Luzimar Campos da Silva

Somatic embryogenesis of a wild passion fruit species Passiflora cincinnata Masters: histocytological and histochemical evidences.

The characterization of cellular changes that occur during somatic embryogenesis is essential for understanding the factors involved in the transition of somatic cells into embryogenically competent cells and determination of cells and/or tissues involved. The present study describes the anatomical and ultrastructural events that lead to the formation of somatic embryos in the model system of the wild passion fruit (Passiflora cincinnata). Mature zygotic embryos were inoculated in Murashige and Skoog induction media supplemented with 2,4-dichlorophenoxyacetic acid and 6-benzyladenine. Zygotic embryo explants at different development stages were collected and processed by conventional methods for studies using light, scanning, and transmission electron microscopy (TEM). Histochemical tests were used to examine the mobilization of reserves. The differentiation of the somatic embryos began in the abaxial side of the cotyledon region. Protuberances were formed from the meristematic proliferation of the epidermal and mesophyll cells. These cells had large nuclei, dense cytoplasm with a predominance of mitochondria, and a few reserve compounds. The protuberances extended throughout the abaxial surface of the cotyledons. The ongoing differentiation of peripheral cells of these structures led to the formation of proembryogenic zones, which, in turn, dedifferentiated into somatic embryos of multicellular origin. In the initial stages of embryogenesis, the epidermal and mesophyll cells showed starch grains and less lipids and protein reserves than the starting explant. These results provide detailed information on anatomical and ultrastructural changes involved in the acquisition of embryogenic competence and embryo differentiation that has been lacking so far in Passiflora.

Flúor em chuva simulada: sintomatologia e efeitos sobre a estrutura foliar e o crescimento de plantas arbóreas

Fluoride in simulated rain: symptomatology and effects on leaf structure and growth of wood plants). The objectives of this study was to characterize the effects of fluoride on seedlings and young plants of native woody species from Parque Estadual do Rio Doce (MG) and on the leaf blade structure of the most sensitive of the studied tree species. This work was carried out to gain information on diagnosing criteria for bioindication and to throw some light on the mechanisms of plant resistance and tolerance in order to assist in the revegetation programs of highly affected areas. The employed species were Gallesia gorazema Moq. (Phytolac- caceae), Genipa americana L. (Rubiaceae), Joannesia princeps Vell. (Euphorbiaceae), Peltophorum dubium (Spreng.) Taub. (Legu- minosae, Caesalpinioideae) and Spondias dulcis Forst. f. (Anarcadiaceae). Seedlings and young plants of those species were submitted to rain with dissolved fluoride (30 mg.L -1 ), during 20 min daily, for 10 consecutive days. Apical and marginal necrosis occurred in the leaf blades of all species just after the first simulated rain. Two days after treatment, S. dulcis, in the young plant stage, showed to be the most sensitive species by exhibiting large necrotic areas, while G. americana was the most resistant species. At the young plant stage, the species that accumulated more fluoride were also the most sensitive ones; such a relation was not observed in plants at the seedling stage. The pollutant at the level employed caused drastic alterations in the leaf blade of S. dulcis and also damaged all of its anatomical structure. The high susceptibility of S. dulcis to fluoride suggests that this species possess potential features to be employed as a bioindicator. However, for a better characterization of the responses of S. dulcis to fluoride and its usage in biomonitoring programs, more studies are required. RESUMO - (Fluor em chuva simulada: sintomatologia e efeitos sobre a estrutura foliar e o crescimento de plantas arboreas). Os objetivos deste trabalho foram: caracterizar os efeitos do fluor em especies arboreas nativas, nos estadios de plântula e muda; identificar injurias provocadas pelo fluor na estrutura da lâmina foliar da especie mais sensivel; fornecer subsidios para selecao de caracteristicas a serem utilizadas na bioindicacao; e contribuir com informacoes sobre a resistencia ou tolerância das plantas, visando ao reflorestamento de areas impactadas pela chuva com fluor. As especies analisadas foram Gallesia gorazema Moq. (Phytolaccaceae), Genipa americana L. (Rubiaceae), Joannesia princeps Vell. (Euphorbiaceae), Peltophorum dubium (Spreng.) Taub. (Leguminosae, Caesalpinioideae) e Spondias dulcis Forst. f. (Anacardiaceae). Plântulas e mudas dessas especies, provenientes do Parque Estadual do Rio Doce (MG), foram submetidas a 20 min diarios de chuva com fluor (30 mg.L -1 ), por 10 dias consecutivos. Necroses apicais e marginais foram observadas em todas as especies analisadas, logo apos a primeira chuva simulada. S. dulcis, no estadio de muda, foi a especie mais sensivel ao fluor, pois apresentou extensas necroses com apenas dois dias de tratamento, enquanto que G. americana foi a especie mais resistente. Nas mudas, as especies que acumularam mais fluor foram tambem as que apresentaram maior sensibilidade a esse poluente; essa relacao nao foi verificada nas plântulas. A concentracao de fluor utilizada promoveu alteracoes drasticas na lâmina foliar de S. dulcis com extensas areas necrosadas, danificando toda a sua estrutura anatomica. A sensibilidade ao fluor observada em S. dulcis indica que essa especie apresenta potencial para ser usada como bioindicadora. Entretanto, estudos detalhados serao necessarios para a melhor caracterizacao das respostas de S. dulcis ao fluor visando a sua utilizacao em programas de biomonitoramento ambiental.

Effects of simulated acid rain on leaf anatomy and micromorphology of Genipa americana L. (Rubiaceae)

Experimentos foram conduzidos para avaliar o grau de susceptibilidade e determinar as injurias causadas pela chuva acida simulada na anatomia e micromorfologia foliar de Genipa americana. Plantas foram expostas a chuva com pH 3,0 durante 10 dias consecutivos. No tratamento controle utilizou-se apenas agua destilada (pH 6,0). Amostras foliares foram coletadas e fixadas para microscopia de luz e eletronica de varredura. Foram observados nas folhas expostas a chuva acida: necroses pontuais intervenais, colapso das celulas do mesofilo e da epiderme; hipertrofia do parenquima lacunoso e acumulo de compostos fenolicos e graos de amido. A analise micromorfologica evidenciou, nas areas necrosadas, plasmolise das celulas-guarda e ruptura da cuticula e da crista estomatica. Alteracoes anatomicas ocorreram antes que sintomas visuais fossem observados nas folhas. Estes resultados comprovam a importância de dados anatomicos na diagnose precoce da injuria e na determinacao da sensibilidade de G. americana a chuva acida.

Leaf anatomy as an additional taxonomy tool for 16 species of Malpighiaceae found in the Cerrado area (Brazil).

This work describes the leaf anatomy of 16 species belonging to three genera of the Malpighiaceae family found in the Cerrado (Minas Gerais State, Brazil). The scope of this study was to support the generic delimitation by contributing to the identification of the species and constructing a dichotomous identification key that includes anatomical characters. The taxonomic characters that were considered to be the most important and used in the identification key for the studied Malpighiaceae species were as follows: the presence and location of glands; presence of phloem in the medullary region of the midrib; mesophyll type; presence and type of trichomes; and presence, quantity, and disposition of accessory bundles in the petiole. It was also possible to indicate promising characters for future taxonomic and phylogenetic studies in the Malpighiaceae family, especially for the Banisteriopsis, Byrsonima, and Heteropterys genera.

Iron plaque formation and morphoanatomy of roots from species of restinga subjected to excess iron

The restingas, a sandy coastal plain ecosystem of Brazil, have received an additional amount of iron due to the activity of mining industries. The present study aims to characterize morphoanatomically and histochemically the iron plaque formation on roots of Ipomoea pes-caprae L. and Canavalia rosea DC, cultivated in hydroponic solution with and without excess iron. The iron plaque formation as well as changes in the external morphology of the lateral roots of both species were observed after the subjection to excess iron. Changes in the nutrient uptake, and in the organization and form of the pericycle and cortex cells were observed for both species. Scanning electron microscopy showed evident iron plaques on the whole surface of the root. The iron was histolocalized in all root tissues of both species. The species of restinga studied here formed iron plaque in their roots when exposed to excess of this element, which may compromise their development in environments polluted by particulated iron.

Diagnostic and prognostic characteristics of phytotoxicity caused by fluoride on Spondias dulcis Forst. F. (Anacardiaceae)

The goal of this study was to determine the symptoms and microscopic damage caused by fluoride on Spondias dulcis, a fluoride-sensitive species. The plants were exposed to simulated fog with fluoride (0, 5, 10, 15 and 20 mg L(-1)) for 20 min daily during four consecutive days. Samples from leaflets without any apparent fluoride injury were collected to microscopic analysis. The percentage of necrosed leaf area was measured, and the level of pollutant in the dry matter from the basal and apical portions of the plant was determined. The necroses began 24 h after the first simulation mainly from the base of the leaflets. A higher level of necrosis was observed at the apical portion of the plants, a region of higher fluoride accumulation. The damage on the surface of the leaflets was characterized as plasmolysis, erosion of the epicuticular waxes and epidermal rupture. Structurally, the noticeable accumulation of granules and droplets green stained by toluidine blue in the spongy parenchima and the boundaries of ending veinlets was observed. The limb thickness reduction occurred due to plasmolysis in the mesophyll, showing an apparent correlation with the damage observed on the surface. The parameters observed in the laboratory are promising for field biomonitoring studies.

Morphoanatomy and development of leaf secretory structures in Passiflora amethystina Mikan (Passifloraceae)

Extrafloral nectaries (EFNs) are commonly found in Passiflora L. Reports have been made on the occurrence of resin-producing structures morphologically similar to EFNs in the genus. The objective of this study was to characterise the morphoanatomy and development of leaf secretory structures in Passiflora amethystina and to use chemical and histochemical tests to detect the presence of sugars in the exudates. Samples of leaf blade and petioles in different developmental stages were collected and subjected to usual techniques using light and scanning electron microscopy. Secretion samples were analysed by high performance liquid chromatography. The concentration of total sugars in the secretion amounted to 39.67% for blade EFNs and 52.82% for petiolar EFNs. EFNs consist of a secretory, uni- or bistratified palisade epidermis, arising from the protoderm by means of anticlinal and periclinal divisions, glandular parenchyma originated from the ground meristem, and xylem and phloem elements formed from the procambium. Exudate accumulated in a subcuticular space formed outside the epidermal cells from where it was then released. Histochemical tests showed a positive reaction for neutral polysaccharides. The results confirm that the leaf secretory structures are indeed extrafloral nectaries, and these findings constitute important information for studies on the taxonomy and ecology of this species.

Effects of simulated acid rain on the growth of five Brazilian tree species and anatomy of the most sensitive species (Joannesia princeps)

Seedlings and young saplings of some woody species were exposed to simulated low-pH acid rain, in order to develop a response screening for tropical tree species by determination of the symptoms of foliar injury and growth responses, as well as to identify anatomical alterations in the leaf blade of the most sensitive species. Gallesia integrifolia (Spreng.) Harms, Genipa americana L., Joannesia princeps Vell., Mimosa artemisiana Heringer & Paula and Spondias dulcis Forst.f. were exposed daily to 20 min of acid rain, pH 3.0, for 10 consecutive days. The degree of leaf damage and the anatomical alterations observed were efficient parameters to determine the sensitivity to acid rain. At the end of the experiment J. princeps was the most sensitive species as determined by foliar injury and seedling growth. The degree of leaf damage was similar among the seedlings, except in S. dulcis, which showed reduced percentage of foliar injury. Necrotic and chlorotic spots on the leaf blade occurred. In the most sensitive species, J. princeps, necrotic blade tissues showed accumulation of phenolic compounds, hypertrophy and collapsed cells. Most of the structural alterations were observed in the adaxial epidermis, the palisade parenchyma and spongy parenchyma and the abaxial epidermis. Long-term experiments with seedlings of S. dulcis and saplings of G. integrifolia are suggested, to characterise the response of these species that presented fewer symptoms but whose growth was affected under acid rain.

Arsenic accumulation in Brassicaceae seedlings and its effects on growth and plant anatomy

We wished to evaluate the effects of arsenic on the morphology and anatomy of Brassica oleracea, Raphanus sativus, Brassica juncea, Brassica oleracea var. capitata and Brassica oleracea var. italica. Seeds were subjected to concentrations 0µM, 250µM, 350µM and 450µM arsenic in the form of sodium arsenate (Na2HAsO4·7H2O) during 12 days. All species accumulated more arsenic in the roots than in the shoots, except for B. oleracea var. capitata. There was no difference of translocation factor between species and treatments. Growth decrease was observed in roots of B. oleracea and R. sativus, and in shoots of R. sativus and B. oleracea var. italica. All species presented anatomical alterations in the roots, such as: cell hypertrophy, protoplast retraction, cellular plasmolysis, and necrotic regions. B. juncea presented collapse and hypertrophy of cells from the leaf blade tissues. Quantitative anatomical analyses performed on the root and leaves of B. oleracea and B. juncea revealed that arsenic interfered on the root vascular cylinder diameter and on height of epidermal cells of the adaxial leaf surface of both species. We concluded that arsenic was absorbed from the culture medium and induced alterations both on root and shoot growth of the seedlings. Retention of arsenic within the root was responsible for major damage in this organ.

Anatomical characteristics as taxonomic tools for the species of Curtia and Hockinia (Saccifolieae–Gentianaceae Juss.)

The delimitation of species and even support of the genera Curtia and Hockinia, monotypic genus, are still controversial. To define anatomical characteristics that help resolve taxonomic problems, the stem and leaf anatomy of seven species of Curtia and Hockinia montana are described. Some characteristics are found in both genera: leaves with uniseriate epidermis, dorsiventral mesophyll, cells of the palisade parenchyma with invaginations, basal acrodromous venation with an incomplete marginal ultimate venation, presence of colleters and nectaries, stem with small wings and internal phloem. The types of stomata, the midrib vascular bundle, and the type and distribution of nectaries were used to separate the genera. The presence/absence of crystals, the sinuosity of the anticlinal walls of the epidermal cells, the type and distribution of stomata, and the distribution of nectaries distinguish species of Curtia. Our results suggest that these anatomical characteristics are potentially useful for clarifying the taxonomy of Gentianaceae, and that they represent an additional tool for distinguishing genera and identifying species of Curtia, particularly pointing out the need to revise the synonymy between C. patula and C. tenuifolia. We also highlight the first report of leaf nectaries and colleters in the Saccifolieae and the first record of stem nectaries in the Gentianaceae.