted sense. Its flowers have a complex pollination mechanism and unique, co-adapted to pollination by moths and butterflies and their fruits are dispersed by mammals large and medium-sized. It has high economic potential and its use has been conducted in an exploratory way. To implement conservation programs, breeding and enabling the commercial use of species, knowledge about the genetic variability, the spatial genetic structure, the reproductive system and gene flow in natural populations are needed. In this context, in the present study was performed the molecular characterization of the genetic variability in three stages of individuals (adults, juveniles and seedlings) and the assessment of genetic structure spatial (adults and juveniles), of the system of crossing and of the gene flow via pollen in a subpopulation of Hancornia speciosa located in the State Park of Serra de Jaragua, Jaragua-GO. To this end, 113 adults and 100 juveniles were sampled and georeferenced in an area of approximately 2.5ha. Of the total number of adults, 20 trees were selected matrices to obtain seedlings and formation of families of open pollination. Genomic DNA was obtained from the leaves of all individuals (adults, juveniles and seedlings) and was amplified with the use of seven microsatellite loci for obtaining of genotypes. The analyzes of genetic diversity, of spatial structure, rates of cross-fertilization and distance of dispersal of pollen were obtained from these genotypes. The total number of alleles at seven loci evaluated was 125, with an average of 17.8 alleles per locus. For adults the mean allele was 15.8, for the juveniles was 13.5and the seedlings were 11alleles. The average total values of heterozigosidade expected (He) and observed (Ho) were equal to 0.750 and 0.698, respectively. In adults He = 0.750 and Ho =0.714, in juveniles He =0.744 and Ho =0.679 and in seedlings He = 0. 712 and Ho =0. 763. These values indicate that the subpopulation evaluated presents high levels of genetic diversity. The fixation index(f) waspositive and significantforthe generations ofadults(0.052, p <0.05)andjuvenile(0.087, p <0.05), indicating the existence of inbreeding in this subpopulation. The analysis of spatial autocorrelation evidenced that kinship is weakly related to the geographical distance in in bothstagesof lifeevaluated (adults: b= -0.00223, R2 = 0. 000514, p < 0.05 and juveniles: b: - 0.00440, R2 = 0.00148489; p < 0.001).Concomitantly the values of Sp were low and the size of neighborhoods (Nb) were high for the two generations. This result shows that there is no restriction of gene flow via seed and corroborates the hypothesis that the dispersal by animals have high potential to disperse the seeds over long distances.The rates of crossfertilization multilocus (tm= 1.000) and single locus (ts = 0.972 a 1.29) were high and significantly different from zero in all families. The difference in the rate of the crossfertilization multilocus and single locus combined for all families analyzed (tm-ts = 0. 077) was also positive and significant, suggesting that 7% of crossings that occur in this population are between related individuals. The correlation of selfing negative(rs=-0.999), indicates absence of selfing and the correlation of paternity(rp=0.107) not significantly different from zero (SD = 0.135) shows that this subpopulation no full siblings. Every subpopulation resulting outcrossing and individuals are evaluated relatives at least to the 15 level of half-brothers. The paternity analysis assigned pollen donors to 75.2% (64) seedlings, and 24 (37.5%) assignments at a confidence level of 85%, 30 (46.8%) assignments at 95% probability assignments and 10(15.6%) at a confidence level of 99%.This low assignment can be explained by sampling: due to the size of the total area of the population and distribution in aggregate, many individuals may not have been sampled. Moreover, the loci battery used in this analysis could not demonstrated the optimal values for the combined exclusion. Although the maximum distance of pollination has been of 292m and covers the entire area evaluated, most events of pollination (77%) occurred at distances less than 200m. The flowering in mass, the distribution in aggregate and the floral structure are the main responsible for predominance of events of cross-pollination at short distances. / Hancornia speciosa (Apocynaceae) é uma espécie frutífera que apresenta distribuição ampla em áreas de fitofisionomia de Cerrado sentido restrito. Suas flores possuem um mecanismo de polinização complexo e único, adaptado á polinização por mariposas e borboletas. Seus frutos são dispersos por mamíferos de grande e médio porte. Possui alto potencial econômico e sua utilização tem sido realizada de forma exploratória. Para implementar programas de conservação e melhoramento, conhecimentos a cerca da variabilidade genética, da estrutura genética espacial, do sistema reprodutivo e do fluxo gênico nas populações naturais são necessários. Nesse contexto, no presente estudo foi realizada a caracterização molecular da variabilidade genética em três estágios de indivíduos (adultos, juvenis e plântulas) e a avaliação da estrutura genética espacial (adultos e juvenis), do sistema de cruzamento e do fluxo gênico via pólen em uma subpopulação de Hancornia speciosa localizada na região do Parque Estadual da Serra de Jaraguá, Jaraguá-GO. Para tanto, 113 indivíduos adultos e 100 juvenis foram amostrados e georeferenciados em uma área de aproximadamente 2,5 ha. Do total de adultos, 20 árvores matrizes foram selecionadas para obtenção de plântulas e formação das famílias de polinização aberta. O DNA genômico foi obtido a partir das folhas de todos os indivíduos (adultos, juvenis e plântulas) e foi amplificado com o uso de sete locos microssatélites para obtenção dos genótipos. As análises de diversidade genética, de estrutura espacial, taxas de fecundação cruzada e distância de dispersão de pólen foram obtidas a partir desses genótipos. O número total de alelos nos sete locos avaliados foi de 125, com média de 17,8 alelos por loco. Para os indivíduos adultos a média de alelos foi de 15,8, para os juvenis foi de 13,5 e nas plântulas foi de 11 alelos. Os valores totais médios de heterozigosidade esperada (He) e observada (Ho) foram iguais a 0, 750 e 0, 698. Nos adultos He = 0, 750 e Ho =0, 714, nos juvenis He =0, 744 e Ho =0, 679 e nas plântulas He = 0, 712 e Ho =0, 763. Esses valores indicam que a subpopulação avaliada apresenta altos níveis de diversidade genética. O índice de fixação (f) foi positivo e significativo para as gerações de indivíduos adultos (0, 052; p < 0, 05) e juvenis (0, 087; p < 0, 05), indicando a existência de endogamia nessa subpopulação. A análise de autocorrelação espacial evidenciou que o parentesco está fracamente relacionado com a distância geográfica em ambas as gerações de indivíduos avaliadas (adultos: b= -0, 00223, R2 = 0, 000514, p < 0,05 e juvenis: b: - 0, 00440, R2 = 0, 00148489; p < 0, 001). Concomitantemente os valores de Sp foram baixos e o tamanho das vizinhanças (Nb) foram altos para as duas gerações. Esse resultado mostra que não existe restrição ao fluxo gênico via semente e corrobora com a hipótese de que a dispersão por animais tem alto potencial para dispersar as sementes a longas distâncias. As taxas de cruzamento multiloco (tm = 1, 000) e uniloco (ts =0, 972 a 1, 29) foram altas e significativamente diferentes de zero em todas as famílias. A diferença da taxa de fecundação cruzada multiloco e uniloco combinada para todas as famílias analisadas (tm-ts = 0, 077) também foi positiva e significativa, sugerindo que 7% dos cruzamentos que ocorrem nessa população são entre indivíduos aparentados. A correlação de autofecundação negativa (rs = -0,999) indica ausência de autofecundação e a correlação de paternidade (rp= 0,107) não significativamente diferente de zero (SD = 0,135) mostra que 13 nessa subpopulação não existem irmãos completos. Toda a subpopulação é resultante de fecundação cruzada e os indivíduos avaliados são parentes no mínimo ao nível de meiosirmãos. A análise de paternidade atribuiu doadores de pólen para 75,2 % (64) plântulas, sendo 24 (37,5 %) atribuições ao nível de confiança de 85%, 30 (46,8%) atribuições ao nível de 95% de probabilidade e 10 atribuições, 15,6% ao nível de confiança de 99%. Essa baixa atribuição pode ser explicada pela amostragem: devido ao tamanho da área total da população e a distribuição em agregado, muitos indivíduos podem não ter sido amostrados. Além disso, a bateria de locos usada nessa análise não apresentou os valores considerados ótimos de probabilidade de exclusão combinada. Embora a distância máxima de polinização tenha sido de 292m e abrange toda a área avaliada, a maior parte dos eventos de polinização (77%) ocorreu a distâncias inferiores a 200m. A floração em massa, a distribuição em agregado e a estrutura floral são os principais responsáveis pelo predomínio de eventos de polinização cruzada a curtas distâncias.

L’intérêt pour la dispersion des gènes via le pollen a augmenté avec les cultures de plantes génétiquement modifiées. A ce jour, les données expérimentales ainsi que la modélisation portant sur les mouvements du pollen de colza, Brassica napus L., à l’échelle du paysage ne différencie pas clairement la part du vent et des insectes dans cette dispersion. Cependant, l’estimation de la dispersion des gènes par le pollen reste une condition nécessaire pour la gestion des risques d’échappement des (trans-)gènes vers l’environnement et les cultures conventionnelles. A travers cette thèse, nous avons pu mettre en évidence qu’une grande diversité d’insectes pollinisateurs pouvait transporter du pollen viable entre différentes plantes de colza sur des distances importantes (>1.1 km). La diversité d’insectes varie d’une région à l’autre et d’une année sur l’autre. Cependant, bien que la majorité des insectes sur une zone de grande production de colza ait du pollen de cette culture sur leur corps, seulement 39,4 % des insectes capturés sur des plantes mâle-stériles transportent du pollen de colza viable. Bien que nous n’ayons pas pu déterminer avec précision la part du vent et des insectes dans le pollinisation du colza, il semblerait que les insectes participent de façon plus importante à la pollinisation de plantes présentes en bordures de champs, augmentant ainsi le taux de pollinisation croisée. Nos résultats fournissent des données fiables pour améliorer les modèles de dispersion pollinique pour des cultures entomophiles à l’échelle du paysage. Ces modèles sont essentiels pour l’aide à la gestion afin de réduire la dispersion des gènes par le pollen des cultures génétiquement modifiées vers les plantes sauvages ou les cultures conventionnelles / Interest in pollen-borne gene dispersal has grown with the cultivation of genetically modified plants. To date both experimental data and models of oilseed rape (OSR) Brassica napus pollen movement at the landscape scale do not clearly differentiate between wind- and insect-mediated dispersal. Estimations of pollen-borne gene dispersal would be valuable for managing potential escapes of transgenes. Our study provides clear evidence that a large variety of insect species can transfer viable pollen between oilseed rape plants over considerable distances (>1.1 km). Insect’s diversity according to geographical site and years. However, the majority of pollinator have OSR pollen in their body hairs, only 39.4% of the insects caught on male-sterile flowers carried OSR pollen. Although we could not determine with precision the role of the wind and the insects in the OSR pollination, it would seem that insects take part in a more important way in pollination of plants present in edges of fields, thus increasing cross pollination rate. Our results provide valuable data to improve models of pollen dispersal for entomophilous crops at the landscape scale. These models are essential to help land-managers reduce pollen-borne gene dispersal from genetically modified plants to wild relative and field planted with non-GM crops

Plants often occur aggregated into clusters and this spatial pattern is supposed to affect pollinator behaviour and pollen dispersal. Such pollinator reaction may influence reproductive success of zoogamous plant species both in terms of number of available mates and their genetic diversity (nearby growing plant individuals are also often closely related, especially in clonal plants). In the present thesis, I investigated the influence of plant spatial aggregation on pollinator behaviour and how this translates into pollen transfer. For that purpose, I carried three experiments. In the Experiment 1, I used potted plants placed into arrays and aggregated into four patches in order to track the pollen dispersal by means of a UV-dye pollen analogue. I manipulated distances between plants within clusters (dense × loose) and between clusters (near × far). I conducted this experiment for three plant species differing in their pollinator spectra. In the Experiment 2, I observed pollinator foraging sequences (sequences of visited plant individuals) under the same experimental design as for the first experiment, but I carried out this experiment for five plant species. In addition in one study species, Dianthus carthusianorum, I conducted the Experiment 3 to get better insight into pollination effectiveness...