The Orchidaceae is characterized by a remarkably diverse range of life forms and some of the most highly specialized interactions with soil fungi and insect pollinators found in the flowering plants. Many species are rare or threatened with extinction either directly through loss of habitat or over-collection or, indirectly through debilitation or loss of mycorrhizal association or pollinator capacity. Australian temperate terrestrial orchids represent one of the most threatened groups in the Australian flora with many taxa clinging to existence in urban and rural bushland remnants, road verges and unprotected bushland. The aim of this study is to research and develop integrated conservation based on critical aspects of terrestrial orchid biology and ecology, towards the recovery of the rare and endangered Western Australian terrestrial orchid Caladenia huegelii. This study identified key aspects involved in an integrated conservation approach and research focused on conservation genetics, mycorrhizal interactions and in situ and ex situ conservation strategies for this species. Using polymorphic microsatellite molecular markers, high levels of genetic diversity were found within remnant populations of C. huegelii, while weak differentiation was observed among populations over the species geographic range. These results indicate historic genetic exchange between C. huegelii populations, a possible consequence of the sexually deceptive pollination strategy and the capacity for widespread seed dispersal. Symbiotic germination studies revealed compatibility barriers to C. huegelii germination with the orchid possessing a highly specific orchid-mycorrhizal association relative to common sympatric congeners. These results were reflected in a phylogenetic analysis of DNA sequences, revealing C. huegelii associates with only one endophyte species within the fungal family Sebacinaceae across its geographic range. Large scale in situ seed baiting demonstrated that endophytes compatible with C. huegelii were limited in distribution relative to common and widespread orchid species, a feature for C. huegelii that may be a major contributing factor in limiting the distributional range of the species. Detailed, within site seed baiting methods identified hotspots for mycorrhizal fungus compatible with C. huegelii that were unoccupied by the orchid. These mycorrhizal hotspots where used to investigate the effect of endophyte presence on survival of transplanted mature plants and seedling outplants. The in situ survival of glasshouse propagated seedlings was further optimized by incubating seedlings in growth containers before transfer to soil and outplanting seedlings in their second growing season. The findings of this study will substantially advance the recovery of C. huegelii and provide benchmark knowledge for similar projects with other rare and threatened terrestrial orchid species.

published_or_final_version / Fine Arts / Master / Master of Philosophy

The Orchidaceae makes up the largest and most diverse family of flowering plants. Orchids are popular, often expensive ornamentals, with a broad range of ethnobotanical applications. There is very limited documented information on South African medicinal orchid species; no formal pharmacopoeia outlining ethnobotanical uses; and ethnobotanical and distribution records are either scarce or inconsistent and plant populations are becoming gradually smaller. There have been significant developments in medicinal orchid research worldwide with medicinal use and corresponding pharmacological and phytochemical properties being extensively investigated. It is evident from the literature that there is no pharmacological research on South African medicinal orchids; hence the need to explore biological activity and chemical composition of South African medicinal orchid species. The ethnobotanical approach used to select the orchid species for pharmacological and phytochemical research elsewhere, yielded valuable biological compounds. Thus, a similar approach was applied to South African medicinal orchids. There are approximately 20 000 species and 796 genera of orchids distributed across the world. In southern Africa, orchids are widely represented with 55 genera and 494 species. Approximately 75% are endemic to this region. As part of the current investigation a review of available ethnobotanical literature on South African medicinal orchids was prepared. The review revealed that an estimated 49 indigenous orchid species from 20 orchid genera are currently being informally traded and used in South African traditional medicine. They are used primarily for medicinal and cultural purposes, especially by the Zulu community in South Africa. Medicinal uses of orchid species include: treatment of inflammatory, intestinal, neurological and reproductive disorders and emetics are used to cause emesis. Non-medicinal uses of orchid species include: love, fertility, protective and lethal charms. Based on their ethnobotanical uses and endemism, South African orchids were considered to be one of the untapped sources of bioactive compounds that needed to be researched. The current investigation addressed the broader aims of medicinal plant research by determining the efficacy, safety and chemical profile of seven indigenous orchid species used in South African traditional medicine and practices. The biological and toxic effects of orchid plant extracts were assessed using established pharmacological bioassays. The phytochemical evaluation of the seven orchid plant extracts provided insight into the classes of chemical compounds present and their possible role in the observed biological activities. The potential of plant extracts from seven orchid species used in South African traditional medicine, as sources of natural bioactive products, are discussed. The current investigation determined the biological activity and chemical profile of seven orchid species commonly traded in KwaZulu-Natal herbal markets: Ansellia africana Lindl., Bulbophyllum scaberulum (Rolfe) Bolus, Cyrtorchis arcuata (Lindl.) Schltr., Eulophia hereroensis Schltr., Eulophia petersii (Rchb.f.) Rchb.f., Polystachya pubescens (Lindl.) Rchb.f. and Tridactyle tridentata (Harv.) Schltr. Well established in vitro micro-dilution bioassays were used to determine the antibacterial, antifungal, anthelmintic activities of crude orchid extracts. A minimum inhibitory and/or lethal effect of organic and aqueous crude orchid extracts was observed against Bacillus subtilis, Escherichia coli, Klebsiella pneumoniae, Staphylococcus aureus, Candida albicans and Caenorhabditis elegans. Tridactyle tridentata aqueous root extract produced the most effective antibacterial activity against S. aureus (0.049 mg/ml). All T. tridentata organic root extracts produced significant inhibitory activities against B. subtilis and S. aureus. Eulophia petersii DCM pseudobulb extracts significantly inhibited all bacterial strains tested (0.39 mg/ml against S. aureus and 0.78 mg/ml against B. subtilis, E. coli, and K. pneumoniae). Eulophia hereroensis 80% EtOH root extract was the only other extract to exhibit significant inhibitory effects against K. pneumoniae (0.65 mg/ml). After 48 h C. albicans was most susceptible to P. pubescens aqueous pseudobulb extract (0.0816 mg/ml). Eulophia petersii DCM pseudobulb extract however, exhibited significant activity against C. albicans (0.65 mg/ml) over 72 h. Cyrtorchis arcuata leaf and root extracts were the most effective anthelmintic extracts with MLCs of 0.041 mg/ml for 80% EtOH leaf and root extracts; 0.049 mg/ml for aqueous leaf extracts and 0.78 mg/ml for aqueous and DCM root extracts. Caenorhabditis elegans was most susceptible to all A. africana and T. tridentata organic root extracts. A similar significant effect was observed for all E. petersii organic pseudobulb extracts, DCM extracts and organic root extracts of B. scaberulum. Only the DCM tuber and root extracts of E. hereroensis exhibited lethal effects on C. elegans. All of the P. pubescens extracts showed poor anthelmintic activity. Similarly, in vitro enzyme based cyclooxygenase (COX) 1 and 2 and acetylcholinesterase (AChE) inhibitory bioassays, revealed significant inhibition of COX-1, COX-2 and AChE enzymes by crude organic and certain aqueous orchid extracts. Out of a total of 53 evaluated extracts, 21 and 13 extracts exhibited significant anti-inflammatory activity in the COX-1 and COX-2 assays respectively. The DCM tuber extract of E. hereroensis was the only extract to significantly inhibit both COX enzymes, 100.02±0.11% and 87.97±8.38% respectively. All B. scaberulum root extracts (DCM, EtOH and water) exhibited COX-2 selective inhibitory activity (100.06±0.01, 93.31±2.33 and 58.09±3.25%). Overall, the DCM root extract of A. africana was found to be the most potent extract (EC50 0.25±0.10 mg/ml). The 80% EtOH root extract of B. scaberulum was the most potent in the COX-2 assay (EC50 0.44±0.32 mg/ml). Generally the root extracts exhibited greater AChE inhibitory activity; where the most active extract was B. scaberulum DCM root extract (EC50 0.02±0.00 mg/ml). All aqueous extracts, except that of A. africana roots and B. scaberulum pseudobulbs, showed poor or no COX-1 and COX-2 inhibition. The antioxidant capacity of crude orchid extracts was determined using: hydrogen atom transfer (HAT) (β-carotene/linoleic acid assay) and single electron transfer (SET) (2,2‟-diphenylpicrylhydrazyl (DPPH) free radical scavenging assay and ferric reducing antioxidant power (FRAP) assay) reaction-based assays. Potent antioxidant effects were observed for certain crude methanolic orchid extracts. Generally, there was a dose-dependent change in radical scavenging activities of crude extracts from which EC50 values were determined. The root extracts of all species, except that of E. petersii, had consistently more effective radical scavenging activity than that of other plant parts within each species. The pseudobulb extract of E. petersii, was the most potent extract (EC50 1.32±0.86 mg/ml). In the β-carotene-linoleic acid assay, based on the oxidation rate ratio (ORR), the leaf extract of T. tridentata and the root extracts of C. arcuata and E. hereroensis exhibited the best antioxidant effects (0.02, 0.023 and -0.15 respectively). Similarly, the average antioxidant activity (%ANT) of these samples was greater than that of BHT (95.88±6.90%) and all other samples. Bulbophyllum scaberulum leaf, pseudobulb and root extracts, E. petersii pseudobulb extract and T. tridentata root extract also exhibited a greater capacity to prevent β-carotene oxidation when compared to BHT. All crude orchid extracts tested demonstrated a general dose-dependent response in the ferric reducing power assay. The reducing power of ascorbic acid (0.08 mM) and BHT (0.05 mM), as measured as absorbance, was 1.12±0.12 and 0.73±0.08 respectively. At 6.25 mg/ml, A. africana root and E. petersii pseudobulb extracts were the most effective in reducing power activity. The short-term bacterial reverse mutation Ames Salmonella/microsome mutagenicity (ASMM) assay, which makes use of mutant histidine-dependent Salmonella typhimurium strains, was used to determine the mutagenicity and toxicity of crude orchid extracts. In the presence of a mutagen S. typhimurium TA98 strain detects frameshift events while the TA100 and TA102 strains detect base-pair substitutions. In the absence of metabolic activation, mutagenic extracts were observed against the TA98 strain only. All A. africana DCM leaf and stem extracts tested, the DCM root extract (0.5, 0.05 mg/ml) and EtOH leaf, stem and root extracts at 5 mg/ml exhibited mutagenic effects. The EtOH root extracts (5, 0.5 mg/ml) of B. scaberulum exhibited mutagenic indices (MI) comparable to that of 4NQO (17.00 and 13.00, respectively). Eulophia petersii PE pseudobulb extract demonstrated mutagenic potential at 5 mg/ml. The ethanolic root extracts of T. tridentata showed mutagenic effects at 5 and 0.5 mg/ml. The mutagenicity index (MI) with metabolic activation (S9) was determined using only the TA98 strain; where no mutagenic effects were observed. In the phytochemical evaluation of crude methanol orchid extracts, the Folin-Ciocalteu assay for total phenolics, butanol-HCl assay for condensed tannins, rhodanine assay for gallotannins and vanillin assay for flavonoids revealed a quantitative chemical profile of the tested samples. The correlation between observed biological effects and chemical compounds present was found to be generally significant. The significant antimicrobial, anthelmintic, anti-inflammatory and antioxidant activity of E. petersii pseudobulb extracts and E. hereroensis tuber and root extracts may be attributed to their high total phenolic content. Alternatively, the significant levels of gallotannin content in E. hereroensis may have contributed to the bioactivity. The flavonoid content of B. scaberulum and T. tridentata may explain the potent activity observed in the anti-inflammatory, antioxidant and acetylcholinesterase inhibitory assays; while the flavonoid content C. arcuata may have contributed to the potent anthelmintic and antioxidant activities. The significantly higher levels of gallotannin content may explain the significant anti-inflammatory and anthelmintic activity of A. africana. A number of biologically active compounds have been isolated from certain Orchidaceae species around the world on the basis of their traditional medicinal uses. The traditional uses of these orchid species were scientifically validated. No pharmacological research has been previously conducted on South African medicinal orchids; therefore the current investigation has produced novel findings on the efficacy and safety of these orchid species and promotes the continued research of medicinal orchids in South Africa. / Thesis (Ph.D.)-University of KwaZulu-Natal, Pietermaritzburg, 2012.

The point of that experiment was checking on status of selected areas with appearance of Dactylorhiza majalis and Dactylorhiza fuchsii around České Budějovice in south Bohemia. On 4 areas there were finding of biometric charakteristics always 40 selected plants and theirs mutual correlation in fact of more detailed knowing of status of individual population those kinds in particular area. There was always completed also phytocoenotic characteristics of the area. The graduation theses also includes updated information about few others tenths of areas of different kinds of orchids in České Budějovice and Český Krumlov area. Part of the experiment is map display of areas of the orchids. In map there are picture only areas with GPS not others.

Submitted by (edna.saturno@ufrpe.br) on 2016-07-06T11:28:13Z No. of bitstreams: 1 Maiza de Azevedo Dantas.pdf: 1426194 bytes, checksum: 248ab81985fd9f5b4bd3a17adefea341 (MD5) / Made available in DSpace on 2016-07-06T11:28:13Z (GMT). No. of bitstreams: 1 Maiza de Azevedo Dantas.pdf: 1426194 bytes, checksum: 248ab81985fd9f5b4bd3a17adefea341 (MD5) Previous issue date: 2014-02-06 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / Orchids stand out among the ornamental tropical due to the exotic beauty of flowers, exuberant colors and shapes. Due to the high market demand, depredation of habitat, low rate of seed germination in natural environment and slow growth, tissue culture has been presented as an important tool for the production of seedlings of orchids on a large scale, in a reduced period of time and with greater seedling quality over conventional methods. The polysaccharide chitosan has been tested as an additive in the culture medium of some species as it stand out as a growth promoter and by acting as an antioxidant. This study aimed to evaluate the influence of dietary supplementation of chitosan to the nutrient medium in vitro cultivation of Dendrobium Phalaenopsis and Cattleya labiata in liquid and solid medium. Shoots from in vitro seedlings of both species were inoculated into vials containing 30 ml of MS medium with half the ionic strength. The experiment consisted of the combination of three chitosan concentrations (0, 15 and 20 mg.L-1) with nutrient medium in solid and liquid state. Each treatment had four replicates, each with two plants. The experimental design was completely randomized with a factorial scheme of 3x2 (chitosan concentration x physical state of the medium). For 50 days were evaluated the total fresh weight, number of shoots and roots, and were determined activities of superoxide dismutase enzyme (SOD), ascorbate peroxidase (APX) and catalase (CAT). In cultivation in liquid medium, D. phalaenopsis fresh weight showed higher treatment with 20 mg.L-1 of chitosan as the highest number of shoots was observed in both media, at the same concentration. C. labiata also exhibited higher fresh weight and number of shoots in liquid medium, but at the concentration of 15 mg.L-1 of chitosan. The solid medium favored the emission of roots in both species, but inhibited the formation of shoots of C. labiata regardless of the chitosan concentration. The best development of seedlings of both species in liquid medium coincided with increased activity of SOD directly proportional to the chitosan. This activation of SOD was not accompanied by an increase in the activity of APX or CAT, showing the action of chitosan in the dismutation of superoxide without forming hydrogen peroxide. On solid medium, the action of chitosan on the antioxidative mechanism is also provided by the activation of the APX and CAT. Verified the prevalence of the liquid medium for micropropagation of the species studied and confirmed the combination of chitosan in controlling levels of reactive oxygen species in joint action with the antioxidant enzyme system action. / As orquídeas destacam-se entre as ornamentais tropicais, devido à beleza exótica das flores, exuberância de cores e formas. Em virtude da alta demanda do mercado, depredação do habitat, baixa taxa de germinação das sementes em ambiente natural e o lento crescimento, a cultura de tecidos vem se apresentando como uma importante ferramenta para a produção de mudas de orquídeas em larga escala, em período reduzido de tempo e com maior qualidade das mudas em relação aos métodos convencionais. O polissacarídeo quitosana tem sido testado como aditivo no meio de cultura de algumas espécies por se destacar como promotor de crescimento e por sua ação como antioxidante. O presente trabalho teve por objetivo avaliar a influência da suplementação de quitosana ao meio nutritivo no cultivo in vitro de Dendrobium phalaenopsis e Cattleya labiata, em meio líquido e sólido. Brotações provenientes de mudas in vitro de ambas as espécies foram inoculadas em frascos com 30 ml de meio MS com metade da força iônica. O experimento constou da combinação de três concentrações de quitosana (0, 15 e 20 mg L-1 de quitosana) em meio nutritivo em estado sólido e líquido. Cada tratamento contou com quatro repetições, cada uma com duas plantas. O delineamento experimental foi inteiramente casualizado em esquema fatorial de 3x2 (concentrações de quitosana x estado físico do meio). Aos 50 dias avaliaram-se o peso fresco total, número de brotos e de raízes, e foram determinadas as atividades das enzimas superóxido dismutase (SOD), ascorbato peroxidase (APX) e catalase (CAT). No cultivo em meio líquido, D. phalaenopsis apresentou maior peso fresco no tratamento com 20 mg.L-1 de quitosana, enquanto o maior número de brotos foi observado em ambos os meios, nessa mesma concentração. C. labiata também exibiu maior peso fresco e número de brotos em meio líquido, porém na concentração de 15 mg.L-1 de quitosana. O meio sólido favoreceu a emissão de raízes das duas espécies, mas inibiu a formação de brotos de C. labiata, independente das doses de quitosana. O melhor desenvolvimento das mudas das duas espécies em meio líquido coincidiu com aumento da atividade de SOD diretamente proporcional ao da concentração de quitosana. Essa ativação da SOD não foi acompanhada pelo aumento na atividade da APX nem da CAT, evidenciando a ação da quitosana na dismutação do superóxido sem formação de peróxido de hidrogênio. Em meio sólido, a ação da quitosana no mecanismo antioxidativo se deu pela ativação da APX e CAT. Constatou-se a prevalência do meu líquido para a micropropagação das espécies estudadas e confirmou-se a ação combinada da quitosana no controle dos níveis de espécies reativas de oxigênio, em ação conjunta com o sistema enzimático antioxidativo.

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / A micropropagação de orquídeas é uma técnica utilizada com vários objetivos, podendo-se destacar a multiplicação rápida de plantas, propagação de novos híbridos, produção em larga escala e obtenção de matrizes livres de patógenos. A utilização de extratos naturais com ação antimicrobiana na micropropagação mostra-se como uma forma alternativa, ecológica e econômica de obtenção de plantas isentas de micro-organismos. Este trabalho teve como objetivo avaliar a utilização de extratos de Allium sativum (alho), de Syzygium aromaticum (cravo) e de própolis para evitar a contaminação com micro-organismos durante o processo de subcultivo in vitro de Epidendrum ibaguense Kunth e as possíveis interferências desses extratos no desenvolvimento vegetativo das plântulas. Foram isolados e identificados os micro-organismos Aspergillus sp., Candida sp., bactéria Gram-negativa, bactéria Gram-positiva e Penicillium sp. presentes em frascos com meio de cultura contaminados, oriundos do cultivo in vitro e do ambiente. Avaliou-se a ação antimicrobiana dos extratos sobre os micro-organismos isolados e observou-se que, o extrato de cravo inibiu o crescimento dos mesmos. Foram utilizados sete tratamentos compostos pelo meio Murashige; Skoog (1962) com metade da concentração de macronutrientes, acrescido de carvão vegetal e dos extratos de alho a 1,0 mgL-1 (T1), alho a 0,5 mgL-1 (T2), cravo a 1,0 mgL-1 (T3), cravo a 0,5 mgL-1 (T4), própolis a 1,0 mgL-1 (T5), própolis a 0,5 mgL-1 (T6) e o controle (T7) em um delineamento inteiramente casualizado, com cinco repetições e três plântulas por repetição. Após sessenta dias do subcultivo nos meios com os extratos avaliou-se a altura da parte aérea, o comprimento da maior raiz, o número de raízes, o número folhas, o comprimento da maior folha, o peso da massa fresca, o peso da massa seca e a taxa de multiplicação das plântulas. Houve influência dos extratos de alho, cravo e própolis no desenvolvimento in vitro de plântulas de E. ibaguense. O extrato de cravo apresentou ação antimicrobiana frente aos micro-organismos Aspergillus sp., Candida sp., Escherichia coli, Penicillium sp. e Streptococcus pneumoniae. Já os extratos de alho e própolis não demonstraram atividade antimicrobiana sobre nenhuma das cepas testadas. Através deste trabalho foi possível estabelecer um protocolo para o cultivo in vitro de E. ibaguense visando o controle da contaminação. / The micropropagation of orchids is a technique used for various purposes, may be noted the rapid multiplication of plants, propagation of new hybrid, large-scale production and obtaining free arrays of pathogens. The use of natural extracts with antimicrobial activity in micropropagation shows up as an alternative, environmentally friendly and economical way to obtain plants free of microorganisms. This study aimed to evaluate the use of extracts of Allium sativum (garlic), Syzygium aromaticum (clove) and propolis to prevent contamination with micro-organisms during the process of in vitro subculture of Epidendrum ibaguense Kunth and the possible interference of these extracts on vegetative growth of seedlings. The microorganisms Aspergillus sp. Candida sp. Gram-negative bacteria, Gram-positive and Penicillium sp. present in the contaminated bottles with culture, derived from in vitro culture and the environment, were isolated and identified. We evaluated the antimicrobial activity of the extracts on the isolated microorganisms and it was observed that the extract of clove inhibited their growth. Seven compound treatments were used by Murashige; Skoog (1962) with half the concentration of macronutrients, plus charcoal and garlic extracts to 1.0 mgL-1 (T1), garlic 0.5 mgL-1 (T2), cloves 1.0 mgL- 1 (T3), cloves 0.5 mgL-1 (T4), propolis to 1.0 mgL-1 (T5), propolis 0.5 mgL-1 (T6) and control (T7) on a randomized complete randomized with five replications and three seedlings per replication. Sixty days of subculture in the media with the extracts evaluated the shoot height, the length of roots, number of roots, leaves number, the length of the longest leaf, the weight of the fresh weight, the weight of the mass drought and the multiplication rate of the seedlings. There were significant effects of the extracts of garlic, clove and propolis in the development of in vitro plantlets of E. ibaguense. Clove extract showed antimicrobial activity against microorganisms Aspergillus sp., Candida sp., Escherichia coli, Penicillium sp. and Streptococcus pneumoniae and the extracts of garlic and propolis showed no antimicrobial activity against any of the tested strains. Through this work, it was possible to establish a protocol for the in vitro cultivation of E. ibaguense for the contamination control.

Na natureza, as orquídeas são totalmente dependentes de fungos micorrízicos para germinar. Estes fungos podem penetrar nas células das raízes e formar pélotons, os quais, quando digeridos pela planta, providenciam açúcares simples para o embrião. Durante a fase aclorofilada de desenvolvimento da plântula, orquídeas são obrigatoriamente dependentes dos fungos; algumas continuam assim por toda vida enquanto outras se tornam facultativamente responsivas à colonização. O objetivo deste trabalho foi identificar quantos clados de fungos podem estabelecer associação micorrízica com Coppensia doniana (sin. Oncidium donianum), uma orquídea amplamente distribuída nos arredores de Campos do Jordão e, demonstrar como as características morfológicas dos isolados, quando analisadas com ferramentas de estatística multivariada, podem ser úteis para a taxonomia destes fungos. Dez plantas foram amostradas em um sítio com vegetação típica de campos de altitude, junto ao Parque Estadual de Campos do Jordão. Fungos foram isolados pela transferência asséptica de cortes de raízes contendo pélotons para meios de cultura BDA modificados. Três clados de fungos foram formados, tanto analisando as características qualitativas das culturas quanto as quantitativas. Os clados foram identificados como dois morfotipos do gênero Ceratorhiza (fase anamórfica de Ceratobasidium) e uma Rhizoctonia-uninucleada. O sequenciamento da região ITS produziu resultados idênticos a estes, mostrando os mesmos três clados. Todas as sequências tiveram alta correlação com sequências de Ceratobasidium depositadas no Genbank, o que sugere uma alta afinidade de Coppensia doniana com este gênero. Também ficou demonstrado que os dados morfológicos, quando associados à estatística multivariada são uma ferramenta útil na taxonomia polifásica de Rhizoctonia spp. As sequências dos isolados de Coppensia doniana também foram comparadas com as de isolados de outras orquídeas, dentro da subtribo Oncidiinae, incluindo: Ionopsis utricularioides e Psygmorchis pussila, coletadas na região do Valle del Cauca Colômbia e isolados de 10 Ionopsis utricularioides, Oncidium altissimum e Tolumnia variegata, estudados por Otero (2002, 2004, 2007), em diferentes regiões de Porto Rico, Costa Rica, Cuba e Panamá. Esta última análise veio a comprovar a preferência de orquídeas da subtribo Oncidiinae por fungos do gênero Ceratobasidium, apesar de que os clados obtidos no Brasil e na Colômbia foram distantes filogeneticamente dos clados previamente estudados na América Central. Representantes dos três clados obtidos de C. doniana em Campos do Jordão foram também testados quanto à capacidade de induzir germinação em suas sementes. Todos isolados testados tiveram sucesso na germinação das sementes, levando as plântulas a estádios avançados de desenvolvimento após 30 dias, o que indica um alto potencial para utilização biotecnológica destes isolados para a germinação das sementes destas orquídeas, tanto para a floricultura comercial quanto para programas de reintrodução de espécies de orquídeas ameaçadas de extinção. / In nature, orchids are fully dependent on mycorrhizal fungi for germination. These fungi can penetrate root cells and form pelotons, whose digestion provides simple sugars for the embryo. During the achlorophyllous seedling stage, orchids are obligatory dependent on the fungi, and some species remain so through life, while others become facultatively responsive to fungal infection. The aim of this study was to identify how many fungal clades can establish mycorrhizal associations with Coppensia doniana, a widespread orchid from Campos do JordãoBrazil, and to demonstrate how their morphological features, analyzed with multivariate statistics, can be useful for classification. Ten plants were sampled in an Araucaria forest near Campos do Jordão. Fungi were isolated by transferring surface disinfected root segments containing pelotons to PDA culture medium. Three main fungal clades were formed by qualitative and quantitative morphological data. They were identified as two morphotypes of Ceratorhiza (anamorphic stage of Ceratobasidium) and one uninucleated Rhizoctonia. The ITS sequencing corroborates this identification, since the same three clades were found. All sequences were highly correlated to Ceratobasidium ITS data deposited at the Genebank, suggesting a high affinity between this species of Oncidiinae and Ceratobasidium. It also could be shown that morphological data associated with multivariate statistics can be a useful tool in fungal multi-level taxonomy. C. doniana sequences were also compared to sequences obtained from isolates of other orchids, belonging to the sub-tribe Oncidiinae, including: Ionopsis utricuarioides and Psygmorchis pussila, collected in Valle del Cauca Colombia and isolated from I. utricularioides, Oncidium altissimum and Tolumnia variegata, studied by Otero (2002, 2004, 2007) in different regions of Puerto Rico, Costa Rica and other Caribbean islands. This last analysis confirmed the preference of this Oncidiinae sub-tribe for Ceratobasidium, although isolates obtained in Brazil or Colombia belong to different clades from those previously studied in Puerto Rico, Costa Rica, Panama and Cuba. 12 Fungi representing the three clades obtained from C. doniana in Campos do Jordão were also tested for their ability to induce germination of C. doniana seeds, with a positive response for all of them, being able to bring the seedlings to advanced development stages in 30 days. These results suggest a high biotechnological potential of these isolates, to be used in orchid symbiotic germination for commercial flower production or for the reintroduction of endangered Brazilian orchid species.

Made available in DSpace on 2014-08-20T14:33:08Z (GMT). No. of bitstreams: 1 Dissertacao_Tangela_Denise_Perleberg.pdf: 8691099 bytes, checksum: 1f0a2e55e57d45a97d2b32e72803b425 (MD5) Previous issue date: 2009-05-04 / Rio Grande do Sul state south region is not well-known as for its forests floristic composition. This work conducted a taxonomic treatment of Orchidaceae species occurring in Morro Quilongongo, a seasonal semi-deciduous forest fragment located in Southeast Slope, in the municipality of Pelotas, Rio Grande do Sul state. The work shows descriptions, identification key and illustrations of the species, complemented with data about habitat, phenology and geographic distribution. Forty species of Orchidaceae were found distributed in 22 genders. The most representative genders in number of species were Oncidium (8 spp.), Acianthera (4 spp.), Anathallis and Specklinia (3 spp. for each of the plants), however, 14 genders (64%) presented only one specie. The most found species, 29 (72%), are epiphyte, 9 of them are frequently observed upon the rocks. Eleven (27.5%) are terrestrial, seven are frequently found occurring in rocks. The number of species in bloom by month varied from 8 to 14. In September and October a larger number of species come into bloom. In April and July a smaller number of species come into bloom. The regions of Rio Grande do Sul which presented the largest number of these species were the Superior Northeast Slope (30 spp.), Central Depression (27 spp.) and Coast (26 spp.). The Orchidaceae flora of the region is not much represented in the consulted herbaria; just only 11 (27.5%) species found in Morro Quilongongo have a representative exsiccate of the region of Pelotas. Therefore, this work contributes to the knowledge of the South region flora of Rio Grande do Sul, providing data that justify the preservation of this area. / A região sul do Rio Grande do Sul é pouco conhecida quanto à composição florística de suas florestas. Neste trabalho foi realizado o tratamento taxonômico das espécies de Orchidaceae ocorrentes no Morro Quilongongo, um fragmento de floresta estacional semidecidual, localizado na Encosta do Sudeste, no município de Pelotas, Estado do Rio Grande do Sul. São apresentadas descrições, chave de identificação e ilustrações das espécies, complementadas com dados sobre habitat, fenologia e distribuição geográfica. Foram encontradas 40 espécies de Orchidaceae distribuídas em 22 gêneros. Os gêneros mais representativos, em número de espécies, foram Oncidium (8 spp.), Acianthera (4 spp.) e Anathallis e Specklinia (3 spp. cada), porém, 14 gêneros (64%), apresentam apenas uma espécie. A maioria das espécies encontradas, 29 (72,5%), é epífita, sendo 9 destas também observadas frequentemente sobre rochas. Onze (27,5%) são terrestres, sendo 7 frequentemente encontradas como rupícolas. O número de espécies floridas por mês variou de 8 a 14, sendo setembro e outubro os meses com maior número de espécies floridas e abril e julho os meses com menor número de espécies floridas. As regiões do Rio Grande do Sul que apresentaram o maior número de espécies comuns à área de estudo foram a Encosta Superior do Nordeste (30 spp.), Depressão Central (27 spp.) e Litoral (26 spp.). A flora de Orchidaceae da região é pouco representada nos herbários consultados, sendo que apenas 11 (27,5%) espécies encontradas no Morro Quilongongo possuem uma exsicata representativa da região de Pelotas. Portanto, o trabalho contribui para o conhecimento da flora da região sul do Rio Grande do Sul gerando dados que justificam a preservação da área.

O gênero Burkholderia é composto por bactérias de elevada diversidade fisiológica e genética, sendo alvo para uma série de aplicações biotecnológicas. A linhagem Burkholderia seminalis TC3.4.2R3 foi capaz de controlar fungos e bactérias patogênicas, bem como produzir antibióticos. A análise de perfil fisiológico da planta hospedeira indica que não há a elicitação de respostas de defesa no controle dos sintomas da podridão mole por B. seminalis. Foi observado que o sucesso do controle depende do contato direto do agente de biocontrole e o patógeno B. gladioli, e que B. seminalis possui atividade antimicrobiana in vitro contra com a formação de um halo de inibição e redução de 40% da densidade de B. gladioli em co-cultura líquida com B. seminalis. A análise do mutante M01 defectivo no controle da podridão mole, que apresenta a região intergênica da patatina-ferritina nocauteada pelo transposon, não mostrou diferenças significativas comparado ao isolado selvagem para os mesmos ensaios. / The genus Burkholderia is composed of bacteria with high physiological and genetic diversity, and it is being goal for a number of biotechnological applications. The Burkholderia seminalis TC3.4.2R3 strain was able to control fungal and bacterial pathogens as well as produce antibiotics. The physiological profile analysis of the host plant indicates that there is no defense responses elicitation induced by B. seminalis involved in controlling symptoms of soft rot. It was observed that the success of control depends on the full contact of the biocontrol agent and the pathogen B. gladioli. B. seminalis has an antimicrobial activity in vitro showing a inhibition halo and reducing the density of B. gladioli in co-culture with B. seminalis by 40%. The M01 mutant, wich is defective in control of soft rot and showing a insertion of transposon in a intergenic region between patatin-ferritin, presented no significant differences compared to the wild strain for the same assays.

No description available.