Molecular characterization of two rocio flavivirus strains isolated during the encephalitis epidemic in são paulo state, brazil and the development of a one-step rt-pcr assay for diagnosis

Rocio virus (ROCV) was responsible for an explosive encephalitis epidemic in the 1970s affecting about 1,000 residents of 20 coastland counties in São Paulo State, Brazil. ROCV was first isolated in 1975 from the cerebellum of a fatal human case of encephalitis. Clinical manifestations of the illness are similar to those described for St. Louis encephalitis. ROCV shows intense antigenic cross-reactivity with Japanese encephalitis complex (JEC) viruses, particularly with Ilheus (ILHV), St. Louis encephalitis, Murray Valley and West Nile viruses. In this study, we report a specific RT-PCR assay for ROCV diagnosis and the molecular characterization of the SPAn37630 and SPH37623 strains. Partial nucleotide sequences of NS5 and E genes determined from both strains were used in phylogenetic analysis. The results indicated that these strains are closely related to JEC viruses, but forming a distinct subclade together with ILHV, in accordance with results recently reported by Medeiros et al. (2007).

Neurotoxic activity and ultrastructural changes in muscles caused by the brown widow spider Latrodectus geometricus venom

Brown widow spider (Latrodectus geometricus) venom (BrWSV) produces few local lesions and intense systemic reactions such as cramps, harsh muscle pains, nausea, vomiting and hypertension. Approximately 16 protein bands under reducing conditions and ~ 14 bands under non-reducing conditions on a 12.5% sodium dodecyl sulfate-polyacrylamide gel electrophoresis were observed. Neurotoxic clinical manifestations were confirmed in vivo, while proteolytic activity was demonstrated on gelatine film. Severe ultrastructural damages in mice skeletal muscles were observed at 3, 6, 12 and 24 h postinjection with at total of 45 µg of venom protein. Infiltration of eosinophils and ruptures of the cellular membranes were observed in the muscles along with swelling of the nuclear cover and interruption of the collagen periodicity. Altered mitochondrias and autophage vacuoles, nuclear indentation and mitochondria without cristae, slight increment of intermyofibrillar and subsarcolemic spaces and myelinic figures formation were also observed. In the capillary, endothelial membrane unfolding into the lumen was noticed; along with myelinic figures compatible with a toxic myopathy. Swollen sarcotubular systems with lysis of membrane, intense mitochondria autophagia and areas without pinocytic vesicles were observed. Swollen mitochondria surrounded by necrotic areas, myofibrillar disorganization and big vacuolas of the sarcotubular system, degenerated mitochondrium with formation of myelinic figure was seen. Glycogenosomes with small particulate, muscle type glycogen was noticed. Autophagic vacuole (autophagolysosomes) and necrotic areas were also noticed. These damages may be due to interactive effects of the multifactorial action of venom components. However, Latrodectus geometricus venom molecules may also be utilized as neuro therapeutic tools, as they affect neuronal activities with high affinity and selectivity. To our knowledge, the present study is the first ultrastructural report in the literature of muscle injuries and neurological and proteolytic activities caused by BrWSV.

In vitro antimalarial activity and cytotoxicity of some selected cuban medicinal plants

Terrestrial plants have been demonstrated to be sources of antimalarial compounds. In Cuba, little is known about antimalarial potentials of plant species used as medicinals. For that reason, we evaluated the antimalarial activity of 14 plant species used in Cuba as antimalarial, antipyretic and/or antiparasitic. Hydroalcoholic extracts were prepared and tested in vitro for the antimalarial activity against Plasmodium falciparum Ghana strain and over human cell line MRC-5 to determine cytotoxicity. Parasite multiplication was determined microscopically by the direct count of Giemsa stained parasites. A colorimetric assay was used to quantify cytotoxicity. Nine extracts showed IC50 values lower than 100 µg/mL against P. falciparum, four extracts were classified as marginally active (SI < 4), one as partially active (Parthenium hysterophorus) exhibiting SI equal to 6.2 and two extracts as active (Bambusa vulgaris and Punica granatum), showing SI > 10. B. vulgaris showed the most potent and specific antiplasmodial action (IC50 = 4.7 µg/mL, SI = 28.9). Phytochemical characterization of active extracts confirmed the presence of triterpenoids in B. vulgaris and polar compounds with phenol free groups and fluorescent metabolites in both extracts as major phytocompounds, by thin layer chromatography. In conclusion, antimalarial use of B. vulgaris and P. hysterophorus was validated. B. vulgaris and P. granatum extracts were selected for follow-up because of their strong antimalarial activity.

Induction of phagocytic activity and nitric-oxide production in natural populations of Trypanosoma Cruzi I and II from the state of Paraná, Brazil

Twelve strains of Trypanosoma cruzi isolated from wild reservoirs, triatomines, and chronic chagasic patients in the state of Paraná, southern Brazil, and classified as T. cruzi I and II, were used to test the correlation between genetic and biological diversity. The Phagocytic Index (PI) and nitric-oxide (NO) production in vitro were used as biological parameters. The PI of the T. cruzi I and II strains did not differ significantly, nor did the PI of the T. cruzi strains isolated from humans, triatomines, or wild reservoirs. There was a statistical difference in the inhibition of NO production between T. cruzi I and II and between parasites isolated from humans and the strains isolated from triatomines and wild reservoirs, but there was no correlation between genetics and biology when the strains were analyzed independently of the lineages or hosts from which the strains were isolated. There were significant correlations for Randomly Amplified Polymorphic Deoxyribonucleic acid (RAPD) and biological parameters for T. cruzi I and II, and for humans or wild reservoirs when the lineages or hosts were considered individually.

Leishmanicidal activity and cytotoxicity of compounds from two Annonacea species cultivated in Northeastern Brazil

INTRODUCTION: Visceral leishmaniasis is endemic in 88 countries, with a total of 12 million people infected and 350 million at risk. In the search for new leishmanicidal agents, alkaloids and acetogenins isolated from leaves of Annona squamosa and seeds of Annona muricata were tested against promastigote and amastigote forms of Leishmania chagasi. METHODS: Methanol-water (80:20) extracts of A. squamosa leaves and A. muricata seeds were extracted with 10% phosphoric acid and organic solvents to obtain the alkaloid and acetogenin-rich extracts. These extracts were chromatographed on a silica gel column and eluted with a mixture of several solvents in crescent order of polarity. The compounds were identified by spectroscopic analysis. The isolated compounds were tested against Leishmania chagasi, which is responsible for American visceral leishmaniasis, using the MTT test assay. The cytotoxicity assay was evaluated for all isolated compounds, and for this assay, RAW 264.7 cells were used. RESULTS: O-methylarmepavine, a benzylisoquinolinic alkaloid, and a C37 trihydroxy adjacent bistetrahydrofuran acetogenin were isolated from A. squamosa, while two acetogenins, annonacinone and corossolone, were isolated from A. muricata. Against promastigotes, the alkaloid showed an IC50 of 23.3 µg/mL, and the acetogenins showed an IC50 ranging from 25.9 to 37.6 µg/mL; in the amastigote assay, the IC50 values ranged from 13.5 to 28.7 µg/mL. The cytotoxicity assay showed results ranging from 43.5 to 79.9 µg/mL. CONCLUSIONS: These results characterize A. squamosa and A. muricata as potential sources of leishmanicidal agents. Plants from Annonaceae are rich sources of natural compounds and an important tool in the search for new leishmanicidal therapies.

Evaluation of larvicidal activity and brine shrimp toxicity of rhizome extracts of Zingiber zerumbet (L.) Smith

Introduction In this study, we used dichloromethane (DCM) and methanol (MeOH) extracts of the Zingiber zerumbet rhizome to evaluate brine shrimp lethality and larvicidal activity on Aedes aegypti and Anopheles nuneztovari mosquitoes. Methods Bioassays were performed by exposing third-instar larvae of each mosquito species to the DCM or MeOH extracts. Results Probit analysis with DCM and MeOH extracts demonstrated efficient larvicidal activity against A. aegypti and A. nuneztovari larvae. Conclusions The DCM and MeOH extracts showed higher activity against A. nuneztovari larvae than against A. aegypti larvae, suggesting that the extracts have species-specific activity.

Geomorphology and land development in the Maraca area of northern Roraima, Brazil.

The geomorphological materials and forms of the Maraca area of Roraima, Brazil are described, an their sgnificance for land development examined. Significant contrasts are noted in areas presently under rainforest and savanna vegetation. Lateritic gravels and extensive shetwash accumulations in savanna areas constrast with incipient or absent plinthite development, few gravels and limited evidence of colluvium under rainforest. Terrain is in general relatively highly-dissected. Slope profiles are characterised, particularly within the savanna zone, by a relatively steep lower concavity. These contrasts are sharply-demarcated by the present savanna/rainforest bondary, unexpectedly in view of the generally accepted hypothesis of repeated contraction an expansion of Amazonian rainforest throughout the Pleistocene. It is concluded that geomorphological conditions in the Maraca area are not favorable for land develoment.

The relationship between stem vessel parameters and the development of strata in the early stages of secondary forest succession in Amazonia

In order to compare the development of strata in the early stages of secondary forest succession with vessel parameters of the tree species, a forest inventory was carried out in 4-year (Q1: 48 m2), 11-year (Q2: 400 m2) and 20-year (Q3: 400 m2) forests and vessel parameters were investigated from stem cross sections of 18 species obtained in Q2. Thirty three species (21 families), 77 species (35 families), 39 species (20 families) were found in Ql, Q2, Q3, respectively. The percentage of dead individuals, dead stems and the percentage of individuals with multiple stems increased with time after clear cutting. Also, the total D2H of Q3 was 26.1 times that of Q1, and the development of strata started in Q2 and Q3. The image analysis of vessel size, area and number of vessels revealed that species which reach the forest canopy had a large D2H value, vessel diameter and area, while species which remain near the forest floor had smaller ones. Poecilanthe effusa (Huber) Ducke is an example of the latter case, with a large number of individuals and abundant sprouting of new stems from stumps, but with high mortality.

In vitro screening of Amazonian plants for hemolytic activity and inhibition of platelet aggregation in human blood

In the present study, different aerial parts from twelve Amazonian plant species found in the National Institute for Amazon Research's (INPA's) Adolpho Ducke Forest Reserve (in Manaus, Amazonas, Brazil) were collected. Separate portions of dried, ground plant materials were extracted with water (by infusion), methanol and chloroform (by continuous liquid-solid extraction) and solvents were removed first by rotary evaporation, and finally by freeze-drying which yielded a total of seventy-one freeze-dried extracts for evaluation. These extracts were evaluated initially at concentrations of 500 and 100 µg/mL for in vitro hemolytic activity and in vitro inhibition of platelet aggregation in human blood, respectively. Sixteen extracts (23 % of all extracts tested, 42 % of all plant species), representing the following plants: Chaunochiton kappleri (Olacaceae), Diclinanona calycina (Annonaceae), Paypayrola grandiflora (Violaceae), Pleurisanthes parviflora (Icacinaceae), Sarcaulus brasiliensis (Sapotaceae), exhibited significant inhibitory activity towards human platelet aggregation. A group of extracts with antiplatelet aggregation activity having no in vitro hemolytic activity has therefore been identified. Three extracts (4 %), all derived from Elaeoluma nuda (Sapotaceae), exhibited hemolytic activity. None of the plant species in this study has known use in traditional medicine. So, these data serve as a baseline or minimum of antiplatelet and hemolytic activities (and potential usefulness) of non-medicinal plants from the Amazon forest. Finally, in general, these are the first data on hemolytic and inhibitory activity on platelet aggregation for the genera which these plant species represent.

Body image dissatisfaction and its relationship with physical activity and body mass index in Brazilian adolescents

Objective: To evaluate body image dissatisfaction and its relationship with physical activity and body mass index in a Brazilian sample of adolescents. Methods: A total of 275 adolescents (139 boys and 136 girls) between the ages of 14 and 18 years completed measures of body image dissatisfaction through the Contour Drawing Scale and current physical activity by the International Physical Activity Questionnaire. Weight and height were also measured for subsequent calculation of body mass index. Results: Boys and girls differed significantly regarding body image dissatisfaction, with girls reporting higher levels of dissatisfaction. Underweight and eutrophic boys preferred to be heavier, while those overweight preferred be thinner and, in contrast, girls desired to be thinner even when they are of normal weight. Conclusion: Body image dissatisfaction was strictly related to body mass index, but not to physical activity.

Daily activity and microhabitat use of sympatric lizards from Serra do Cipó, southeastern Brazil

We studied the influence of seasonality on the daily activity pattern and microhabitat use of three sympatric lizard species, Cnemidophorus ocellifer Spix, 1825 (Teiidae), Tropidurus montanus Rodrigues, 1987 and Eurolophosaurus nanuzae (Rodrigues, 1981) (Tropiduridae), in an area of campos rupestres (rocky fields) habitat in state of Minas Gerais, Brazil. Cnemidophorus ocellifer exhibited low density and activity concentrated within the hottest hours of the day, and was observed mainly on shaded rocks. Tropidurus montanus and E. nanuzae had similar activity patterns that did not vary between seasons. Activity of T. montanus was related to environmental temperatures. However, we did not find such relationships for E. nanuzae during the dry season. Both T. montanus and E. nanuzae were sighted mainly on exposed rocks. Extension of activity varied between seasons, shorter for C. ocellifer and longer for T. montanus and E. nanuzae during the rainy season.

Thermal biology, activity, and population parameters of Cnemidophorus vacariensis (Squamata, Teiidae), a lizard endemic to southern Brazil

We investigated the following aspects of the biology of a population of Cnemidophorus vacariensis Feltrim & Lema, 2000 during the four seasons: thermal biology, relationship with the thermal environment, daily and seasonal activity, population structure and growth rate. Cnemidophorus vacariensis is restricted to rocky outcrops of the "campos de cima da serra" grasslands on the Araucaria Plateau, southern Brazil, and is currently listed as regionally and nationally threatened with extinction. Data were collected from October 2004 through September 2007 in the state of Rio Grande do Sul. Sampling was conducted randomly from 08:00 a.m. to 6:00 p.m. The capture-mark-recapture method was employed. The lizards were captured by hand, and their cloacal temperature, sex, snout-ventral length (SVL), mass, and the temperature of their microhabitat (substrate temperature and air temperature) were recorded. Individuals were then marked by toe-clipping and released at the site of capture. Body temperatures were obtained for 175 individuals, activity data for 96 individuals, and data on population structure and growth for 59 individuals. All data were obtained monthly, at different times of the day. Cnemidophorus vacariensis average body temperature was 23.84ºC, ranging between 9.6 and 38.2ºC. Temperatures ranged between 21 and 29ºC. The correlation between external heat sources, substrate and air were positive and significant and there was a greater correlation between lizard's temperature and the temperature of the substrate (tigmothermic species). The relatively low body temperatures of individuals are associated with the climate of their environment (altitude up to 1,400 m), with large variations in temperature throughout the day and the year, and low temperatures in winter. The average body temperature observed for C. vacariensis was low when compared with that of phylogenetically related species, suggesting that the thermal biology of this species reflects adaptations to the temperate region where it lives. The monthly rates of activity of lizards were related to monthly variations in the ambient temperatures. Our data suggest that the daily and seasonal activity of C. vacariensis result from the interaction between two factors: changes in the environment temperature and the relationship between individuals and their thermal environment. The population structure of C. vacariensis varied throughout the study period, with maximum biomass in January and maximum density in February (recruitment period). The sex ratio diverged from the expected 1:1. The growth analysis showed a negative relationship between the growth rate of individuals and the SVL, revealing that young individuals grow faster than adults, a typical pattern for short-lived species. The population studied showed a seasonal and cyclical variation associated with the reproductive cycle. The life strategy of C. vacariensis seems to include adaptations to the seasonal variations in temperature, typical of its environment.

Influence of seasonal changes in daily activity and annual life cycle of Geotrigona mombuca (Hymenoptera, Apidae) in a Cerrado habitat, São Paulo, Brazil

The foraging activity of Geotrigona mombuca Smith, 1863 was studied under natural conditions aiming to verify the influence of seasonal changes on daily flight activity and annual cycle of the colony. Daily flight activity was monitored for a year based on the observation and counting of foragers leaving and entering the hive, as well as the kind of material transported and meteorological factors such as day time, temperature and relative humidity. The influence of seasonal changes was evidenced by alterations on daily rhythm of flight activity and by differences on transportation of food resources, building material and garbage. These data indicate that forager behavior is related to daily microclimate conditions and it is synchronized with the requirements of colony annual cycle, which determines an intense pollen collection in the summer. Thus, the recomposition of the intranidal population in spring and summer can be ensured, which is characterized both for a higher intensity of flight activity and increase in garbage and resin transport, as well as the swarming process in the spring. In this way, an action targeting the preservation or management of the species in a natural environment should consider that survival and reproduction of the colony depends greatly on the amount of available pollen in late winter.