Factors affecting Cyperus difformis seed germination and seedling emergence

Specific knowledge about the dormancy, germination, and emergence patterns of weed species aids the development of integrated management strategies. Laboratory studies were conducted to determine the effect of several environmental factors on seed germination and seedling emergence of Cyperus difformis. Germination of freshly harvested seeds was inhibited by darkness; however, when seeds were subsequently transferred to complete light they germinated readily. Our results showed that 2 wk of cold stratification overcome the light requirement for germination. Seeds of C. difformis were able to germinate over a broad range of temperatures (25/15, 30/20, 35/25, and 40/30 ºC day/night). The response of germination rate to temperature was described as a non-linear function. Based on model outputs, the base, the optimum and the ceiling temperatures were estimated as 14.81, 37.72 and 45 ºC, respectively. A temperature of 120 ºC for a 5 min was required to inhibit 50% of maximum germination. The osmotic potential and salinity required for 50% inhibition of maximum germination were -0.47 MPa and 135.57 mM, respectively. High percentage of seed germination (89%) was observed at pH=6 and decreased to 12% at alkaline medium (pH 9) pH. Seeds sown on the soil surface gave the greatest percentage of seedling emergence, and no seedlings emerged from seeds buried in soil at depths of 1 cm.

Globe fringerush (Fimbristylis miliacea) cross resistance to als-inhibitor herbicides under field conditions in irrigated rice in the south of Brazil

ALS-inhibiting herbicides usually provide adequate weed control in irrigated rice fields. After consecutive years of use, the Cyperaceae species, globe fringerush (Fimbristylis miliacea) began to show resistance to ALS (acetolactate synthase) inhibitors. Globe fringerush is one of the most problematic herbicide-resistant weeds in irrigated rice in the state of Santa Catarina in the South of Brazil. The objective of this research was to examine cross resistance of globe fringerush to ALS inhibitors, under field conditions. Two experiments were conducted in a rice field naturally infested with ALS-resistant globe fringerush in Santa Catarina, in the 2008/09 and 2009/10 cropping seasons. The experimental units were arranged in randomized complete block design, with five replicates, consisting of two factors (herbicide and dose) in a 4 x 5 factorial arrangement. ALS herbicides included bispyribac-sodium, ethoxysulfuron, pyrazosulfuron-ethyl and penoxsulam. Six-leaf globe fringerush was sprayed with herbicide doses of 0, 0.5, 1, 2 and 4X the recommended doses in a spray volume of 200 L ha-1. The number of rice culm, filled and sterile grains, plant height, dry shoot biomass and grain yield were recorded. Globe fringerush control was evaluated 28 and 70 days after herbicide application (DAA); shoots were harvested at 13 weeks after herbicide application and dry weight recorded. Competition with globe fringerush reduced the number of culm and rice grain yield. The globe fringerush biotype in this field was resistant to all ALS herbicides tested. Penoxsulam had the highest level of activity among treatments at 28 and 70 DAA, but the control level was only 50% and 42%, respectively, in the second year of assessment. This was not enough to prevent rice yield loss. Alternative herbicides and weed control strategies are necessary to avoid yield losses in rice fields infested with ALS-resistant biotypes of globe fringerush.

Occurrence of weeds in Cassava savanna plantations in Roraima

A phyto-sociological survey is the first step to implement integrated weed management in crops. In this study, weed occurrence was evaluated in cassava plantations in the savannah of Roraima in northern Brazil. Harvest was performed randomly 80 times in 10 crops over four seasons (January, February, March, and April 2012). The harvested plants were cut at ground level, sorted out per species, identified, quantified, and weighed on a 0.01 g precision scale. A descriptive analysis was conducted of the phyto-sociological parameters (frequency, density, abundance, total number of individuals per species, relative frequency, relative density, relative abundance and importance value index) for the collected species. A description was also made of the botanical classes, families, species, type of propagation, life cycle, growth habit, total number of species and dry weight ha-1. The community in the surveyed area was considered to have a heterogeneous composition, comprising 27 species. The species presenting the highest density per hectare were Digitaria sanguinalis (210,500), Brachiaria brizantha (111,000), Brachiaria decumbens (86,500) and Brachiaria humidicola (69,000). Digitaria sanguinalis had the highest relative density (28.08), relative abundance (26.16) and importance value index (65,34). Most weeds had herbaceous growth habit.

Weed Community in a Raw Sugarcane Renovation Area Submitted to Different Soil Managements

The objective of this study was to evaluate the weed community in a raw sugarcane renovation area with three soil managements and peanut sowing in succession. The experiment was conducted during the 2007/08 season on a raw sugarcane area harvested without prior burning in the last five cuts. A randomized block design with treatments arranged in a split plot and arranged in four replications was used. The main treatments consisted of three cropping systems: conventional tillage, minimum tillage and direct planting, and subplots consisted in the absence (resting) or presence of crop rotation with peanuts. After 135 days from planting peanuts and 180 days of sugarcane harvest, the number of weeds m-2 was counted and the shoot dry biomass of the weeds collected was determined. The data were interpreted by analysis of variance and the means were compared by Tukey's test at 5% probability so that phytosociological indices a, b, c e d were calculated. The use of soil conservation tillage and peanut in rotation with sugarcane in the renovation areas is effective in controlling weeds and suppression of weed species difficult to control like Cyperus rotundus, Commelina bengalensis, Urochloa plantaginea, and Digitaria nuda.

High irradiance and temperature inhibit the germination of spores of the fern Rumohra adiantiformis (Forst.) Ching (Dryopteridaceae)

Rumohra adiantiformis (Forst.) Ching is a fern (Dryopteridaceae) which is used to compose floral arrangements. Fertile fronds were harvested in the "Permanently Protected Area" of Ilha Comprida, São Paulo, Brazil. Sterilized spores were germinated in Mohr liquid medium modified by Dyer. The effect of 72%, 54%, 17% and 9% of total irradiance on germination under field conditions, was analyzed. Experiments were carried out in March (I), April (II) and August of 2000 (III). Under 54% and 72% of total irradiance in Experiment I (March) the germination was completely inhibited and partially inhibited under 72% of total irradiance in Experiment II (April). The lowest mean germination time () was observed under 9% of total irradiance in Experiments II (11.62 days) and III (8.80 days) respectively, followed by 17% in Experiment III (10.12 days) and 9% of total irradiance in the Experiment I (11.62 days ). The effect of temperatures of 15 ± 1, 20 ± 1, 25 ± 1 and 30 ± 1 ºC on germination was also analyzed. The lowest mean germination time (7.93 days) was observed at 25 ± 1 °C followed by 20 ± 1 °C. The highest mean germination time was observed at 15 ± 1 °C (12.10 days) followed by 30 ± 1 °C (10.63 days), which inhibited germination. The germination of R. adiantiformis was photoinhibited by high irradiance and partially inhibited by the highest temperature tested.

Harvesting of Protium heptaphyllum (Aubl.) March. seeds (Burseraceae) by the leaf-cutting ant Atta sexdens L. promotes seed aggregation and seedling mortality

The role played by leaf-cutting ants as seed dispersers of non-myrmecochorous plants remains poorly understood. Here we document the harvesting of Protium heptaphyllum (Aubl.) March. seeds (Burseraceae) by the leaf-cutting ant Atta sexdens L. and its consequences for (1) seed deposition pattern; (2) seed germination; and (3) seedling mortality. The study was carried out at Dois Irmãos, a 390 ha reserve of Atlantic forest, northeast Brazil. Ant-seed harvesting on the ground was detected in 18.5% of all fruiting trees and ants harvested 41.1% ± 19.7% of the seed crop (mean ± s). In average, ants piled seeds 3.4 ± 2.2 m away from the trunk of parent trees and seed density in these piles reached 128.8 ± 138.8 seeds 0.25 m² during the peak of seed discarding by ants. During a 13 month period, mean seedling mortality varied from 0.54% up to 10.6% in ant-made seed piles vs. 0.05-4.2% in control samples, what resulted in a total seedling mortality of 97.7% vs. 81%. Ants systematically cut seedling epicotyls, accounting for 55% of seedling mortality in seed piles, whereas only 14 seedlings (4.2%) were cut by ants in the control samples. Our results suggest that seed harvesting by A. sexdens (1) affects approximately 20% of fruiting P. heptaphyllum trees and their seed crops; (2) promotes short-distance seed dispersal and high levels of seed aggregation; and (3) reduces seedling survival beneath parents.

Leaf anatomy of two Anemia Sw. species (Schizaeaceae-Pteridophyte) from a rocky outcrop in Niterói, Rio de Janeiro, Brazil

The ferns Anemia tomentosa (Sav.) Sw. var. anthriscifolia (Schrad.) Mickel and Anemia villosa Humb. & Bonpl. ex Willd. are widely associated with vegetation islands on rocky outcrops in Rio de Janeiro. Both species are desiccation tolerant. The leaf anatomy of these species was examined aiming to identify morphological characteristics that would allow the establishment of these species in water-scarce environments. The plants were harvested on "Pedra de Itacoatiara" and prepared according to the usual procedures. The petiole has a uniseriate epidermis with lignified cell walls, conical stegmata, and uniseriate multicelular and glandular trichomes. In A. villosa, the stomata protrude in a respiratory line. Under the epidermis the cells have thick, lignified walls. The parenchyma has phenolic compounds and starch grains. The petiole vascular bundles are surrounded by endodermis with Casparian strips and the xylem is V-shaped (A. villosa) or arc-shaped (A. tomentosa var. anthriscifolia). The leaf blades have a uniseriate epidermis with sinuous anticlinal and convex periclinal walls, conical stegmata and chloroplasts on both surfaces. The leaf margins of A. villosa have lignified cells. The guard cells of the stomata on the abaxial surface are on the same level or are raised above ordinary epidermal cells. Multicelular uniseriate trichomes and glandular hairs were observed. The dorsiventral mesophyll has loosely packed chlorenchyma with arm-shaped and H-shaped cells. The vascular bundles are surrounded by endodermis with Casparian strips and with parenchymatic extensions towards the epidermis. Anatomical results were analyzed considering the interaction of these plants with abiotic factors.

Assessment of the degree of contamination of rat germ cell preparations using specific cDNA probes

Recent reports showing a decrease in sperm count in men have brought new concerns about male infertility. Animal models have been widely used to provide some relevant information about the human male gamete, and extrapolations are made to men and to the clinical context. The present study assesses one of the methods used for separation of germ cells of the adult rat testis, namely centrifugal elutriation followed by density gradients (Percoll®). This method was chosen since it presents the best results for cell purity in separating germ cells from the rat testis. A comparison between continuous and discontinuous Percoll® gradients was performed in order to identify the best type of gradient to separate the cells. Maximal cell purity was obtained for spermatocytes (81 ± 8.2%, mean ± SEM) and spermatids (84 ± 2.6%) using centrifugal elutriation followed by continuous Percoll® gradients. A significant difference in purity was observed between elongating spermatids harvested from continuous Percoll® gradients and from discontinuous gradients. Molecular analysis was used to assess cell contamination by employing specific probes, namely transition protein 2 (TP2), mitochondrial cytochrome C oxidase II (COX II), and sulfated glycoprotein 1 (SGP1). Molecular analysis of the samples demonstrated that morphological criteria are efficient in characterizing the main composition of the cell suspension, but are not reliable for identifying minimal contamination from other cells. Reliable cell purity data should be established using molecular analysis

Cytokine profiles during experimental Chagas' disease

People infected with Trypanosoma cruzi remain so for life, yet only 30-40% of these individuals develop characteristic chagasic cardiomyopathies. Similarly, when infected with the Brazilian strain of T. cruzi, DBA/2 mice develop severe cardiac damage while B10.D2 mice do not. To better understand the immunological parameters that may be involved in the disease process, we have used this murine model (DBA/2 vs B10.D2) and compared the changes in cytokine production during the course of infection with T. cruzi. Concanavalin A (Con A) stimulation of spleen cells harvested during the acute phase (day 30) resulted in similarly high levels of IFN-g in both mouse strains. However, the amount of IFN-g in supernatants from cultures of B10.D2 spleen cells initiated during the chronic phase (day 72) was at subacute levels, whereas secretion by chronic DBA/2 spleen cells remained high. In addition, Con A-stimulated spleen cells from acute DBA/2 mice produced approximately twice as much IL-10 and significantly more IL-4 than cells from B10.D2 mice. IL-4 secretion remained low by cells from chronic B10.D2 mice, but when using cells from chronic DBA/2 mice, levels continued to increase beyond the already high levels secreted by cells harvested during the acute phase. Proliferative responses to Con A stimulation by spleen cells from DBA/2 mice were significantly higher than those from B10.D2 mice in both the acute and chronic phases. These data suggest that enhanced responses in DBA/2 mice, which may be related to a higher parasite burden, a lack of down-regulation, and/or the onset of autoimmune phenomena, correlate with the more severe cardiomyopathy seen in pathopermissive mice.

Immunologic patterns associated with cure in human American cutaneous leishmaniasis

Patients with American cutaneous leishmaniasis were studied before therapy (active lesion) and at the end of therapy (cured patients). Assays of lymphocyte proliferative responses of peripheral blood mononuclear cells induced in vitro by Leishmania braziliensis promastigote antigens (Lb) were performed. Antigen-stimulated cells were harvested for CD4 and CD8 phenotype analysis and the levels of gamma interferon (IFN-g) and interleukin 4 (IL-4) produced were also determined in the culture supernatants. Two different patterns of Lb-induced T cell responses were observed: a) predominance of responding CD4+ cells and mixed type 1 and type 2 cytokine production (IFN-g and IL-4) during the active disease, and b) similar proportions of responding CD4+ and CD8+ cells, and type 1 cytokine production (presence of IFN-g and very low IL-4) at the end of therapy (healed lesions). This last pattern is probably associated with a beneficial T cell response

Cyanobacterial toxins in Portugal: effects on aquatic animals and risk for human health

Toxic cyanobacteria are common in Portuguese freshwaters and the most common toxins are microcystins. The occurrence of microcystin-LR (MCYST-LR) has been reported since 1990 and a significant number of water reservoirs that are used for drinking water attain high levels of this toxin. Aquatic animals that live in eutrophic freshwater ecosystems may be killed by microcystins but in many cases the toxicity is sublethal and so the animals can survive long enough to accumulate the toxins and transfer them along the food chain. Among these, edible mollusks, fish and crayfish are especially important because they are harvested and sold for human consumption. Mussels that live in estuarine waters and rivers where toxic blooms occur may accumulate toxins without many significant acute toxic effects. In this study data are presented in order to understand the dynamics of the accumulation and depuration of MCYST-LR in mussels. The toxin is readily accumulated and persists in the shellfish for several days after contact. In the crayfish the toxin is accumulated mainly in the gut but is also cleared very slowly. In carps, although the levels of the toxins found in naturally caught specimens were not very high, some toxin was found in the muscle and not only in the viscera. This raises the problem of the toxin accumulation by fish and possible transfer through the food chain. The data gathered from these experiments and from naturally caught specimens are analyzed in terms of risk for human consumption. The occurrence of microcystins in tap water and the incidence of toxic cyanobacteria in fresh water beaches in Portugal are reported. The Portuguese National Monitoring Program of cyanobacteria is mentioned and its implications are discussed.

Efficacy, safety, quality control, marketing and regulatory guidelines for herbal medicines (phytotherapeutic agents)

This review highlights the current advances in knowledge about the safety, efficacy, quality control, marketing and regulatory aspects of botanical medicines. Phytotherapeutic agents are standardized herbal preparations consisting of complex mixtures of one or more plants which contain as active ingredients plant parts or plant material in the crude or processed state. A marked growth in the worldwide phytotherapeutic market has occurred over the last 15 years. For the European and USA markets alone, this will reach about $7 billion and $5 billion per annum, respectively, in 1999, and has thus attracted the interest of most large pharmaceutical companies. Insufficient data exist for most plants to guarantee their quality, efficacy and safety. The idea that herbal drugs are safe and free from side effects is false. Plants contain hundreds of constituents and some of them are very toxic, such as the most cytotoxic anti-cancer plant-derived drugs, digitalis and the pyrrolizidine alkaloids, etc. However, the adverse effects of phytotherapeutic agents are less frequent compared with synthetic drugs, but well-controlled clinical trials have now confirmed that such effects really exist. Several regulatory models for herbal medicines are currently available including prescription drugs, over-the-counter substances, traditional medicines and dietary supplements. Harmonization and improvement in the processes of regulation is needed, and the general tendency is to perpetuate the German Commission E experience, which combines scientific studies and traditional knowledge (monographs). Finally, the trend in the domestication, production and biotechnological studies and genetic improvement of medicinal plants, instead of the use of plants harvested in the wild, will offer great advantages, since it will be possible to obtain uniform and high quality raw materials which are fundamental to the efficacy and safety of herbal drugs.

Insulin impairs the maturation of chondrocytes in vitro

The precise nature of hormones and growth factors directly responsible for cartilage maturation is still largely unclear. Since longitudinal bone growth occurs through endochondral bone formation, excess or deficiency of most hormones and growth factors strongly influences final adult height. The structure and composition of the cartilaginous extracellular matrix have a critical role in regulating the behavior of growth plate chondrocytes. Therefore, the maintenance of the three-dimensional cell-matrix interaction is necessary to study the influence of individual signaling molecules on chondrogenesis, cartilage maturation and calcification. To investigate the effects of insulin on both proliferation and induction of hypertrophy in chondrocytes in vitro we used high-density micromass cultures of chick embryonic limb mesenchymal cells. Culture medium was supplemented with 1% FCS + 60 ng/ml (0.01 µM) insulin and cultures were harvested at regular time points for later analysis. Proliferating cell nuclear antigen immunoreactivity was widely detected in insulin-treated cultures and persisted until day 21 and [³H]-thymidine uptake was highest on day 14. While apoptosis increased in control cultures as a function of culture time, terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL)-labeled cells were markedly reduced in the presence of insulin. Type II collagen production, alkaline phosphatase activity and cell size were also lower in insulin-treated cultures. Our results indicate that under the influence of 60 ng/ml insulin, chick chondrocytes maintain their proliferative potential but do not become hypertrophic, suggesting that insulin can affect the regulation of chondrocyte maturation and hypertrophy, possibly through an antiapoptotic effect.

Interleukin-5 mediates peritoneal eosinophilia induced by the F1 cell wall fraction of Histoplasma capsulatum

An alkali-insoluble fraction 1 (F1), which contains mainly ß-glucan isolated from the cell wall of Histoplasma capsulatum, induces eosinophil recruitment into the peritoneal cavity of mice. The present study was carried out to determine the participation of interleukin-5 (IL-5) in this process. Inbred C57BL/6 male mice weighing 15-20 g were treated ip with 100 µg of anti-IL-5 monoclonal antibody (TRFK-5, N = 7) or an isotype-matched antibody (N = 7), followed by 300 µg F1 in 1 ml PBS ip 24 h later. Controls (N = 5) received only 1 ml PBS. Two days later, cells from the peritoneal cavity were harvested by injection of 3 ml PBS and total cell counts were determined using diluting fluid in a Neubauer chamber. Differential counts were performed using Rosenfeld-stained cytospin preparations. The F1 injection induced significant (P < 0.01) leukocyte recruitment into the peritoneal cavity (8.4 x 10(6) cells/ml) when compared with PBS alone (5.5 x 10(6) cells/ml). Moreover, F1 selectively (P < 0.01) induced eosinophil recruitment (1 x 10(6) cells/ml) when compared to the control group (0.07 x 10(6) cells/ml). Treatment with TRFK-5 significantly (P < 0.01) inhibited eosinophil recruitment (0.18 x 10(6) cells/ml) by F1 without affecting recruitment of mononuclear cells or neutrophils. We conclude that the F1 fraction of the cell wall of H. capsulatum induces peritoneal eosinophilia by an IL-5-dependent mechanism. Depletion of this cytokine does not have effect on the recruitment of other cell types induced by F1.

Maturation of pig oocytes in vitro in a medium with pyruvate

The aim of in vitro maturation oocyte systems is to produce oocytes of comparable quality to those derived in vivo. The present study was designed to examine the surface morphological changes of the cumulus-oocyte complex (COC) and nuclear maturation in a culture system containing pyruvate. Ovaries were obtained from a slaughterhouseand transported to the laboratory within 2 h at 35-39ºC,and rinsed three times in 0.9% NaCl. The COCs were harvested from the ovaries and in vitro maturation was evaluated in San Marcos (SM) medium, a chemically defined culture system containing 22.3 mM sodium pyruvate. Oocytes were cultured in SM, SM + porcine follicular fluid (pFF) and in SM + pFF + gonadotropins (eCG and hCG) for 20-22 h and then without hormonal supplements for an additional 20-22 h. After culture, the degree of cumulus expansion and frequency of nuclear maturation were determined. Oocytes matured in SM (40.9%) and SM + pFF (42.9%) showed moderate cumulus expansion, whereas oocytes matured in SM + pFF + gonadotropins (54.6%) showed high cumulus expansion. The maturation rate of cultured oocytes, measured in function of the presence of the polar corpuscle, did not differ significantly between SM (40.9 ± 3.6%) and SM + pFF (42.9 ± 3.7%). These results indicate that pig oocytes can be successfully matured in a chemically definedmedium and suggest a possible bifunctional role of pyruvate as an energy substrate and as an antioxidant protecting oocytes against the stress of the in vitro environment.