O déficit hídrico decorrente da seca se estabelece quando a absorção de água pelas raízes não consegue atender as demandas da planta, entre as quais estão a fotossíntese e transpiração, essenciais ao crescimento vegetal. Para avaliar a influência de déficits hídricos de diferentes intensidades nas relações hídricas, trocas gasosas, crescimento e acúmulo de carboidratos de Bauhinia forficata Link, plantas foram cultivadas em casa de vegetação durante três meses em vasos de 20 L e submetidas aos seguintes regimes: regas diária (controle) e regas a cada 7 (7D) e 15 dias (15D) retornando as regas diárias nos regimes 7D e 15D aos 75 dias de experimento. Em intervalos quinzenais (15, 30, 45, 60, 75, 90 dias) foram avaliados a umidade do solo (Usolo), potencial hídrico foliar (wf), a fotossíntese em resposta a radiação fotossinteticamente ativa (AxPAR) para obtenção da assimilação liquida máxima (Amax) e do ponto de saturação luminoso (PARsat), área foliar total (AFT) e massas secas (MS) de folhas (MSf), caules (MSc) e raízes (MSr). Foi também coletado material vegetal para as análises bioquímicas quantitativas dos teores de açúcares solúveis totais (AST), redutores (AR), amido (S) e prolina (Pro) por métodos colorimétricos e qualitativos da composição de carboidratos solúveis neutros por HPAEC/PAD e análise de açúcares alcoóis por GC/MS. O déficit hídrico afetou as relações hídricas, fotossíntese e crescimento das plantas, sendo que os menores valores de Usolo observados nos tratamentos 7D e 15D respectivamente, coincidiram com os menores wf, Amax e acúmulo de MS. Alterações nos PARsat em resposta ao déficit hídrico foram observadas apresentando valores médios de 665, 275 e 254 mol fótons m-2 s-1 no Controle, 7D e 15D respectivamente. O retorno das regas diárias após 75 dias de experimento, promoveu a recuperação de Amax (7,8 e 9,6 mol CO2 m-2 s-1) e dos PARsat (588 e 643 mol fótons m-2 s-1) das plantas 7D e 15D respectivamente aos 90 dias, com valores maiores daqueles observados nas plantas controle (4,7 mol CO2 m-2 s-1 e 631 mol fótons m- 2 s-1), sugerindo forte dependência da fotossíntese de B. forficata à disponibilidade hídrica no solo. Os baixos teores de amido em folhas nos tratamentos 7D e 15D podem explicar a manutenção dos teores de AST semelhante ao controle, bem como, o aumento dos ASTs após o retorno das regas diárias, os quais poderiam ser prontamente utilizados na osmorregulação e concomitante retomada do crescimento das plantas. A alta detecção de mio-inositol nas análises de HPAEC/PAD sugerem que os açúcares alcoóis podem constituir grande parte dos ASTs não redutores em folhas e raízes de B. forficata. Nas raízes, os aumentos na concentração de glicerol, galactinol e galactosilglicerol detectados por GC/MS seguem o gradiente de disponibilidade hídrica dos tratamentos, indicando uma função osmorreguladora destes açúcares em B. forficata. / The water deficit caused by drought occurs when root water uptake cannot attend plant demands to maintain photosynthesis and transpiration, two essential processes for plant growth. Aiming to evaluate the influence of water deficits of different intensities on water relations, gas exchange, growth, and carbohydrate accumulation, potted plants of Bauhinia forficata Link were grown under greenhouse conditions during three months and submitted to the following water regimes: daily irrigated (control) and irrigated every 7 (7D) or 15 days (15D), recovering the daily irrigations after 75 days of the beginning of the experiment. At fortnightly intervals (15, 30, 45, 60, 75, 90 days) we evaluated soil moisture (Usolo), leaf water potential (wf), photosynthesis in response to photosynthetically active radiation (AxPAR) to obtain the maximum net assimilation (Amax) and light saturation point (PARsat), total leaf area (AFT) and dry mass (MS) of leaves (MSf), stems (MSc) and roots (MSr). Leaves and roots were also sampled for analyses of total soluble sugars (AST), reducing sugars (AR), starch (S), and free proline (Pro) by colorimetric methods. The analyses of the soluble sugar composition were carried out by HPAEC/PAD and GC/MS. The imposed water stress affected the water relations, photosynthesis, and plant growth. Decreases in Usolo values observed in the treatments 7D and 15D, respectively, coincided with the lowest wf, Amax and accumulation of MS. Changes in PARsat in response to water deficit showed mean values of 665, 275, and 254 mol photons m-2 s-1 in control, 7D, and 15D, respectively. The re-watering after 75 days led to the recovery of Amax (7.8 and 9.6 mol CO2 m-2 s-1) and PARsat (588 and 643 mol photons m-2 s-1) of plants 7D and 15D, respectively, at 90 days, with higher values than those observed in the control plants (4.7 mol CO2 m-2 s-1 e 631 mol photons m-2 s-1), suggesting a strong dependence of the photosynthesis of B. forficata of the soil water availability. The low leaf starch contents in the 7D and 15D treatments could explain the maintenance of the concentration of AST similar to control, as well as the increase in AST contents after re-watering. These sugars could be promptly used for osmoregulation and concomitant re-start of plant growth. The presence of high amounts of of myo-inositol detected by HPAEC/PAD analysis suggests that sugar alcohols can constitute the major part of non-reducing ASTs in leaves and roots of B. forficata. In roots, increases in the contents of glycerol, galactinol, and galactoglycerol detected by GC/MS indicating that these sugars can play a role in the osmoregulation of B. forficata.

As espécies Cattleya labiata Lindley e a Cattleya eldorado Linden, nativas da Mata Atlântica e Amazônia, respectivamente, encontram-se sob forte risco de desaparecimento na natureza, sendo, portanto, necessário o desenvolvimento de métodos eficientes de propagação de suas mudas, dentre os quais se destaca a micropropagação. A eficiência do cultivo in vitro de orquídeas está associada aos meios de cultura utilizados e, principalmente, aos reguladores de crescimento. O trabalho teve como objetivo avaliar a influência de paclobutrazol (PBZ) no desenvolvimento das microplântulas cultivadas in vitro. Plântulas provenientes de germinação in vitro, com 1,0 ± 0,2 cm de comprimento, foram utilizadas no estudo. No primeiro experimento, utilizou-se como controle o meio de cultura MS isento de reguladores de crescimento vegetal. Nos tratamentos, o meio MS foi suplementado com quatro concentrações de PBZ 1,0; 2,0; 4,0 e 6,0 mg.L-1; com ácido naftalenoacético (ANA) 1,0 mg.L-1 associado a 6-benzilaminopurina (BAP) 1,0 mg.L-1 e com a combinação de 4,0 mg.L-1 PBZ, com 1,0 mg.L-1 ANA e 1,0 mg.L-1 BAP. As avaliações morfológicas, número de brotos, comprimento da parte aérea (CPA), comprimento da maior raiz (CMR) e comprimento da lâmina foliar (CLF), foram realizadas no início do experimento e nos subcultivos a cada 40 dias, durante 160 dias. O delineamento experimental foi inteiramente casualizado em arranjo trifatorial com parcelas subdivididas no tempo, com seis repetições por tratamento. Um segundo experimento foi realizado utilizando a técnica de pulse. Como controle foi utilizado meio de cultura MS, e para os tratamentos foi utilizado o mesmo meio acrescido de concentrações crescentes de PBZ com diferentes tempos de exposição, sendo: T0= controle (MS); T1= 1,0 mg.L-1 PBZ por 40 dias; T2= 10,0 mg.L-1 PBZ por 4 dias e T3= 100,0 mg.L-1 PBZ por 4 horas. Foram feitas as avaliações de CPA, CLF, CMR, número de raízes, massa fresca e massa seca da parte aérea, massa fresca e massa seca de raiz, massa fresca e massa seca total, contagem e caracterização estomática e avaliação anatômica do tecido radicular. O experimento foi conduzido no delineamento inteiramente casualizado totalizando quatorze repetições por tratamento. O tratamento com 1,0 mg.L-1 de PBZ por 40 dias promoveu maior desenvolvimento e vigor do sistema radicular, aumentando a espessura das raízes, podendo contribuir para a fase de aclimatização das microplântulas. Os tratamentos com pulse reduziram significativamente a parte aérea das microplântulas, não sendo recomendado na micropropagação da C. labiata. / The Species Cattleya labiata Lindley and Cattleya eldorado Linden, native to the Atlantic Forest and Amazon, respectively, are under high risk of extinction in nature, therefore, necessary to develop efficient methods of spreading their seedlings, among which highlights micropropagation. The efficiency of in vitro cultivation of orchids is associated with culture media, and especially the growth regulators. The study aimed to evaluate the influence of paclobutrazol (PBZ) in the development of in vitro microplantlets. Seedlings from germination in vitro, with 1.0 ± 0.2 cm in length, were used in the study. In the first experiment, we used to control the MS culture medium free of plant growth regulators. In the treatments, MS medium was supplemented with four concentrations of PBZ 1.0, 2.0, 4.0 and 6.0 mg L-1, with naphthaleneacetic acid (NAA) 1.0 mg L-1 associated with 6-benzylaminopurine (BAP) 1.0 mg L-1 and with the combination of 4.0 mg L-1 PBZ with 1.0 mg.L-1 NAA and 1.0 mg L-1 BAP. The morphological evaluations, number of shoots, shoot length (CPA), longest root length (CMR) and leaf blade length (CLF) were performed at the beginning of the experiment and in subcultures every 40 days during 160 days. The experimental design was completely randomized in three-factor split-plot arrangement in time, with six replicates per treatment. A second experiment was performed using the technique of \"pulse\". It was used as control MS medium, and the treatments we used the same medium plus increasing concentrations of PBZ with different exposure times, as follows: T0 = control (MS), T1 = 1.0 mg L-1 PBZ for 40 days, T2 = 10.0 mg L-1 PBZ for 4 days and T3 = 100.0 mg L-1 PBZ for 4 hours. Assessments were made CPA, CLF, CMR, number of roots, fresh and dry mass of shoots, fresh weight and dry weight of root, fresh weight and total dry matter, stomatal counting and characterization and evaluation of the anatomic root tissue. The experiment was conducted in a completely randomized a total of fourteen replicates per treatment. Treatment with 1.0 mg L-1 of PBZ for 40 days promoted further development and vigor of the root system, increasing the thickness of the roots and may contribute to the acclimatization phase of microplantlets. Treatments with \"pulse\" significantly reduced the shoot microplantlets and Its not recommended to the micropropagation of C. labiata.

As orquídeas são a maior família das plantas monocotiledôneas, sendo o Brasil um dos países contém grande diversidade de espécies. As orquídeas são, em sua maioria, alógamas e possuem mecanismos sofisticados para evitar a autopolinização. Os insetos são os agentes polinizadores mais comuns, mas também podem ser polinizados por aves como beija-flores. Tradicionalmente as espécies Cattleya coccinea e C. mantiqueirae tem sido reconhecidas como distintas de acordo com caracteres morfológicos, distribuição geográfica e época de floração. Esses critérios, entretanto, não permitem a identificação clara de espécies, uma vez que muitos indivíduos apresentam morfologia e fenologia intermediárias. Nesse contexto, esse estudo tem como objetivo contribuir para o conhecimento taxonômico e evolutivo de orquídeas brasileiras do gênero Cattleya. Especificamente, propõe-se rever a atual delimitação entre as espécies C. coccinea e C. mantiqueirae e caracterizar a diversidade genética entre e dentro de populações dessas espécies a partir de marcadores moleculares ISSR. Para testar se a atual delimitação de espécies corresponde a linhagens filogenética distintas, foram realizadas coletas em seis localidades da região Sudeste. Foram testados 20 iniciadores ISSR, dos quais 13 foram otimizados para obtenção de dados. Os géis de ISSR obtidos foram utilizados para construção de uma matriz binária representando a presença/ausência de fragmentos amplificados. A matriz contendo 173 indivíduos e 295 caracteres foi analisada com algoritmo de neighbor-joining e o critério de parcimônia máxima para obtenção de hipóteses filogenéticas. Os resultados indicam que as espécies tradicionalmente reconhecidas, C. coccinea e C. mantiqueirae, não constituem grupos monofiléticos e, portanto, não podem ser reconhecidas como espécies distintas de acordo com o conceito filogenético de espécies. Os resultados também apontam que as populações amostradas constituem grupos monofiléticos com altos valores de confiança e que o complexo C. coccinea-C. mantiqueirae não constitui um grupo monofilético. O parafiletismo do grupo é determinado pela posição da população de Lima Duarte/MG, que constitui um clado irmão da espécie C. brevipedunculata (ocorrente na Serra do Espinhaço) e C. wittigiana (restrita ao Estado do Espírito Santo). Os resultados de análises de genética de populações corroboram com os resultados da análise filogenética e indicam que as populações possuem baixos índices de diversidade genética entre indivíduos e que a maior diversidade encontra-se entre populações. Por serem plantas com alto valor ornamental e sofrerem com ações antrópicas constantes, esse estudo foi de fundamental importância para permitir estratégias viáveis para a manutenção e conservação da diversidade genética dessas populações de orquídeas. / Orchids represent the largest family of monocots, with great diversity of species in Brazil. These plants are generally allogamous and bear sofisticated mechanisms to avoid self-pollination. Insects are by far the most common pollinators, but birds (i.e hummingbirds) may also be important. Within Cattleya, the species C. coccinea and C. mantiqueirae have been distinguished by morphological characters, geographical distribution and flowering period. Such criteria, however, do not allow a clear identification of species, since many specimens show intermediate morphological and phenological variation. The goal of this study is to contribute to the understanding of taxonomical and evolutionary aspects of Brazilian orchids, especially within the genus Cattleya. In order to achieve that I revised current species limits within the C. coccinea-C. mantiqueirae species complex. The study was based on phylogenetic and genetic diversity analyses among and within populations considering ISSR molecular markers. Six populations from Southeastern Brazil were considered. I tested 20 ISSR primers, of which 13 were used in this study. Presence/absence of fragments visualized in agarose gels were used to built a binary matrix. The analyses considered 173 individuals and 295 caracters (fragments). Phylogenetic analyses were performed according to distance (neigbor-joining) and parsimony criteria. According to the results, the species C. coccinea and C. mantiqueirae do not constitute monophyletic groups and, therefore, cannot be recognized as distinct according to the phylogenetic species criterion. Also the C. coccinea-C. mantiqueirae species complex is paraphyletic considering the closely related species C. brevipedunculata (from Serra do Espinhaço) and C. wittigiana (from Espírito Santo State). The population of Lima Duarte/MG is phylogenetically more closely related to such species than to other populations of C. coccinea and C. mantiqueirae. On the other hand, the studied populations comprise strong monophyletic groups. Population genetics analyses agree with phylogenetic results. All populations show low diversity indices among individuals. Also, the greatest portion of genetic diversity was found between populations. Orchids belonging to the C. coccinea-C. mantiqueirae species complex are high ornamental species, with great anthropogenic pressure. For this reason this study was important to allow conservation strategies to maintain and monitor genetic and morphological diversity of populations.

A key question in biology is how changes in gene function or regulation produce new morphologies during evolution. The nectar spur is an evolutionarily labile structure known to influence speciation in a broad range of angiosperm taxa. Here, the genetic basis of nectar spur development, and the evolution of differences in nectar spur morphology, is investigated in Linaria vulgaris and two closely related species of orchid, the primitively longer-spurred Dactylorhiza fuchsii, and more derived short-spurred D. viridis (Orchidinae, Orchidaceae). Despite considerable morphological and phylogenetic differences, nectar spur ontogeny is fundamentally similar in each of the study species, proceeding from an abaxial bulge formed on the ventral petal relatively late in petal morphogenesis. However, spur development is progenetically curtailed in the short-spurred orchid D. viridis. In each case spur development involves class 1 KNOTTED1-like homeobox (KNOX) proteins. KNOX gene expression is not restricted to the spur-bearing petal, indicating that additional components are required to define nectar spur position, e.g. canonical ABC genes, determinants of floral zygomorphy, and additional (currently unknown) factors. However, constitutive expression of class 1 KNOX proteins in transgenic tobacco produces flowers with ectopic outgrowths on the petals, indicating that KNOX proteins alone are, to some degree, capable of inducing structures similar to nectar spurs in a heterologous host. Interestingly, KNOX gene expression is high in the ovary of all study taxa, suggesting that KNOX proteins may also have been involved in the evolution of this key angiosperm feature. Although principally involved in maintaining indeterminacy in the shoot apical meristem (SAM), members of the KNOX gene family have been co-opted in the evolution and development of compound leaves where they suppress differentiation and extend the morphogenetic potential of the leaf. A similar model is presented here to explain the role of KNOX proteins in nectar spur development. Co-option of KNOX gene expression to the maturing perianth delays cellular differentiation, facilitating the development of the nectar spur but requiring additional, unknown factors, to determine nectar spur fate. As facilitators of nectar spur development, changes in the spatio-temporal patterns of KNOX gene expression may alter the potential for nectar spur development and explain the critical length differences observed between the orchids D. fuchsii and D. viridis (and among other angiosperm taxa). Taken together, the available data indicate that KNOX genes confer a meristematic state upon plant tissues in a variety of morphogenetic contexts, making the gene family a potentially versatile tool to mediate a wide variety of evolutionary transformations.

Orientador: Luiz Fernando Rolim de Almeida / Resumo: Orquídeas epífitas estão expostas a diferentes microclimas, que são caracterizados pela alta irradiância imposta ao dossel e baixa irradiância em plantas situadas no sub-bosque. Além disso, este grupo de plantas frequentemente lida com a restrição hídrica. Na produção comercial, o estabelecimento de orquídeas é em quase sua totalidade limitada pela distribuição e intensidade destes mesmos fatores disponibilizados em casas de vegetação. Gomesa flexuosa Sims e Oncidium sphacelatum Lindley pertencem à subtribo Oncidiinae, e quando hibridizadas dão origem a O. ‘Aloha’ (uma orquídea CAM facultativa e importante híbrida comercial). O objetivo deste estudo foi elucidar a performance fotossintética e as estratégias de balanço hídrico de G. flexuosa e O. sphacelatum sob condições de estresse hídrico e luminoso, focando tanto nas diferenças interespecíficas como nas funções dos diferentes órgãos em um mesmo indivíduo. Para isso, analisamos trocas gasosas, fluorescência da clorofila a, quantificação de lipoperóxidos, discriminação isotópica, acidez titulável, conteúdo relativo de água e potencial hídrico em tecidos foliares, de pseudobulbos e raízes, em diferentes tratamentos envolvendo restrição hídrica e variação luminosa. Depois de restrição hídrica por 60 dias, G. flexuosa apresentou conversão de metabolismo C3 para CAM, o que deu subsídios para uma melhor performance das trocas gasosas, fluorescência da clorofila a, mecanismo de fotoproteção e relações hídricas. Quando submetida a ... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Epiphyte orchids are exposed to different microclimates, which are characterized by high irradiance imposed on the canopy and low irradiance in plants located on the understory. In addition, this group of plants frequently deals with water restriction. In commercial production, the distribution and intensity of these same factors inside the greenhouses also limit the orchids establishment. Gomesa flexuosa Sims and Oncidium sphacelatum Lindley are Oncidiinae species and when hybridized, it gave origin to O. ‘Aloha’ (a facultative CAM orchid and na important commercial hybrid). The aim of this study was to elucidate the photosynthetic performance and the water balance strategies of G. flexuosa and O. sphacelatum under water and light stress conditions, focusing on an interspecific diferences as well as on different organs functions in the same individual. For that, we analyzed the gas exchange, chlorophyll a fluorescence, lipoperoxides quantification, isotopic discrimination, titratable acidity, water relative content and water potential of leaf, pseudobulb and root tissues in different treatments involving water restriction and light variation. After 60 days of water restriction, G. flexuosa converted the photosynthetic metabolism from C3 to CAM, which provided a better performance on gas exchanges, chlorophyll a fluorescence, photoprotection mechanism and water relations. When G. flexuosa was submitted to two events of water restriction, it showed a capacity to store informat... (Complete abstract click electronic access below) / Doutor

All orchid species studied so far have been shown to participate in orchideoid mycorrhizal symbiosis. Morover, this symbiosis is absolutely vital component of their life cycle. Exchange of nutrients occurs between symbionts where the fungi provides the orchid with energy and carbon supply at least in its early developmental stages. This study focuses on the possible role of trehalose in this transfer. In vitro experiments have showed in five species from three different subfamilies of Orchidaceae family that they can utilize trehalose comparably with sucrose and glucose. Thus, the ability of trehalose utilization seems to be conserved among orchids. Trehalase enzyme activity was localized histochemically in orchid mycorrhizas. The activity strongly colocalized with colonized tissue supporting a hypothesis that trehalose transfer occurs in this site and is mediated by trehalase. Using bioinformatic methods, trehalase gene duplications were identified in many taxons of Embryophyta including three orchid species. Interestingly, highest number of trehalase gene copies was identified in genome of orchid Dactylorhiza majalis. Trehalose utilization, high trehalase activity in mycorrhizas and trehalase gene duplications in some orchids together indicate that trehalose transfer in orchid myccorhizas...

Perhaps all orchids are mycotrophic at early developmental stages, while majority of species photosynthesize at adulthood and only about 200 species remain fully mycotrophic for the whole life. Mycotrophy affects orchids at many levels. In this thesis, I focus on four aspects of orchid biology, which could be connected with mycotrophy: (i) systematics, (ii) genome size and endoreduplication, (iii) regulation of seed germination and (iv) mechanism of transfer of carbon and energy from fungi to orchids. There are over 27,000 recently recognized orchid species, nevertheless new ones are still discovering and old ones are revisiting. In this work I present a description of new species, Cleisostoma yersinii, and its morphological, anatomical, ecological and systematic characterization. Phylogeny reconstruction confirmed relationship with C. birmanicum. In the subtribe Podochileae, I reappraised the genus Campanulorchis to establish monophyletic but also morphologically defined group. For both abovementioned genera I prepared the artificial identification key. In the genus Dactylorhiza I revised taxa present in our country and I prepared an identification key which firstly mentions D. maculata subsp. elodes from Czech Republic. Orchid species diversity is probably reflected in genome structure. Results...

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Perhaps all orchids are mycotrophic at early developmental stages, while majority of species photosynthesize at adulthood and only about 200 species remain fully mycotrophic for the whole life. Mycotrophy affects orchids at many levels. In this thesis, I focus on four aspects of orchid biology, which could be connected with mycotrophy: (i) systematics, (ii) genome size and endoreduplication, (iii) regulation of seed germination and (iv) mechanism of transfer of carbon and energy from fungi to orchids. There are over 27,000 recently recognized orchid species, nevertheless new ones are still discovering and old ones are revisiting. In this work I present a description of new species, Cleisostoma yersinii, and its morphological, anatomical, ecological and systematic characterization. Phylogeny reconstruction confirmed relationship with C. birmanicum. In the subtribe Podochileae, I reappraised the genus Campanulorchis to establish monophyletic but also morphologically defined group. For both abovementioned genera I prepared the artificial identification key. In the genus Dactylorhiza I revised taxa present in our country and I prepared an identification key which firstly mentions D. maculata subsp. elodes from Czech Republic. Orchid species diversity is probably reflected in genome structure. Results...