WUSCHEL-related genes are expressed during somatic embryogenesis of the basal angiosperm Ocotea catharinensis Mez. (Lauraceae)

Ocotea catharinensis is a basal angiosperm and an endangered tree species from the Brazilian Atlantic Rain Forest. Despite its economical and ecological importance, mass-propagation of this species is hampered by seldom-produced short-lived seeds, and in vitro propagation is challenged by frequently malformed somatic embryos. Therefore, O. catharinensis somatic embryos are also a good experimental material to study the physiological and molecular mechanisms underlying in vitro morphogenesis. In an ongoing effort to characterize genes expressed during somatic embryogenesis of O. catharinensis we have cloned two Ocotea WUSCHEL-related genes. According to our RT-PCR data, both genes were preferentially expressed in embryogenic cell aggregates. One of them, OcWUS, is a possible ortholog of the Arabidopsis WUSCHEL (WUS) gene, which codes for a homeodomain-containing protein involved in the specification and maintenance of the shoot apical meristem. We analyzed the expression patterns of OcWUS and OcWOX4 by RT-PCR, and OcWUS expression was also assessed by in situ hybridization. The expression patterns of OcWUS were very similar to those described for the Arabidopsis WUS. OcWUS transcripts were generally restricted to a small group of cells in the center of the putative shoot apical meristem of O. catharinensis somatic embryos. Perturbed expression of OcWUS might be related to abnormally formed somatic embryos of O. catharinensis obtained through tissue culture.

QUALIDADE FISIOLÓGICA DE SEMENTES DE Tabernaemontana fuchsiaefolia A. DC EM FUNÇÃO DO TESTE DE ENVELHECIMENTO ACELERADO1

Tabernaemontana fuchsiaefolia A. DC é uma espécie da família Apocynaceae, nativa do Brasil, com característica invasora, possuindo potencial para uso em recuperação de áreas degradadas. O trabalho teve o objetivo de avaliar a influência dos métodos de envelhecimento acelerado sobre a qualidade fisiológica e germinação de sementes, utilizando-se o método tradicional e em solução de NaCl, empregando-se a metodologia de gerbox, formando minicâmaras de envelhecimento, em três temperaturas (41, 43 e 45°C) e cinco tempos de envelhecimento (0, 24, 48, 72 e 96 horas). As sementes, após o envelhecimento, foram colocadas para germinar em rolos de papel germitest, em quatro repetições de 25 sementes, em câmaras BOD a temperatura de 30°C, com fotoperíodo de 8 horas. O efeito da temperatura foi comparado pelo teste de Tukey em nível de 5% de probabilidade e o efeito do tempo de envelhecimento, pela análise de regressão. Após 35 dias, foram avaliados percentual, índice de velocidade, tempo médio e frequência da germinação, bem como comprimento de raiz e parte aérea, massa seca de plântulas e percentual de plântulas normais. Observou-se que o método alternativo de envelhecimento com solução de NaCl teve pouca influência na qualidade das sementes, não sendo indicado para testes de vigor para a espécie. O método tradicional, na temperatura de 45°C em período de envelhecimento de 96 horas, promoveu queda na germinação e pode ser utilizado para testar o vigor de sementes de leiteira.

An inventory of the ethnobotanicals used as anti-diabetic by a rural community of Dhemaji district of Assam, Northeast India

Ethnopharmacological relevance: Traditional remedies used for treating diabetic ailments are very important in the primary health care of the people living in rural Dhemaji district of Assam, north-east India. Novel information gathered from the current survey is important in preserving folk indigenous knowledge. Materials and methods: Interviews were conducted amongst 80 households comprising of 240 individuals using semi-structured questionnaires. The focus was on plants used in treating diabetes mellitus. Results: The current survey documented 21 plant species (20 families) which are reportedly used to treat diabetes mellitus by the rural people in the study area. To the best of our knowledge, Amomum linguiforme, Cinnamomum impressinervium, Colocasia esculenta, Dillenia indica, Euphorbia ligularia, Garcinia pedunculata, Solanum indicum, Sterculia villosa and Tabernaemontana divaricata are recorded for the first time based on globally published literature as medicinal plants used for treating diabetes mellitus and related symptoms. Conclusions: The wide variety of plants that are used to treat diabetes mellitus in this area supports the traditional value that medicinal plants have in the primary health care system of the rural people of Dhemaji district of Assam. The finding of new plant uses in the current study reveals the importance of the documentation of such ethnobotanical knowledge. (C) 2011 Elsevier Ireland Ltd. All rights reserved.

Comunidades macrofaunais bênticas associadas às algas de deriva e à grama marinha Halodule wrightii Ascherson na Ilha do Japonês, Cabo Frio, RJ

As extensas pradarias submersas formadas pelas gramas marinhas são importantes habitats da costa, onde ocorrem interações ecológicas entre diversas espécies da vegetação subaquática, invertebrados bentônicos e peixes. As gramas marinhas e algas de deriva são conhecidas como macrófitas marinhas e, por ocuparem o mesmo tipo de substrato, são normalmente encontradas juntas, proporcionando oxigênio, alimento, proteção, abrigo além de sítios de reprodução e pastagem para os animais associados a essas pradarias. Amostras de algas de deriva e de H. wrightii foram coletadas, ao longo de transectos fixos de 50 m paralelos à Ilha do Japonês, a fim de analisar a existência de relações positivas entre as espécies de macrófitas marinhas e sua macrofauna associada, comparar as duas comunidades e avaliar a estruturação da comunidade macrofaunal bêntica do local. Os transectos foram alocados de acordo com a posição do banco de grama marinha. Observou-se que a densidade de eixos e a biomassa de H. wrightii não explicam a variação da biomassa, riqueza de espécies e diversidade (Índice de Simpson) das algas de deriva. A grande movimentação das algas de deriva ao longo do banco de grama marinha faz com que elas se homogeneízem e ocupem diferentes lugares ao acaso na pradaria, muitos desses locais com baixa biomassa de H. wrightii devido à grande variabilidade na distribuição dessa espécie no local de estudo. Os descritores ecológicos da grama marinha também não tiveram relações positivas com sua macrofauna bêntica associada. A comunidade macrofaunal associada às gramas marinhas foi mais densa, rica e diversa do que a comunidade macrofaunal associada às algas de deriva. Os moluscos Anomalocardia flexuosa, Cerithium atratum, Ostrea sp, Tellina lineata e Divalinga quadrissulcata dominaram o ambiente de gramas marinhas. A maior complexidade estrutural das algas de deriva forneceu um habitat protegido mais atrativo para os crustáceos como, Pagurus criniticornis, Cymadusa filosa e Batea catharinensis. A malacofauna associada às algas não foi abundante, mas um novo registro foi a ocorrência do bivalve invasor Lithopaga aristatus, perfurando uma concha de Ostrea sp. As relações entre os descritores da biomassa algal foram comprovadas para a maioria dos descritores de sua fauna associada. As relações das macrófitas marinhas com a macrofauna total associada seguiram o mesmo padrão das relações das algas de deriva. As análises de agrupamento e ordenação mostraram que as comunidades macrofaunais bênticas do local são estruturadas de acordo com os táxons dos organismos associados mais dominantes influenciados pelo tipo de vegetação basibionte (algas de deriva ou grama marinha). Destaca-se com o presente estudo a importância de medidas de maior proteção no local para a preservação e manutenção do ecossistema da Ilha do Japonês, RJ, Brasil

Localization of monoterpenoid indole alkaloid (MIA) biosynthesis by in situ hybridization (ISH)

Monoterpenoid indole alkaloids (MIA) are among the largest and most complex group of nitrogen containing secondary metabolites that are characteristic of the Apocynaceae plant family including the most notable Catharanthus roseus. These compounds have demonstrated activity as successful drugs for treating various cancers, neurological disorders and cardiovascular conditions. Due to the low yields of these compounds and high pharmacological value, their biosynthesis is a major topic of study. Previous work highlighting the leaf epidermis and leaf surface as a highly active area in MIA biosynthesis and MIA accumulation has made the epidermis a major focus of this thesis. This thesis provides an in-depth analysis of the valuable technique of RNA in situ hybridization (ISH) and demonstrates the application of the technique to analyze the location of the biosynthetic steps involved in the production of MIAs. The work presented in this thesis demonstrates that most of the MIAs of Eurasian Vinca minor, African Tabernaemontana e/egans and five Amsonia species, including North American Amsonia hubrichitii and Mediterranean A. orienta/is, accumulate in leaf wax exudates, while the rest of the leaf is almost devoid of alkaloids. Biochemical studies on Vinca minor displayed high tryptophan decarboxylase (TOe) enzyme activity and protein expression in the leaf epidermis compared to whole leaves. ISH studies aimed at localizing TOe and strictosidine synthase suggest the upper and lower epidermis of V. minor and T. e/egans as probable significant production sites for MIAs that will accumulate on the leaf surface, however the results don't eliminate the possibility of the involvement of other cell types. The monoterpenoid precursor to all MIAs, secologanin, is produced through the MEP pathway occurring in two cell types, the IPAP cells (Gl0H) and epidermal cells (LAMT and SLS). The work presented in this thesis, localizes a novel enzymatic step, UDPG-7-deoxyloganetic acid glucosyltransferase (UGT8) to the IPAP cells of Catharanthus longifolius. These results enable the suggestion that all steps from Gl0H up to and including UGT8 occur in the IPAP cells of the leaf, making the IPAP cells the main site for the majority of secologanin biosynthesis. It also makes the IPAP cells a likely cell type to begin searching for the gene of the uncharacterized steps between Gl0H and UGT8. It also narrows the compound to be transported from the IPAP cells to either 7-deoxyloganic acid or loganic acid, which aids in the identification of the transportation mechanism.

The γ-tocopherol-like family of N-methyltransferases: A taxonomically clustered gene family encoding enzymes responsible for N-methylation of monoterpene indole alkaloids

The plant family Apocynaceae accumulates thousands of monoterpene indole alkaloids (MIAs) which originate, biosynthetically, from the common secoiridoid intermediate, strictosidine, that is formed from the condensation of tryptophan and secologanin molecules. MIAs demonstrate remarkable structural diversity and have pharmaceutically valuable biological activities. For example; a subunit of the potent anti-neoplastic molecules vincristine and vinblastine is the aspidosperma alkaloid, vindoline. Vindoline accumulates to trace levels under natural conditions. Research programs have determined that there is significant developmental and light regulation involved in the biosynthesis of this MIA. Furthermore, the biosynthetic pathway leading to vindoline is split among at least five independent cell types. Little is known of how intermediates are shuttled between these cell types. The late stage events in vindoline biosynthesis involve six enzymatic steps from tabersonine. The fourth biochemical step, in this pathway, is an indole N-methylation performed by a recently identified N-methyltransfearse (NMT). For almost twenty years the gene encoding this NMT had eluded discovery; however, in 2010 Liscombe et al. reported the identification of a γ-tocopherol C-methyltransferase homologue capable of indole N-methylating 2,3-dihydrotabersonine and Virus Induced Gene Silencing (VIGS) suppression of the messenger has since proven its involvement in vindoline biosynthesis. Recent large scale sequencing initiatives, performed on non-model medicinal plant transcriptomes, has permitted identification of candidate genes, presumably involved, in MIA biosynthesis never seen before in plant specialized metabolism research. Probing the transcriptome assemblies of Catharanthus roseus (L.)G.Don, Vinca minor L., Rauwolfia serpentine (L.)Benth ex Kurz, Tabernaemontana elegans, and Amsonia hubrichtii, with the nucleotide sequence of the N-methyltransferase involved in vindoline biosynthesis, revealed eight new homologous methyltransferases. This thesis describes the identification, molecular cloning, recombinant expression and biochemical characterization of two picrinine NMTs, one from V. minor and one from R. serpentina, a perivine NMT from C. roseus, and an ajmaline NMT from R. serpentina. While these TLMTs were expressed and functional in planta, they were active at relatively low levels and their N-methylated alkaloid products were not apparent our from alkaloid isolates of the plants. It appears that, for the most part, these TLMTs, participate in apparently silent biochemical pathways, awaiting the appropriate developmental and environmental cues for activity.

Dinâmica de crescimento e produção de cinco gramíneas nativas do sul do Brasil

Pelo fato de algumas gramíneas nativas apresentarem um bom valor forrageiro objetivou-se com este trabalho caracterizar a dinâmica do crescimento de algumas espécies: Paspalum notatum ecótipos André da Rocha e Comum, P. pauciciliatum, P. lividum, e Axonopus catharinensis, relacionada com as suas características morfogenéticas. O solo foi corrigido de acordo com análise do solo e a adubação nitrogenada foi aplicada para otimizar a disponibilidade deste elemento. Os parâmetros estudados foram: produção de matéria seca verde (MSV), de lâminas foliares, bainhas foliares, colmo e inflorescências, material morto e outras espécies. O filocrono foi menor para o ecótipo André da Rocha no período de primavera e inverno-primavera, sendo o ecótipo Comum no período de verão. A duração de vida da folha, elongação e senescência foliar têm uma grande variabilidade entre os materiais, com grande influência da época do ano, notadamente o outono, quando todos materiais diminuem seu ritmo de crescimento. Axonopus pode ter maior período de acúmulo pois uma senescência significativa ocorre posteriormente aos materiais do gênero Paspalum e apresenta alta duração de vida da folha (> 1.000 GD). Produções ao redor de 10 t/ha de MSV foram alcançadas. Axonopus foi mais produtivo tanto em MSV quanto lâminas foliares. P. pauciciliatum teve um intenso florescimento que dificulta o manejo. P. notatum Comum permitiu o maior crescimento de espécies invasoras. Axonopus permite um maior período de acúmulo líquido de folhas e 400 GD é o limite máximo para o corte dos materiais do gênero Paspalum. Ocorre uma grande estacionalidade da produção nos materiais do gênero Paspalum. Todos os materiais avaliados apresentam um grande potencial de uso como espécie forrageira, podendo ser recomendadas para o plantio, porém com maiores cuidados no manejo do Pauciciliatum.

Epiderme e padrão de venação foliar de espécies de Lauraceae

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Antiviral activity of plants occurring in the state of minas gerais (Brazil): Part III

A total of 24 extracts from 14 plant species collected at the state of Minas Gerais, Brazil, and belonging to five botanical families (Annonaceae, Apocynaceae, Ochnaceae, Polygonaceae and Vitaceae) was screened for cytotoxicity in cultured Vero cells and for antiviral activity against human herpes virus type 1 (HSV-1), vaccinia virus (VACV) and murine encephalomyocarditis virus (EMCV). The highest cytotoxicity (CC 50 < 10 μg/mL) was observed for the ethanol extracts from Annona coriacea fruits and seeds. Extracts from Hancornia speciosa, Ouratea castaneafolia and O. semisrrata were the only ones that have shown activity against all the three viruses assayed. Extracts from Polygonum spectabile, Hancornia speciosa, Himatanthus phagedaenica, Ouratea spectabilis and O. semiserrata were the most active against HSV-1 (EC 50 < 50 mg/mL), with favorable SI values (8.0 to 10.0). Hancornia speciosa and Anaxagorea dolichocarpa were the most active against EMCV (EC 50 50 - 100 μg/mL), with reasonable SI values (5.2 to 6.1), while moderate to low activity (EC 50 > 100 μg/mL) was observed for Ouratea spectabilis and O. semiserrata. A total of 7 plant species, Ouratea semiserrata, O. spectabilis, O. castanaeafolia, Rollinia laurifolia, Cissus erosa, Polygonum spectabile, and Hancornia speciosa, were active against VACV, disclosing EC50 < 50 μg/mL and SI values ranging from 6.6 to 67.3. In total, 10 out of the 14 species were selected from a literature survey on plants used to treat viral diseases in Brazil; these species were responsible for 70% of the positive results.

Diversidade de ácaros em cultivo orgânico de citros e na vegetação natural circundante, e perspectivas para a criação massal de Iphiseiodes zuluagai (Acari: Phytoseiidae)

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Estudos estruturais e cristalização de complexos formados entre toxinas botrópicas e inibidores vegetais

This research presents a study of structural complexes formed between the bothropstoxin BthTX-I and PrTX-I and phospholipase A2 (PLA2s) inhibitor 12- methoxy-4-methyl-voachalotine (MMV) of Tabernamontana catharinensis and rosmarinic acid (Cv-RA) of Rosmarinus officinalis. For both were used cocrystallization experiments with the toxins and inhibitors. In the future, the experimental three-dimensional structures of these complexes will be obtained through the technique of crystallography. Simultaneously, construction occurred in silico of inhibitors Cv-RA and MMV and attempts of docking between these binders and protein structures. These theoretical data provide important information regarding the nature of interactions between inhibitors, toxins and structural changes induced by the protein molecule inhibitors. Therefore, the acquisition of this information has great value to identification, characterization and development (drug design) of compounds with great potential for biotechnology and drug use. Considering further and more relevant aplications, these compounds could be used to control symptoms in many cases of ophidic poisoning and also in combating of pathological processes (degenerative inflammatory diseases and auto-immune diseases, Alzheimer's disease, schizophrenia, among others) whose causes are related to molecules belonging to the same groups of proteins which are classified bothropstoxin that were studied in this project

Pollen analysis of surface sediments from the Gulf of Guinea and offshore NW-Africa

The areas of marine pollen deposition are related to the pollen source areas by aeolian and fluvial transport regimes, whereas wind transport is much more important than river transport. Pollen distribution patterns of Pinus, Artemisia, Chenopodiaceae-Amaranthaceae, and Asteraceae Tubuliflorae trace atmospheric transport by the northeast trades. Pollen transport by the African Easterly Jet is reflected in the pollen distribution patterns of Chenopodiaceae-Amaranthaceae, Asteraceae Tubuliflorae, and Mitracarpus. Grass pollen distribution registers the latitudinal extension of Sahel, savannas and dry open forests. Marine pollen distribution patterns of Combretaceae-Melastomataceae, Alchornea, and Elaeis reflect the extension of wooded grasslands and transitional forests. Pollen from the Guinean-Congolian/Zambezian forest and from the Sudanian/Guinean vegetation zones mark the northernmost extension of the tropical rain forest. Rhizophora pollen in marine sediments traces the distribution of mangrove swamps. Only near the continent, pollen of Rhizophora, Mitracarpus, Chenopodiaceae-Amaranthaceae, and pollen from the Sudanian and Guinean vegetation zones are transported by the Upwelling Under Current and the Equatorial Under Current, where those currents act as bottom currents. The distribution of pollen in marine sediments, reflecting the position of major climatic zones (desert, dry tropics, humid tropics), can be used in tracing climatic changes in the past.