Dressing liposomal particles with chitosan and poly(vinylic alcohol) for oral vaccine delivery

Liposomes have been used as adjuvants since 1974. One major limitation for the use of liposomes in oral vaccines is the lipid structure instability caused by enzyme activities. Our aim was to combine liposomes that could encapsulate antigens (i.e., Dtxd, diphtheria toxoid) with chitosan, which protects the particles and promotes mucoadhesibility. We employed physical techniques to understand the process by which liposomes (SPC: Cho, 3: 1) can be sandwiched with chitosan (Chi) and stabilized by PVA (poly-vinylic alcohol), which are biodegradable, biocompatible polymers. Round, smooth-surfaced particles of REVs-Chi (reversed-phase vesicles sandwiched by Chi) stabilized by PVA were obtained. The REVs encapsulation efficiencies (Dtxd was used as the antigen) were directly dependent on the Chi and PVA present in the formulation. Chi adsorption on the REVs surface was accompanied by an increase of zeta-potential. In contrast, PVA adsorption on the REVs-Chi surface was accompanied by a decrease of zeta-potential. The presence of Dtxd increased the Chi surface-adsorption efficiency. The PVA affinity by mucine was 2,000 times higher than that observed with Chi alone and did not depend on the molecule being in solution or adsorbed on the liposomal surface. The liberation of encapsulated Dtxd was retarded by encapsulation within REVs-Chi-PVA. These results lead us to conclude that these new, stabilized particles were able to be adsorbed by intestinal surfaces, resisted degradation, and controlled antigen release. Therefore, REVs-Chi-PVA particles can be used as an oral delivery adjuvant.

Mechanism of Trypanosoma cruzi death induced by Cratylia mollis seed lectin

Incubation of T. cruzi epimastigotes with the lectin Cramoll 1,4 in Ca(2+) containing medium led to agglutination and inhibition of cell proliferation. The lectin (50 A mu g/ml) induced plasma membrane permeabilization followed by Ca(2+) influx and mitochondrial Ca(2+) accumulation, a result that resembles the classical effect of digitonin. Cramoll 1,4 stimulated (five-fold) mitochondrial reactive oxygen species (ROS) production, significantly decreased the electrical mitochondrial membrane potential (Delta I(m)) and impaired ADP phosphorylation. The rate of uncoupled respiration in epimastigotes was not affected by Cramoll 1,4 plus Ca(2+) treatment, but oligomycin-induced resting respiration was 65% higher in treated cells than in controls. Experiments using T. cruzi mitochondrial fractions showed that, in contrast to digitonin, the lectin significantly decreased Delta I(m) by a mechanism sensitive to EGTA. In agreement with the results showing plasma membrane permeabilization and impairment of oxidative phosphorylation by the lectin, fluorescence microscopy experiments using propidium iodide revealed that Cramoll 1,4 induced epimastigotes death by necrosis.

Spatio-temporal variation in seed production in three Euphorbia species and the role of animals on seed fate

Comparative studies on the reproductive biology of co-occurring related plant species have provided valuable information for the interpretation of ecological and evolutionary phenomena, with direct application in conservation management of plant populations. The aims of this thesis were to identify the causes of pre-dispersal reproductive losses in three Euphorbia species (the Mediterranean E. characias and the narrow endemics E. pedroi and E. welwitschii) and evaluate the variation of their effects in time, space and between individuals and species. Furthermore, we intended to study elaiosomes’ fatty acid profiles for the three Euphorbia and assess the role played by the elaiosome in ant attraction. Finally, we aimed to identify the major seed dispersal agents for each Euphorbia species in each site and study differences in short term seed fate due to differences in ant behaviour. The results indicated that intact seed production differed significantly between the three Euphorbia, mostly due to differences in cyathia production. Losses to pre-dispersal seed predators were proportionately larger for the endemic species which also suffered higher losses resulting in flower, fruit (in E. welwitschii) and seed abortion (in E. pedroi). The elaiosomes of E. pedroi are poor in fatty acids and for this reason seeds of this species were removed in lower proportion by mutualistic dispersers than those of their congeners, being more prone to seed predation. Two larger ant species – Aphaenogaster senilis and Formica subrufa – were responsible for a larger percentage of removals with seeds being transported at larger distances and being discarded in the vicinity of their nests following elaiosome removal. Our results highlight the role of insect-plant interactions as major determinants of seed survival for the three study plants and call for the need to include more information on insect-plant interactions in plant conservation programmes.

A scanning electron microscopy study of the seed and post-seminal development in Angelonia salicariifolia Bonpl. (Scrophulariaceae)

The external morphology of seeds and post-germination developmental stages of Angelonia salicariifolia Bonpl. (Scrophulariaceae) were investigated using scanning electron microscopy. Some structural features of the seed exotesta and seedling in Angelonia are presented for the first time and are of potential taxonomic value for this neotropical genus. The seeds are very small (0.9-1.7 mm long and 0.5-0.9 mm wide), ovate, with a reticulate-crested exotesta, reticules arranged uniformly in longitudinal rows, with a high density of microcilia-like projections on the cell wall of the reticule base and on the edge of the crests. The hilum is located beside the micropyle at the narrow end of the seed. Germination is epigeal. During germination the radicle develops, followed by elongation of the hypocotyl and primary root. At this stage dense root hairs develop on the lower part of the hypocotyl. The apical bud-located between the cotyledons-begins to develop after the cotyledons have unfolded. The cotyledons are equal in size, sessile and ovate. The seedlings have two types of trichomes, one characteristic of the cotyledons and first pair of leaves (glandular, sessile, four-celled head with quadrangular shape) and the other characteristic of the hypocotyl and epicotyl (stalked, erect, elongate and three-celled with dome-shaped unicellular head). (C) 2001 Annals of Botany Company.

Electrical conductivity of the seed soaking solution and soybean seedling emergence

Vigor of soybean [Glycine max (L.) Merrill] seeds can be evaluated by measuring the electrical conductivity (EC) of the seed soaking solution, which has shown a satisfactory relationship with field seedling emergence, but has not had aproper definition of range yet. This work studies the relationship between EC and soybean seedling emergence both in the field and laboratory conditions, using twenty two seed lots. Seed water content, standard germination and vigor (EC, accelerated aging and cold tests) were evaluated under laboratory conditions using -0.03; -0.20; -0.40 and -0.60 MPa matric potentials, and field seedling emergence was also observed. There was direct relationship between EC and field seedling emergence (FE). Under laboratory conditions, a decreasing relationship was found between EC and FE as water content in the substrate decreased, Relationships between these two parameters were also found when -0.03; -0.20 and -0.40 MPa matric potentials were used. EC tests can be used successfully to evaluate soybean seed vigor and identify lots with higher or lower field emergence potential.

Accelerated ageing test to evaluate seed vigor in pumpkin and zucchini seeds

The accelerated ageing (AA) test is widely used to evaluate seed vigor in various species including vegetable crops. The objective of this work was to study procedures for conducting AA to evaluate seed vigor of pumpkin and zucchini seed lots. Five seed lots of commercially acceptable quality (germination) standards were selected from both cv. `Menina Brasileira' (zucchini) and `Barbara' Hybrid (pumpkin). The following tests were conducted: standard germination, the first count of germination, and AA with and without saline solution (periods of 48, 72 and 96 hours). For each combination of temperature and ageing period, seeds were placed in a single layer on a screen in a germination plastic box with either 40 mL deionized water or 40g NaCl/100mL of water. Both AA methods identified lots 2 and 5 of `Menina Brasileira' and lot 10 of `Barbara' hybrid as having the lowest physiological quality. The temperature 41 C promoted a more drastic reduction in germination than did 38 C. This observation was more marked after 96 hours. Based on the results obtained, it can be concluded that the combination 41 degrees C/96 hours should be used to evaluate the physiological potential of pumpkin and zucchini seeds; however, additional studies are necessary to evaluate other combinations and confirm this indication.

The amount of solar energy made available for the production of a sabia (Mimosa caesalpinaefolia Benth.) seed has a determinating effect on its becoming dormant

The amount of solar energy made available for the production of a sabid seed varied as a function of the time of the year, the face of the plant in which, the position in the plant on which and the position in the pod in which it was produced.Variation in solar energy availability as a consequence of the time of the year was a direct consequence of latitude. At 21degrees5'22 S the highest amounts of Global Solar Radiation (GSR) reaching the site where the experiment was conducted took place during the months from November through February. During these months there were no marked differences between any two of the amounts of GSR reaching faces North (N), South (S), West (W) East (E). From February through November (period during which the sabid plants of this study flowered and the resulting seeds matured and were harvested) the total GSR's were the lowest and marked differences were found between faces N and S, with face N receiving much more GSR than face S. During that period, faces W and E received practically the same amount of GSR and it was much less than that received by face N and much more than the one received by face S.The amount of biological energy made available for the development of a seed seemed also to vary according to a dry matter partitioning strategy by the plant -the central third of the plant seemed to be the one receiving the highest amounts of energy, followed either by the upper or the lower third of the plant- it was not very clear which third of the plant immediately followed the central one. The partitioning of biological energy at the pod level also seemed to follow a strategy by which the central seeds would be the ones to receive more, followed by the proximal seeds and these by the distal ones.This availability of energy seemed to have a direct effect on seed size, weight and on the percentage of seeds which showed a degree of dormancy deep enough to prevent their germinating under the conditions of a standard germination test.The implications of these results for the improvement of methods for the overcoming of dormancy of sabia seeds are discussed.

Nitrogênio na produção e qualidade de sementes de feijão

O uso de fertilizante nitrogenado, em campos de produção de sementes, pode beneficiar a produtividade e a qualidade da semente, por afetar a formação do embrião e dos órgãos de reserva. Assim, objetivou-se, com este trabalho, avaliar a produção e o potencial fisiológico de sementes de feijoeiro, em função da aplicação de nitrogênio em cobertura, em duas cultivares. Utilizou-se o delineamento de blocos casualizados, em esquema fatorial 2×6, com 12 tratamentos (cultivares Pérola e IAC Carioca Eté e seis doses de nitrogênio em cobertura: 0 kg ha-1, 30 kg ha-1, 60 kg ha-1, 90 kg ha-1, 120 kg ha-1 e 150 kg ha-1) e quatro repetições. O nitrogênio promoveu aumentos na produção de sementes, com dose ótima de 143 kg de N ha-1, para a cultivar Pérola, e 134 kg de N ha-1, para a cultivar IAC Carioca Eté, com a Pérola apresentando maiores níveis de produtividade. Alterações na qualidade de sementes não foram verificadas com o aumento das doses de nitrogênio.

Temperature during soybean seed storage and the amount of electrolytes of soaked seeds solution

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Seed-borne pathogens and electrical conductivity of soybean seeds

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Electrical conductivity as an indicator of pea seed aging of stored at different temperatures

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Relationship of black layer and milk line development on maize seed maturity

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Seed anatomy and germination of Phoenix roebelenii O'Brien (Arecaceae)

O objetivo desse trabalho foi estudar a morfologia, anatomia e o processo germinativo de sementes de Phoenix roebelenii. Para o levantamento dos dados biométricos foram utilizadas 100 sementes de frutos recém-colhidos, deixados secar ao ar por um dia. Para a germinação, quatro repetições de 50 sementes tratadas com Vitavax-Thiran foram semeadas em bandejas de plástico, contendo Sphagnum sp. como substrato e mantidas sob condições ambientais de laboratório. Detalhes da morfologia da semente foram documentados com o uso de microscopia eletrônica de varredura e esquematizados com auxílio de câmara clara, acoplada ao estereomicroscópio. Foram confeccionadas lâminas permanentes com cortes do embrião, para o estudo de sua anatomia. As dimensões médias das sementes foram: 10,32mm comprimento, 5,21mm largura e 3,91mm espessura. O peso de 1000 sementes foi de 151,1g e 1kg continha 6600 unidades. O início da germinação variou entre 27 e 58 dias. As sementes são albuminosas, com endosperma duro e o embrião é pouco diferenciado, lateral e periférico. A germinação inicia-se pela abertura de um opérculo, através do qual é emitido o pecíolo cotiledonar com o eixo embrionário na extremidade. O pecíolo funciona, internamente, como um haustório, digerindo gradativamente o endosperma. Na sua parte posterior, desenvolve-se a plúmula, que emerge através de uma fenda. Nota-se o aparecimento de raízes secundárias na porção anterior da raiz primária.

Automated system of seedling image analysis (SVIS) and electrical conductivity to assess sun hemp seed vigor

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)