Purification, characterization, crystallization and theoretical molecular modeling of gyroxin fraction from Crotalus durissus terrificus venom (Laurenti, 1768)

The venom of Crotalus durissus terrificus snakes presents various substances, including a serine protease with thrombin-like activity, called gyroxin, that clots plasmatic fibrinogen and promote the fibrin formation. The aim of this study was to purify and structurally characterize the gyroxin enzyme from Crotalus durissus terrificus venom. For isolation and purification, the following methods were employed: gel filtration on Sephadex G75 column and affinity chromatography on benzamidine Sepharose 6B; 12% SDS-PAGE under reducing conditions; N-terminal sequence analysis; cDNA cloning and expression through RT-PCR and crystallization tests. Theoretical molecular modeling was performed using bioinformatics tools based on comparative analysis of other serine proteases deposited in the NCBI (National Center for Biotechnology Information) database. Protein N-terminal sequencing produced a single chain with a molecular mass of similar to 30 kDa while its full-length cDNA had 714 bp which encoded a mature protein containing 238 amino acids. Crystals were obtained from the solutions 2 and 5 of the Crystal Screen Kit (R), two and one respectively, that reveal the protein constitution of the sample. For multiple sequence alignments of gyroxin-like B2.1 with six other serine proteases obtained from snake venoms (SVSPs), the preservation of cysteine residues and their main structural elements (alpha-helices, beta-barrel and loops) was indicated. The localization of the catalytic triad in His57, Asp102 and Ser198 as well as S1 and S2 specific activity sites in Thr193 and Gli215 amino acids was pointed. The area of recognition and cleavage of fibrinogen in SVSPs for modeling gyroxin B2.1 sequence was located at Arg60, Arg72, Gln75, Arg81, Arg82, Lis85, Glu86 and Lis87 residues. Theoretical modeling of gyroxin fraction generated a classical structure consisting of two alpha-helices, two beta-barrel structures, five disulfide bridges and loops in positions 37, 60, 70, 99, 148, 174 and 218. These results provided information about the functional structure of gyroxin allowing its application in the design of new drugs.

A nação pela pedra: coleções de paleontologia no Brasil, 1836-1844

A formação e o estudo de coleções de história natural e de paleontologia participaram da instauração da ordem política do Império do Brasil, delineando também uma ordem científica. A simbiose entre ciência e nação encontrou em Peter W. Lund, iniciador dos estudos de paleontologia em nosso país, um agente ativo e constante. As coleções e escritos desse naturalista deram amparo à visualização do passado e à escrita da história em museus, instituições científicas e culturais brasileiras e europeias. As disputas pelo ordenamento político sob as Regências e a Maioridade foram acompanhadas de perto pelo estudo e a explicação das formas de vida e do globo no passado.

Desempenho de reatores aeróbios de leito fluidizado com circulação interna na remoção de matéria carbonácea tratando diferentes águas residuárias

The organic matter. (OM) removal efficiency in the biochemical oxygen demand (BOD) and chemical oxygen demand (COD) forms for domestic wastewater and semi-intensive fish culture effluents by using three phase aerobic fluidized bed reactors with circulation in concentric tubes was studied. Three different ratios between external and internal areas by different internal diameter configurations (100, 125 and 150mm) to the same external diameter of 250mm were used; sand for filters and granulated activated carbon were used as supporting media. The reactors were tested for three hydraulic retention times: 11.5min to the R100, and 3h for the R125 and R150 reactors. The results demonstrated that this kind of reactors had good performance in the BOD and COD removal for different concentrations of waste waters. BOD mean removal efficiencies obtained were: 47% at the R100 reactor, 57% and 93% of raw and filtered BOD respectively at R125, 48 and 89% of raw and filtered BOD at R150. The COD mean calculated removal efficiencies were: 75% at the R100 reactor, 56 and 86% of raw and filtered COD at R125, and 54 and 86% of raw and filtered COD at R150. In the case of domestic wastewater it is necessary to provide a solids removal system at the reactor outflow in order to increase the removal of suspended OM from the final effluent.

Hematological changes in sheep inoculated with natural and Cobalt60-irradiated Crotalus durissus terrificus venom (Laurenti, 1768)

Natural (NV) and Cobalto60-irradiated (IrV) Crotalus durissus terrificus venom were used to evaluate serum production capacity of sheep and possible hematological and biochemical effects. Freeze-dried venom aliquots were diluted in acidified saline solution (NaCl 150 mM, pH 3.0) and irradiated by a Cobalt 60 source at a dose of 5.54 x 102 Gy/h and a concentration of 2.000 Gy. Twelve sheep were divided into two groups of six animals. One group received irradiated venom (IrV) and the other natural venom (NV). Three antigen doses (venom) were administered at monthly intervals. Blood samples were collected weekly for analysis of serum neutralization potency and capacity, complete blood count (CBC), total plasma protein, fibrinogen, albumin, and globulin. At the end of the experiment, the animals were challenged with a LD50 for sheep and showed no signs of envenoming. The two groups did not present clinical alterations. Results of the total leukocyte count did not present interaction or time factor effect for both groups, but there was a different action between them, with the NV group presenting more cells than the IrV group. The leukocyte increase to 13,000/ml indicates that slight leukocytosis occurred in the week after the first inoculation in the NV group. There was no statistically significant difference between groups in the absolute count of segmented neutrophils, eosinophils, and lymphocytes but there were statistically significant oscillations in values at the different collecting times. The NV group presented an increase in the absolute neutrophil count after the first inoculation that persisted for 5 weeks. In the IrV group, the increase in neutrophils occurred only in the first week returning to normal in the following weeks. The alterations in the neutrophil count are indicative of systemic inflammatory response related to cytokine release; response was more marked in the NV group, showing its greater toxicity.

First record of Amblyomma rotundatum Koch, 1844 (Acari: Ixodidae) parasitizing Paleosuchus palpebrosus Cuvier, 1807 (Reptilia: Crocodylidae), in the western border of Pantanal, Mato Grosso do Sul, Brazil.

This is the first record of Amblyomma rotundatum parasitizing Paleosuchus palpebrosus. This record was made during a herpetofaunal study carried out in high-elevation habitats that are under protection in the mountain range "Serra do Amolar", located in the western border of Brazilian Pantanal, in the state of Mato Grosso do Sul.

Amblyomma triste (Koch, 1844) (Acari : Ixodidae): Morphological description of the ovary and of vitellogenesis

The present study presents the morphology, histology, and the dynamics of vitellogenesis in females of the tick Amblyomma triste. The ovary in this species is of the panoistic type, therefore it lacks nurse cells. It is composed of a layer of epithelial cells that outwardly form the wall of the ovary, but also originate the pedicel, the structure that attaches the oocytes to its external margin, as well the oocytes themselves. In Amblyomma triste, the oocytes develop in four synchronic stages, which differs from the process in other tick species. The classification of the stages of the oocytes was carried out based on the presence of four morphologic characteristics: cytoplasm appearance; site of the germ vesicle; presence, quantity, and constitution of the yolk granules and presence of chorium. (c) 2006 Elsevier B.V. All rights reserved.

Amblyomma triste (Koch, 1844) (Acari : Ixodidae) Ovaries: An ultrastructural analysis

This study presents an ultrastructural analysis of the ovary of the tick, Amblyomma triste. The ovary of this species is of the panoistic type that is, without nursing and follicular cells. It is composed of a layer of epithelial cells forming a wall and of germinative cells that generate the oocytes which remain attached to the external margin of this wall by a multicellular pedicel. The different developmental stages in the oocytes had been described by Oliveira et al. [Oliveira, P.R., Bechara, G.H., Camargo-Mathias, M.I., 2006. Amblyomma triste (Koch, 1844) (Acari: Ixodidae): Morphological description of the ovary and of vitellogenesis. Experimental Parasitology 113, 179-185]. The results of the investigation suggest that besides exogenous production of vitellogenic elements, endogenous production can take place simultaneously, contributing to the development and growth of the oocytes. (c) 2007 Elsevier B.V. All rights reserved.

Ultrastructural detection of proteins, lipids and carbohydrates in oocytes of Amblyomma triste (Koch, 1844) (Acari; Ixodidae) during the vitellogenesis process

The ovary of the tick Ainblyomma triste is classified as panoistic, which is characterized by the presence of oogonia without nurse and follicular cells. The present study has demonstrated that the oocytes in all developmental stages (I-IV) are attached to the ovary through a pedicel, a cellular structure that synthesizes and provides carbohydrate, lipids and proteins supplies for the oocytes during the vitellogenesis process. The lipids are deposited during all oocyte stages; they are freely distributed as observed in stages II, III and IV or they form complexes with other elements. The proteins are also deposited in all stages of the oocytes, however, in lower concentration in the stage IV. There is carbohydrate deposition from oocytes in the stage II as well as in stages III and IV. In addition, the present work has demonstrated that the oocyte yolk of A. triste has a glycolipoprotein nature and the elements are deposited in the following sequence: firstly the lipids and proteins, and finally the carbohydrates. (c) 2007 Elsevier Ltd. All rights reserved.

Vitellogenesis in the tick Amblyomma triste (Koch, 1844) (Acari : Ixodidae) - Role for pedicel cells

This study presents new information on the vitellogenesis of the tick Amblyomma triste. In this species, the ovary consists of a layer of epithelial cells, which form the ovarian wall, oogonia and developing oocytes; and the pedicel, a cellular structure that synthesizes and provides yolk precursors for developing oocytes. The pedicel also attaches oocytes to the external surface of the epithelial wall. In this study, evidence is provided in support of pedicel cells in providing compounds for oocytes during vitellogenesis. (c) 2006 Elsevier B.V. All rights reserved.

Morphological records of oocyte maturation in the parthenogenetic tick Amblyomma rotundatum Koch, 1844 (Acari: Ixodidae)

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

Liver dysfunction in patients bitten by Crotalus durissus terrificus (Laurenti, 1768) snakes in Botucatu (State of São Paulo, Brazil)

Os autores estudaram 32 doentes picados por serpentes venenosas, sendo 16 picados por Bothrops spp. e 16 por Crotalus durissus terrificus. Trinta doentes eram do sexo masculino e dois do feminino com idades variando entre 8 e 63 anos (méda 33±15). A prova da retenção da bromosulfaleína apresentou-se aumentada na maioria dos doentes picados por serpentes Crotalus durissus terrificus. Houve correlação positiva entre a retenção da bromosulfaleína e os níveis séricos de alanina aminotransferase e entre alanina e aspartato aminotransferase apenas nos doentes do grupo Crotalus. Um dos doentes evoluiu para o óbito e apresentou no exame anatomopatológico do fígado degeneração hidrópica e lesões mitocondriais. Os autores concluem que as alterações hepáticas são causadas por pelo menos dois mecanismos a saber: lesão mitocondrial por efeito do veneno crotálico; efeito das citoquinas, especialmente a interleucina-6.

Localization of Antigenic Sites in Unfed Nymphs of Amblyomma triste Koch 1844 (Acari: Ixodidae) Ticks by Immunohistochemistry

P>The reactivity of sera collected from guinea pigs after three infestations with Amblyomma triste nymphs on histological sections of the same tick species was investigated through immunohistochemistry to identify potential target cells and tissues. Six guinea pigs were infested thrice, at 30 day intervals, with 30 nymphs of A. triste per animal per infestation. Blood samples were collected from the guinea pigs 15 days after each infestation for serum separation; normal serum was obtained before the first infestation as control. Unfed A. triste nymphs' histological sections were submitted to indirect immunohistochemistry technique by using normal or hyperimmune guinea pig serum as primary antibody and a goat IgG-alkaline phosphatase-APase conjugate as secondary antibody. A weak to moderate APase activity was observed in cells of salivary glands, midgut and haemolymph of unfed nymphs incubated with hyperimmune serum.

Dinâmica da vitelogêneses de carrapatos Amblyomma triste (Koch, 1844) (Acari: Ixodidae)

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Purificação, caracterização, cristalização e modelagem molecular teórica da fração giroxina do veneno de Crotalus durissus terrificus (Laurenti 1768)

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Assessment of the socioeconomic and environmental impact of hurricane Ivan on Jamaica

Includes bibliography