Molecular systematics of Thorea (Rhodophyta, Thoreales) species in Brazil

This study aimed to evaluate species level taxonomy and phylogenetic relationship among Thorea species in Brazil and other regions of the world using two molecular markers - RUBISCO large subunit plastid gene (rbcL) and nuclear small-subunit ribosomal DNA (SSU rDNA). Three samples of Thorea from Brazil (states of Mato Grosso do Sul and São Paulo) and one sample from Dominican Republic (DR) were sequenced. Analyses based on partial sequences of rbcL (1,282 bp) and complete sequences of SSU (1,752 bp) were essentially congruent and revealed that Thoreales formed a distinct monophyletic clade, which had two major branches with high support, representing the genera Thorea and Nemalionopsis. Thorea clade had four main branches with high support for all analyses, each one representing the species: 1) T. gaudichaudii C. Agardh from Asia (Japan and Philippines) - this clade occurred only in the rbcL analyses; 2) T. violacea Bory from Asia (Japan) and North America (U.S.A. and DR); 3) T. hispida (Thore) Desvaux from Europe (England) and Asia (Japan); 4) a distinct group with the three Brazilian samples (sequence identity: rbcL 97.2%, 1,246 bp; SSU 96.0-98.1%, 1,699-1,720 bp). The Brazilian samples clearly formed a monophyletic clade based on both molecular markers and was interpreted as a separate species, for which we resurrected the name T. bachmannii Pujals. Morphological and molecular evidences indicate that the Thoreales is well-resolved at ordinal and generic levels. In contrast, Thorea species recognized by molecular data require additional characters (e.g. reproductive and chromosome numbers) to allow consistent and reliable taxonomic circumscription aiming at a world revision based on molecular and morphological evidences.

Ensaio in vitro da enzima nitrato redutase e efeito da disponibilidade de nitrato e fosfato em variantes pigmentares de Hypnea musciformis (Wulfen) J. V. Lamour. (Gigartinales, Rhodophyta)

A enzima nitrato redutase (NR) catalisa a redução do nitrato a nitrito e controla a taxa de assimilação do nitrato. O ensaio in vitro da nitrato redutase foi otimizado para a linhagem selvagem (marrom, MA) e para a linhagem deficiente em ficoeritrina (verde-clara, VC) de Hypnea musciformis. As duas linhagens foram cultivadas em temperatura de 23 ± 2°C, fotoperíodo de 14 horas, irradiância de 60-90µmol fótons m-2s-1, e meio composto por água do mar esterilizada (30ups) enriquecida com a solução de von Stosch na concentração de 50% (VSES/2). As condições ótimas de ensaio para ambas as linhagens foram: 40µM de NADH; 10min de incubação do extrato bruto (EB) e 100µL de EB. A atividade ótima da NR ocorreu em 4 e 2mM de nitrato para a linhagem VC e MA, respectivamente. As linhagens VC e MA apresentaram, respectivamente, constante aparente de Michaelis-Menten (K M) para NADH de 0,2068 e 0,0837 µM, e K M para nitrato de 0,0492 e 0,0294mM. Os resultados indicam que a NR da linhagem MA tem maior afinidade pelo substrato do que a NR da linhagem VC de H. musciformis. Os experimentos para avaliar os efeitos da disponibilidade de nitrato (5 a 105µM) e nitrato e fosfato (0,5 a 25,5µM, com a relação N:P de 4:1) mostraram que a atividade da NR das linhagens VC e MA não aumentou com a adição de nitrato no meio, o que pode estar relacionado com o estado nutricional dessas algas. A atividade da NR foi maior nos tratamentos com adição de fosfato do que naqueles com adição de apenas nitrato, indicando que esse nutriente é importante para os processos metabólicos relacionados a atividade da NR.

Phylogenetic analyses of Cladophora vagabunda (L.) C. Hoek (Cladophorales, Chlorophyta) from Brazil based on SSU rDNA sequences

The complete SSU rDNA was sequenced for 10 individuals of Cladophora vagabunda collected along the coast of Brazil. For C. rupestris (L.) Kütz. a partial SSU rDNA sequence (1634 bp) was obtained. Phylogenetic trees indicate that Cladophora is paraphyletic, but the section Glomeratae sensu lato including C. vagabunda from Brazil, Japan and France, C. albida (Nees) Kütz., C. sericea (Hudson) Kütz., and C. glomerata (L.) Kütz. is monophyletic. Within this group C. vagabunda is paraphyletic. The sequence identity for the SSU rDNA varied from 98.9% to 100% for the Brazilian C. vagabunda, and from 98.3% to 99.7% comparing the Brazilian individuals to the ones from France and Japan. Sequence identity of the Brazilian C. vagabunda to C. albida and C. sericea vary from 98.0% to 98.6%. The SSU rDNA phylogeny support partially the morphological characteristics presented by Brazilian populations of C. vagabunda. On the other hand, C. rupestris from Brazil does not group with C. rupestris from France, both sequences presenting only 96.9% of identity. The inclusion of sequences of individuals from Brazil reinforces the need of taxonomical revision for the genus Cladophora and for the complex C. vagabunda.

Diferenciação dos feixes vasculares e dos elementos traqueais no rizoma de algumas Cyperaceae

Foram analisados os rizomas de Bulbostylis paradoxa Ness, Cyperus giganteus Vahl, C. odoratus L., Fuirena umbellata Rottb. e Hypolytrum schraderianum Ness. O corpo primário é resultante da atividade dos meristemas apicais e do meristema de espessamento primário (MEP). Também ocorre crescimento em espessura, que é decorrente da atividade do meristema de espessamento secundário (MES). O procâmbio e o MEP originam feixes colaterais em H. schraderianum e feixes anfivasais nas demais espécies. Entretanto, todos os feixes que têm protofloema e protoxilema são de origem procambial. O MES produz floema e xilema constituindo um tecido vascular único. Elementos de vaso foram encontrados na maioria dos caules em estrutura primária e secundária, com exceção de H. schraderianum que, na estrutura secundária, contém apenas traqueídes, informação que respalda a ocorrência de crescimento secundário nas Cyperaceae. Os elementos de vaso apresentam grande variação morfológica; em estrutura primária, geralmente são mais alongados, com apêndices. Os elementos de vaso do crescimento secundário são relativamente mais curtos, apresentam apêndices e ramificações.

Padrões de distribuição geográfica das espécies de Schefflera J. R. Forst. & G. Forst. (Araliaceae) do Brasil extra-amazônico

Uma análise da distribuição geográfica de Schefflera no Brasil extra-amazônico foi realizada com base em mapas atualizados plotando as ocorrências conhecidas das 26 espécies do gênero encontradas nessa grande área: S. angustissima (Marchal) Frodin, S. aurata Fiaschi, S. botumirimensis Fiaschi & Pirani, S. burchellii (Seem.) Frodin & Fiaschi, S. calva (Cham.) Frodin & Fiaschi, S. capixaba Fiaschi, S. cephalantha (Harms) Frodin, S. cordata (Taub.) Frodin & Fiaschi, S. distractiflora (Harms) Frodin, S. fruticosa Fiaschi & Pirani, S. gardneri (Seem.) Frodin & Fiaschi, S. glaziovii (Taub.) Frodin & Fiaschi, S. grandigemma Fiaschi, S. kollmannii Fiaschi, S. longipetiolata (Pohl ex DC.) Frodin & Fiaschi, S. lucumoides (Decne. & Planch. ex Marchal) Frodin & Fiaschi, S. macrocarpa (Cham. & Schltdl.) Frodin, S. malmei (Harms) Frodin, S. morototoni (Aubl.) Maguire, Steyermark & Frodin, S. racemifera Fiaschi & Frodin, S. ruschiana Fiaschi & Pirani, S. selloi (Marchal) Frodin & Fiaschi, S. succinea Frodin & Fiaschi, S. villosissima Fiaschi & Pirani, S. vinosa (Cham. & Schltdl.) Frodin & Fiaschi e S. aff. varisiana Frodin. Dois centros de endemismo associados com áreas de altitude elevada foram reconhecidos: Cadeia do Espinhaço em Minas Gerais e florestas montanas do Estado do Espírito Santo. Os padrões de distribuição geográfica ilustrados são discutidos com base em dados obtidos para outros grupos de angiospermas e em estudos fitogeográficos das principais fitocórias do Brasil extra-amazônico. São apresentadas também hipóteses acerca de prováveis relações filogenéticas entre alguns táxons, visando à busca de possíveis correlações entre estas e a biogeografia do grupo.

Constituintes químicos e atividade antimicrobiana dos extratos de Dilodendron bipinnatum (sapindaceae)

The phytochemical investigation of ethanolic extracts from leaves, branches and stems of D. bipinnatum afforded the steroids β-sitosterol, stigmasterol, campesterol, sitostenone and sitosterol-3-O-β-D-glycopyranoside, along with two cycloartane triterpenes: cycloeucalenol and 24-methylenecycloartenol. The antimicrobial activity of the extracts was evaluated against Staphylococcus aureus (ATCC 25923), Escherichia coli (ATCC 25922), Bacillus subtilis (ATCC 6623), Pseudomonas aeruginosa (ATCC 15442), Micrococcus luteus (ATCC 9341) and Candida albicans (ATCC 10231). The extracts of the leaves and branches showed moderate activity against Candida albicans. The extract of the branches was active against Micrococcus luteus. This is the first report on the phytochemical study of D. bipinnatum.

Neolignans and sesquiterpenes from leaves and embryogenic cultures of Ocotea Catharinensis (Lauraceae)

The extracts from leaves of Ocotea catharinensis Mez (Lauraceae) were found to contain fourteen neolignans and two sesquiterpenes: nine benzofuran types (including three new compounds 1e, 2f and 4b), one new seco-benzofuran type (3b), two bicyclo[3.2.1]octane types (including the new compound 5c), two new dimers of bicyclo[3.2.1]octane type (7a and 7b) and two sesquiterpenes (including a new humulanol 9). In addition, seven neolignans were also showed to occur in somatic embryos of O. catharinensis including one new bicyclo[3.2.1]octane type (4a).

Two-dimensional gel electrophoretic protein profile analysis during seed development of Ocotea catharinensis: a recalcitrant seed species

The aim of the present work was to characterize changes in the protein profile throughout seed development in O. catharinensis, a recalcitrant species, by two-dimensional gel electrophoresis. Protein extraction was undertaken by using a thiourea/urea buffer, followed by a precipitation step with 10% TCA. Comparative analysis during seed development showed that a large number of proteins were exclusively detected in each developmental stage. The cotyledonary stage, which represents the transition phase between embryogenesis and the beginning of metabolism related to maturation, presents the highest number of stage-specific spots. Protein identification, through MS/MS analysis, resulted in the identification of proteins mainly related to oxidative metabolism and storage synthesis. These findings contribute to a better understanding of protein metabolism during seed development in recalcitrant seeds, besides providing information on established markers that could be useful in defining and improving somatic embryogenesis protocols, besides monitoring the development of somatic embryos in this species.

Comparison of radial growth rate of the mutualistic fungus of Atta sexdens rubropilosa forel in two culture media

In vitro culture of the mutualistic fungus of leaf-cutting ants is troublesome due to its low growth rate, which leads to storage problems and contaminants accumulation. This paper aims at comparing the radial growth rate of the mutualistic fungus of Atta sexdens rubropilosa Forel in two different culture media (Pagnocca B and MEA LP). Although total MEA LP radial growth was greater all along the bioassay, no significant difference was detected between growth efficiencies of the two media. Previous evidences of low growth rate for this fungus were confirmed. Since these data cannot point greater efficiency of one culture medium over the other, MEA LP medium is indicated for in vitro studies with this mutualistic fungus due its simpler composition and translucent color, making the analysis easier.

Reserve carbohydrates and lipids from the seeds of four tropical tree species with different sensitivity to desiccation

Considering the importance of water content for the conservation and storage of seeds, and the involvement of soluble carbohydrates and lipids for embryo development, a comparative study was carried out among the seeds of Inga vera (ingá), Eugenia uniflora (pitanga), both classified as recalcitrant, and Caesalpinia echinata (brazilwood) and Erythrina speciosa (mulungu), considered as orthodox seeds. Low concentrations of cyclitols (0.3-0.5%), raffinose family oligosaccharides (ca. 0.05%) and unsaturated fatty acids (0-19%) were found in the seeds of ingá and pitanga, while larger amounts of cyclitols (2-3%) and raffinose (4.6-13%) were found in brazilwood and mulungu, respectively. These results, in addition to higher proportions of unsaturated fatty acids (53-71%) in orthodox seeds, suggested that sugars and lipids played important role in water movement, protecting the embryo cell membranes against injuries during dehydration.

Physiological and biochemical characterization of the assai palm (Euterpe oleracea Mart.) during seed germination and seedling growth under aerobic and anaerobic conditions

Physiological and biochemical aspects of assai palm during seed germination and early seedling growth were investigated. Seeds collected from plants growing in flooded and upland forests were used to determine the influence of normoxic (aerobic) and anoxic (anaerobic) conditions in germination and the initial and average time of development in the roots and shoots. After 75 days, seedlings germinated under normoxia were transferred to trays and submitted to flooding. Seed reserves (lipids, proteins, soluble sugars and starch) were monitored for quiescent and germinated seeds maintained under normoxic and anoxic conditions, as well as after 5, 10 and 20 days of seedling growth. Alcohol dehydrogenase (ADH) activity was quantified in roots and leaves of seedlings without or with flooding (partial and total). Seeds were not able to germinate under anoxia. Different strategies of storage mobilization of lipids, proteins, soluble sugars and starch were observed in seeds of each environment. ADH activity was induced by anoxia, with the highest level observed in the leaves. This study showed that, under normoxic conditions, the best developmental performance of assai palm seeds, from flooded or upland forest areas, during germination was associated with primary metabolites mobilization and seedling flooding tolerance with increased ADH activity. We conclude that the assai palm is well adapted to the anoxic conditions provoked by flooding.

Polyamine patterns in haploid and diploid tobacco tissues and in vitro cultures

The aim of this work was to determine PAs levels in pith tissues and callus cultures from haploid and diploid tobacco plants, explanted from the apical and basal regions of the stem. These explants were cultured in an RM-64 medium supplied with IAA and kinetin, under light or in the dark, during successive subcultures. PAs levels followed a basipetal decrease in diploid and an increase in haploid, pith tissues. A similar pattern of total PAs (free + conjugated) was observed for the callus of diploid and haploid plants maintained in the light, and for the haploid callus in the dark, whereas the diploid callus in the dark showed a constant increase in total PAs levels until the end of culture. The PA increase in the diploid callus in the dark was related to free Put levels increase. The ploidy status of the plants could express different PA gradients together with the plant pith and in vitro callus cultures.

Morpho-Anatomical Analysis of the Rhizome in Species of Scleria Berg. (Cyperaceae) from Serra do Cipó (MG)

Aspects related to the nature of stem thickening in monocotyledons have been the subject of many studies. Primary thickening has been attributed to the Primary Thickening Meristem (PTM). According to most authors, it gives rise, besides the adventitious roots, to the vascular tissues and part of the cortex. In other words, it has centripetal and centrifugal activity. For some authors, however, it gives rise only to the vascular system, and for others, only to part of the cortex. However, this work demonstrated that PTM corresponds to the pericycle in the meristematic phase or to the pericycle associated with the endodermis, also with meristematic activity. It was observed that the pericycle was responsible for the formation of the vascular system of the rhizome and of the adventitious roots; the endodermis gave rise to cell layers with radial disposition which comprised the inner portion of the stem cortex, and which corresponded to the region known as the derivatives of the meristematic endodermis (DME). A continuity was also demonstrated between the tissues of the stem and root in species of Scleria Berg. (Cyperaceae).

Nutritional reserves of Vochysiaceae seeds: chemical diversity and potential economic uses

Contents of proteins, carbohydrates and oil of seeds of 57 individuals of Vochysiaceae, involving one species of Callisthene, six of Qualea, one of Salvertia and eight of Vochysia were determined. The main nutritional reserves of Vochysiaceae seeds are proteins (20% in average) and oils (21. 6%). Mean of carbohydrate contents was 5. 8%. Callisthene showed the lowest protein content (16. 9%), while Q. cordata was the species with the highest content (30% in average). The contents of ethanol soluble carbohydrates were much higher than those of water soluble carbohydrates. Oil contents lay above 20% for most species (30. 4% in V. pygmaea and V. pyramidalis seeds). The predominant fatty acids are lauric (Q. grandiflora), oleic (Qualea and Salvertia) or acids with longer carbon chains (Salvertia and a group of Vochysia species). The distribution of Vochysiaceae fatty acids suggests for seeds of some species an exploitation as food sources (predominance of oleic acid), for other species an alternative to cocoa butter (high contents or predominance of stearic acid) or the production of lubricants, surfactants, detergents, cosmetics and plastic (predominance of acids with C20 or C22 chains) or biodiesel (predominance of monounsaturated acids). The possibility of exploitation of Vochysiaceae products in a cultivation regimen and in extractive reserves is discussed.

Primary and secondary thickening in the stem of Cordyline fruticosa (Agavaceae)

The growth in thickness of monocotyledon stems can be either primary, or primary and secondary. Most of the authors consider this thickening as a result of the PTM (Primary Thickening Meristem) and the STM (Secondary Thickening Meristem) activity. There are differences in the interpretation of which meristem would be responsible for primary thickening. In Cordyline fruticosa the procambium forms two types of vascular bundles: collateral leaf traces (with proto and metaxylem and proto and metaphloem), and concentric cauline bundles (with metaxylem and metaphloem). The procambium also forms the pericycle, the outermost layer of the vascular cylinder consisting of smaller and less intensely colored cells that are divided irregularly to form new vascular bundles. The pericycle continues the procambial activity, but only produces concentric cauline bundles. It was possible to conclude that the pericycle is responsible for the primary thickening of this species. Further away from the apex, the pericyclic cells undergo periclinal divisions and produce a meristematic layer: the secondary thickening meristem. The analysis of serial sections shows that the pericycle and STM are continuous in this species, and it is clear that the STM originates in the pericycle.The endodermis is acknowledged only as the innermost layer of the cortex.