Molecular analysis of the dengue virus type 1 and 2 in Brazil based on sequences of the genomic envelope-nonstructural protein 1 junction region

The genomic sequences of the Envelope-Non-Structural protein 1 junction region (E/NS1) of 84 DEN-1 and 22 DEN-2 isolates from Brazil were determined. Most of these strains were isolated in the period from 1995 to 2001 in endemic and regions of recent dengue transmission in São Paulo State. Sequence data for DEN-1 and DEN-2 utilized in phylogenetic and split decomposition analyses also include sequences deposited in GenBank from different regions of Brazil and of the world. Phylogenetic analyses were done using both maximum likelihood and Bayesian approaches. Results for both DEN-1 and DEN-2 data are ambiguous, and support for most tree bipartitions are generally poor, suggesting that E/NS1 region does not contain enough information for recovering phylogenetic relationships among DEN-1 and DEN-2 sequences used in this study. The network graph generated in the split decomposition analysis of DEN-1 does not show evidence of grouping sequences according to country, region and clades. While the network for DEN-2 also shows ambiguities among DEN-2 sequences, it suggests that Brazilian sequences may belong to distinct subtypes of genotype III.

Production and diagnostic application of recombinant domain III of West Nile envelope protein in Brazil

INTRODUCTION: West Nile virus (WNV) is a flavivirus with a natural cycle involving mosquitoes and birds. Over the last 11 years, WNV has spread throughout the Americas with the imminent risk of its introduction in Brazil. METHODS: Envelope protein domain III of WNV (rDIII) was bacterially expressed and purified. An enzyme-linked immunosorbent assay with WNV rDIII antigen was standardized against mouse immune fluids (MIAFs) of different flavivirus. RESULTS: WNV rDIII reacted strongly with St. Louis encephalitis virus (SLEV) MIAF but not with other flaviviruses. CONCLUSIONS: This antigen may be a potentially useful tool for serologic diagnosis and may contribute in future epidemiological surveillance of WNV infections in Brazil.

Inflammatory response of endothelial cells to hepatitis C virus recombinant envelope glycoprotein 2 protein exposure

The hepatitis C virus (HCV) encodes approximately 10 different structural and non-structural proteins, including the envelope glycoprotein 2 (E2). HCV proteins, especially the envelope proteins, bind to cell receptors and can damage tissues. Endothelial inflammation is the most important determinant of fibrosis progression and, consequently, cirrhosis. The aim of this study was to evaluate and compare the inflammatory response of endothelial cells to two recombinant forms of the HCV E2 protein produced in different expression systems (Escherichia coli and Pichia pastoris). We observed the induction of cell death and the production of nitric oxide, hydrogen peroxide, interleukin-8 and vascular endothelial growth factor A in human umbilical vein endothelial cells (HUVECs) stimulated by the two recombinant E2 proteins. The E2-induced apoptosis of HUVECs was confirmed using the molecular marker PARP. The apoptosis rescue observed when the antioxidant N-acetylcysteine was used suggests that reactive oxygen species are involved in E2-induced apoptosis. We propose that these proteins are involved in the chronic inflammation caused by HCV.

Recombinant hepatitis C virus-envelope protein 2 interactions with low-density lipoprotein/CD81 receptors

Hepatitis C virus (HCV) envelope protein 2 (E2) is involved in viral binding to host cells. The aim of this work was to produce recombinant E2B and E2Y HCV proteins in Escherichia coli and Pichia pastoris, respectively, and to study their interactions with low-density lipoprotein receptor (LDLr) and CD81 in human umbilical vein endothelial cells (HUVEC) and the ECV304 bladder carcinoma cell line. To investigate the effects of human LDL and differences in protein structure (glycosylated or not) on binding efficiency, the recombinant proteins were either associated or not associated with lipoproteins before being assayed. The immunoreactivity of the recombinant proteins was analysed using pooled serum samples that were either positive or negative for hepatitis C. The cells were immunophenotyped by LDLr and CD81 using flow cytometry. Binding and binding inhibition assays were performed in the presence of LDL, foetal bovine serum (FCS) and specific antibodies. The results revealed that binding was reduced in the absence of FCS, but that the addition of human LDL rescued and increased binding capacity. In HUVEC cells, the use of antibodies to block LDLr led to a significant reduction in the binding of E2B and E2Y. CD81 antibodies did not affect E2B and E2Y binding. In ECV304 cells, blocking LDLr and CD81 produced similar effects, but they were not as marked as those that were observed in HUVEC cells. In conclusion, recombinant HCV E2 is dependent on LDL for its ability to bind to LDLr in HUVEC and ECV304 cells. These findings are relevant because E2 acts to anchor HCV to host cells; therefore, high blood levels of LDL could enhance viral infectivity in chronic hepatitis C patients.

Prediction of exposed domains of envelope glycoprotein in Indian HIV-1 isolates and experimental confirmation of their immunogenicity in humans

We describe the impact of subtype differences on the seroreactivity of linear antigenic epitopes in envelope glycoprotein of HIV-1 isolates from different geographical locations. By computer analysis, we predicted potential antigenic sites of envelope glycoprotein (gp120 and gp4l) of this virus. For this purpose, after fetching sequences of proteins of interest from data banks, values of hydrophilicity, flexibility, accessibility, inverted hydrophobicity, and secondary structure were considered. We identified several potential antigenic epitopes in a B subtype strain of envelope glycoprotein of HIV-1 (IIIB). Solid- phase peptide synthesis methods of Merrifield and Fmoc chemistry were used for synthesizing peptides. These synthetic peptides corresponded mainly to the C2, V3 and CD4 binding sites of gp120 and some parts of the ectodomain of gp41. The reactivity of these peptides was tested by ELISA against different HIV-1-positive sera from different locations in India. For two of these predicted epitopes, the corresponding Indian consensus sequences (LAIERYLKQQLLGWG and DIIGDIRQAHCNISEDKWNET) (subtype C) were also synthesized and their reactivity was tested by ELISA. These peptides also distinguished HIV-1-positive sera of Indians with C subtype infections from sera from HIV-negative subjects.

Green microfinance: the case of the Cresol System in Southern Brazil

Climate change which until recently seemed a luxury for the microfinance sector, now appears to be crucial for the future of the sector. Due to their low adaptive capacity, the millions of MF clients worldwide happen to be the most vulnerable to a changing climate. Adapting previous analysis conducted in Nepal and Bangladesh by Agrawala and Maëlis (2010) to the Brazilian context, in this inductive qualitative study we aim to assess potential synergies between MF and CC actions and what strategies can be harnessed to better respond to CC vulnerabilities at client/MF level. To do so, we investigated the case of the second largest rural microcredit programme in Brazil, Sistema Cresol de Cooperativas de Crédito Rural com Interação Solidária. Albeit important overlaps between Cresol's product envelope and CC strategies exist, there is still room to realise synergies to both mitigate a new potential source of risk to Cresol's portfolio and to increase clients' adaptive capacity.

Evaluation of GBV-C / HVG viremia in HIV-infected women

The present study aimed at standardizing a real-time quantitative polymerase chain reaction assay to evaluate the presence of GBV-C/HGV RNA. A "TaqMan" assay using primers and probe derived from the 5¢ NCR region was developed and validated. Two hundred and fifty-three plasma samples from HIV-infected women were tested for GBV-C viremia and antibody against the envelope protein 2. GBV-C RNA was detected in 22.5% of the patients whereas the antibody was identified in 25.3% of the cohort. Detection of viral RNA and of antibodies was mutually exclusive. Viral loads showed a mean of 1,777 arbitrary units / mL, being 1.1 and 13,625 arbitrary units / mL respectively the lowest and highest values measured. We conclude that the real-time quantitative polymerase chain reaction method developed is appropriate for the investigation of GBV-C RNA since it was shown to be highly specific and sensitive, as well as requiring few steps, preventing contamination and providing additional information as to the relative viremia of carriers, a parameter that must be included in studies evaluating the co-factors influencing the clinical outcome of HIV/AIDS.

Production and characterization of monoclonal antibodies to the edta extract of Leptospira interrogans, serovar icterohaemorrhagiae

Monoclonal antibodies (MABs) ivere produced against an etbylenediaminetetraacetate (EDTA) extract of Leptospira interrogans serovar icterohaemorrhagiae being characterized by gel precipitation as IgM and IgG (IgGl and IgG2b). The EDTA extract was detected as several bands by silver staining in SDS-PAGE. In the Western blot the bands around 20 KDa reacted with a monoclonal antibody, 47B4D6, and was oxidized by periodate and was not digested by pronase, suggesting that the determinant is of carbohydrate nature, lmmunocytochemistry, using colloidal gold labeling, showed that an EDTA extract determinant recognized by monoclonal antibody 47B4D6, is localized under the outer envelope of serovar icterohaemorrhagiae. Hoe AIAB raised against the EDTA extract was not able to protect hamsters from lethal challenge with virulent homologous leptospires.

O que os psiquiatras brasileiros esperam das classificações diagnósticas?

OBJETIVO: Realizou-se um levantamento postal anônimo com psiquiatras brasileiros afiliados à Associação Brasileira de Psiquiatria (ABP) sobre uso, percepção em termos de utilidade clínica dos sistemas diagnósticos multiaxiais CID-10 e DSM-IV e, ainda, expectativas sobre as próximas revisões. MÉTODOS: A carta-convite, o questionário padronizado e um envelope selado foram enviados para 1.050 dos 3.062 psiquiatras associados da ABP, selecionados de forma randomizada. Em virtude da taxa relativamente baixa de retorno: 160 respostas (15,2%), não se pode afirmar que a amostra obtida seja representativa. resultados: Os principais resultados mostraram que, entre os respondentes, a CID-10 é o sistema diagnóstico mais utilizado, sobretudo seu eixo clínico. O DSM-IV também é amplamente empregado, principalmente os eixos I e II. A principal característica esperada de um sistema classificatório foi a confiabilidade para comunicação. As classificações são vistas como instrumentos a ser manejados pelos psiquiatras e, em menor proporção, por outros membros de equipes multidisciplinares de saúde mental. Os atuais sistemas foram considerados confiáveis para o uso transcultural no contexto brasileiro. Em relação às expectativas quanto aos futuros sistemas, os psiquiatras preferem classificações com menos de 100 rubricas diagnósticas e dividem-se entre os que acham que estas devam expressar preferencialmente causa/patogênese dos transtornos mentais e aqueles que esperam que tais categorias, sobretudo, sejam construídas de modo a subsidiar o planejamento terapêutico. CONCLUSÕES: Os sistemas diagnósticos desempenham um importante papel na prática psiquiátrica cotidiana dos psiquiatras respondentes. As opiniões dos psiquiatras brasileiros parecem refletir bem algumas controvérsias existentes acerca de questões nosográficas decisivas.

Efeitos cardiovasculares da anestesia local com vasoconstritor durante exodontia em coronariopatas

OBJETIVO: Avaliar a ocorrência de variáveis detectoras de isquemia miocárdica, durante ou após o tratamento odontológico, sob anestesia com vasoconstritor (adrenalina). MÉTODOS: Foram incluídos 54 pacientes coronariopatas submetidos a exodontia sob anestesia local com ou sem vasoconstritor, divididos em dois grupos (sorteio por envelope): grupo I, composto por 27 que receberam anestésico com vasoconstritor; e grupo II, composto por 27 que receberam anestésico sem vasoconstritor. Todos os pacientes foram submetidos a monitoração eletrocardiográfica com Holter por 24 horas, a Doppler-ecocardiografia realizada antes e após intervenção odontológica, e a dosagem dos marcadores bioquímicos antes e 24 horas após a exodontia (creatina cinase fração MB [CK-MB] massa, CK-MB atividade e troponina T). A freqüência cardíaca e a pressão arterial nas fases pré-anestesia, pós-anestesia e pós-exodontia também foram aferidas. A Doppler-ecocardiografia teve como objetivo avaliar a contratilidade segmentar do ventrículo esquerdo e a eventual ocorrência de insuficiência mitral. Em todos os casos foi mantido o protocolo farmacológico habitual prescrito pelo cardiologista. RESULTADOS: Três pacientes do grupo I apresentaram depressão do segmento ST (1,0 mm) durante a aplicação da anestesia, dois outros pacientes do mesmo grupo tiveram elevação da CK-MB massa, e em nenhum caso foi verificada presença de isquemia avaliada pelos demais métodos. Não houve registro, neste estudo, de precordialgia, arritmias e ocorrência ou agravamento de hipocontratilidade segmentar do ventrículo esquerdo ou insuficiência mitral. CONCLUSÃO: A exodontia praticada sob uso de anestesia com adrenalina 1:100.000 não implica riscos isquêmicos adicionais quando realizada com boa técnica anestésica e manutenção do tratamento farmacológico prescrito pelo cardiologista.

Dois teoremas sôbre a função gama

This paper proves the following theorems on the gamma function: Theorem I The integral ∫O∞ t u e-t dt = Γ ( u + 1 ) , where u, real or complex, is such that R (u) > -1, will not change its value if we substitute z = Q (cos φ + i sen φ) for the real variable t, being jconstant and such that - Π/2 < φ < Π/2 , Theorem II The integral ∫-∞∞ w2u + 1 e -w² dw = Γ ( u + 1 ) , where 2u + 1 is supposed to be a non negative even integer, will not change its value if we substitute z = w + fi, f being a real constant, for the real variable w. The proof of both theorems is obtained by means of the well known Cauchy theorem on contour integrals on the complex plane, as suggested by CRAMÉR (1, p. 126) and LEVY (3, p. 178).

Macroecologia de carnívoros do Novo Mundo (Mammalia): envelopes de restrição e análise de padrões filogenéticos

The relationship between body size and geographic range was analyzed for 70 species of terrestrial Carnivora ("fissipeds") of the New World, after the control of phylogenetic patterns in the data using phylogenetic eigenvector regression. The analysis from EcoSim software showed that the variables are related as a triangular envelope. Phylogenetic patterns in data were detected by means of phylogenetic correlograms, and 200 simulations of the phenotypic evolution were also performed over the phylogeny. For body size, the simulations suggested a non-linear relationship for the evolution of this character along the phylogeny. For geographic range size, the correlogram showed no phylogenetic patterns. A phylogenetic eigenvector regression was performed on original data and on data simulated under Ornstein-Uhlenbeck process. Since both characters did not evolve under a simple Brownian motion process, the Type I errors should be around 10%, compatible with other methods to analyze correlated evolution. The significant correlation of the original data (r = 0.38; P < 0.05), as well as the triangular envelope, then indicate ecological and adaptive processes connecting the two variables, such as those proposed in minimum viable population models.

Isolation and antigenic characterization of human immunodeficiency virus (HIV) in Brazil

A retrovirus infecting a Brazilian AIDS patient was isolated and characterized in terms of its reactivity with sera from individuals infected with human immunodeficiency viruses 1 and 2 (HIV-1 and HIV-2). The Western blot analysis revealed that the Brazilian isolate is very similar to the well characterized HIV-1 strain. The serum of the patient from whom the virus was isolated did not react with the 140 kDa envelope glycoprotein specific for HIV-2.