Atual estágio de desenvolvimento da tecnologia GTL e perspectivas para o Brasil

Although the production of liquid fuels from coal, natural gas (CTL and GTL processes) and other carbon sources has been discovered 90 years ago, the interest was renewed in the last years motivated by stranded gas reserves and by petroleum market instability. This review intends to show the scenario of these technologies nowadays, discussing the applied technologies, the steps in an industrial plant, the ways to produce syngas and liquid fuels, the catalysts used, the type of reactors and the operating plants with their respective capacities, besides the technical, economical and environmental viability, challenges and perspectives for Brazil.

Controle genético da antracnose foliar em milho

Em experimento realizado em casa de vegetação com quatro linhagens e dois híbridos de milho (Zea mays), verificou-se variabilidade para resistência a queima foliar causada por Colletotrichum graminicola avaliada por meio das variáveis área total de lesão por planta (ATL), área média de lesão (AML), comprimento total de lesão por planta (CTL), comprimento médio de lesão (CML) e número de lesões por planta (NL). Quando dois isolados de C. graminicola foram inoculados nessas linhagens e em híbridos de milho não foi verificada interação diferencial entre isolados e genótipos do hospedeiro. Em estudos de herança da resistência, as linhagens genitoras L186 e L64 comportaram-se como suscetível e resistente, respectivamente, ao passo que o híbrido L186 x L64 apresentou resistência. A análise das freqüências observadas de plantas resistentes e suscetíveis na população F2 resultantes da autofecundação do híbrido, indicou ocorrência de herança monogênica com dominância completa para o alelo que confere resistência à doença.

Bloqueio do receptor AT1 da Angiotensina II reduz o número de folículos antrais em ratas com obesidade induzida por dieta de cafeteria

OBJETIVO: Avaliar o desenvolvimento folicular em ratas Wistar com obesidade induzida por dieta de cafeteria (DCAF) submetidas à administração de losartan (LOS), um antagonista do receptor AT1 da Angiotensina II. MÉTODOS: Aos 21 dias de vida, as ratas foram separadas aleatoriamente em dois grupos: controle (CTL), que recebeu ração padrão, e cafeteria (CAF), que recebeu a DCAF, altamente palatável e calórica. Aos 70 dias de vida, início da idade reprodutiva, animais do grupo CAF foram subdivididos em dois grupos (n=15/grupo): CAF, que recebeu água, e CAF+LOS, que recebeu 30 mg/kg de peso corporal (PC) de LOS por gavagem durante 30 dias. O grupo CTL também recebeu água por gavagem. Aos 100 dias de vida foi realizada a eutanásia dos animais e o PC e das gorduras retroperitoneal, perigonadal e subcutânea foi avaliado. Os ovários direitos foram retirados para contagem do número dos diferentes tipos de folículos ovarianos. As concentrações plasmáticas dos hormônios folículo-estimulantes (FSH), luteinizante (LH), prolactina (PRL) e progesterona foram avaliadas. Os resultados foram expressos como média±erro padrão da média. Para análise estatística, foi utilizado one-way ANOVA, seguido pelo pós-teste de Newman-Keuls (p<0,05). RESULTADOS: O PC e das gorduras, assim como o número de folículos antrais, foi elevado no grupo CAF em relação ao CTL. Todavia, as concentrações de FSH e LH foram mais baixas entre os animais CAF. A administração de LOS reduziu o PC e das gorduras retroperitoneal e subcutânea, bem como o número de folículos antrais. O tratamento com LOS atenuou a redução das concentrações de FSH e de LH. As concentrações de progesterona e PRL foram semelhantes entre os grupos estudados. CONCLUSÃO: O uso de LOS pode favorecer o desenvolvimento folicular em fêmeas obesas e pode possibilitar sua utilização como fármaco coadjuvante no tratamento da infertilidade associada à obesidade.

Genetic and antigenic analysis of Babesia bigemina isolates from five geographical regions of Brazil

A molecular epidemiological study was performed with Babesia bigemina isolates from five geographical regions of Brazil. The genetic analysis was done with random amplification of polymorphic DNA (RAPD), repetitive extragenic palindromic elements-polymerase chain reaction (REP-PCR) and enterobacterial repetitive intergenic consensus sequences-polymerase chain reaction (ERIC-PCR) that showed genetic polymorphism between these isolates and generated fingerprinting. In RAPD, ILO872 and ILO876 primers were able to detect at least one fingerprinting for each B. bigemina isolate. The amplification of B. bigemina DNA fragments by REP-PCR and ERIC-PCR gave evidence for the presence in this haemoprotozoan of the sequences described previously in microorganisms of the bacterial kingdom. For the first time it was demonstrated that both techniques can be used for genetic analysis of a protozoan parasite, although the ERIC-PCR was more discriminatory than REP-PCR. The dendogram with similarity coefficient among isolates showed two clusters and one subcluster. The Northeastern and Mid-Western isolates showed the greatest genetic diversity, while the Southeastern and Southern isolates were the closest. The antigenic analysis was done through indirect fluorescent antibody technique and Western blotting using a panel of monoclonal antibodies directed against epitopes on the merozoite membrane surface, rhoptries and membrane of infected erythrocytes. As expected, the merozoite variable surface antigens, major surface antigen (MSA)-1 and MSA-2 showed antigenic diversity. However, B cell epitopes on rhoptries and infected erythrocytes were conserved among all isolates studied. In this study it was possible to identify variable and conserved antigens, which had already been described as potential immunogens. Considering that an attenuated Babesia clone used as immunogen selected populations capable of evading the immunity induced by this vaccine, it is necessary to evaluate more deeply the cross-protection conferred by genetically more distant Brazilian B. bigemina isolates and make an evaluation of the polymorphism degree of variable antigens such as MSA-1 and MSA-2.

Immunocytochemistry of the mucilage cells of Araucaria angustifolia (Bertol.) Kuntze (Araucariaceae)

The use of monoclonal antibodies for specific pectic epitopes is an important tool in the study of the cell wall. Throughout the development of mucilage cells of Araucaria angustifolia (Bertol.) Kuntze, a gradient of distribution was observed in relation to the pectic de-esterification, as well as to the increase of galactan and arabinan epitope distribution, and to the reduction of arabinogalactans proteins (AGPs) epitope at maturity. AGP and methyl-esterified homogalacturonan (HGA) were present in the mucilage. Galactans and arabinans were also observed in the mucilage, though with weak labelling. Degradation of AGP in the maturity of mucilage cells, in cell wall, as well as in the secretion, could be involved in the programmed cell death (PCD). Different labellings found among parenchyma and mucilage cells suggested differences in the cell wall properties of the mucilage cells.

Molecular mimicry between cardiac myosin and Trypanosoma cruzi antigen B13: identification of a B13-driven human T cell clone that recognizes cardiac myosin

Previous reports from our group have demonstrated the association of molecular mimicry between cardiac myosin and the immunodominant Trypanosoma cruzi protein B13 with chronic Chagas' disease cardiomyopathy at both the antibody and heart-infiltrating T cell level. At the peripheral blood level, we observed no difference in primary proliferative responses to T. cruzi B13 protein between chronic Chagas' cardiopathy patients, asymptomatic chagasics and normal individuals. In the present study, we investigated whether T cells sensitized by T. cruzi B13 protein respond to cardiac myosin. T cell clones generated from a B13-stimulated T cell line obtained from peripheral blood of a B13-responsive normal donor were tested for proliferation against B13 protein and human cardiac myosin. The results showed that one clone responded to B13 protein alone and the clone FA46, displaying the highest stimulation index to B13 protein (SI = 25.7), also recognized cardiac myosin. These data show that B13 and cardiac myosin share epitopes at the T cell level and that sensitization of a T cell with B13 protein results in response to cardiac myosin. It can be hypothesized that this also occurs in vivo during T. cruzi infection which results in heart tissue damage in chronic Chagas' disease cardiomyopathy

Malaria vaccine: roadblocks and possible solutions

Malaria remains the most prevalent and devastating parasitic disease worldwide. Vaccination is considered to be an approach that will complement other strategies for prevention and control of the disease in the future. In the last 10 years, intense studies aimed at the development of a malaria vaccine have provided important knowledge of the nature of the host immunological mechanisms of protection and their respective target antigens. It became well established that protective immune responses can be generated against the distinct stages of Plasmodium. However, in general, protective immune responses are directed at stage-specific antigens. The elucidation of the primary structure of these antigens made possible the generation of synthetic and recombinant proteins that are being extensively used in experimental immunizations against the infection. Today, several epitopes of limited polymorphism have been described and protective immunity can be generated by immunization with them. These epitopes are being tested as primary candidates for a subunit vaccine against malaria. Here we critically review the major roadblocks for the development of a malaria vaccine and provide some insight on how these problems are being solved

Trypanosoma cruzi: amastigote polymorphism defined by monoclonal antibodies

We have raised monoclonal antibodies (mAbs) directed towards amastigote forms of Trypanosoma cruzi, and shown that mAbs 1D9 and 4B9 are carbohydrate while mAb 4B5 activity is resistant to periodate oxidation of the antigen. Here we used an ELISA to quantitate and compare the expression of surface epitopes on fixed parasites among different parasite isolates. The expression of markers varied among T. cruzi amastigotes isolated from infected cells or after extracellular differentiation of trypomastigotes. Moreover, we also observed an extensive polymorphic expression of these epitopes among amastigotes derived from different strains and clones. For instance, mAb 2C2 strongly and evenly reacted with 9 strains and clones (G, Y, CL, Tulahuen, MD, and F, and clones Sylvio X-10/4, D11, and CL.B), with absorbance at 492 nm (A492 nm) from 0.6 to 0.8. By contrast, mAb 4B5 had a higher expression in Tulahuen amastigotes (around 0.9 at 492 nm) whereas its reactivity with amastigotes from clones CL.B, Sylvio X-10/4 and D11 was much lower (around 0.4). mAb 1D9 displayed an interesting pattern of reactivity with amastigotes of the different strains and clones (A492 nm of G>D11³Sylvio X-10/4 = MD>Tulahuen = F = Y>CL>CL.B). Finally, we observed that mAb 4B9 had the lowest reaction with the parasites studied, with higher values of A492 nm with Y strain (around 0.6) and lower values with Tulahuen, F and CL.B strains (around 0.2). Immunoblotting analysis also showed extensive variations among amastigotes of the various parasite isolates and mAbs 4B9, 1D9 and 4B5 revealed significant differences in expression between clones and parental strains. These data describe a previously uncharacterized polymorphism of T. cruzi amastigote surface components.

MHC-restricted antigen presentation and recognition: constraints on gene, recombinant and peptide vaccines in humans

The target of any immunization is to activate and expand lymphocyte clones with the desired recognition specificity and the necessary effector functions. In gene, recombinant and peptide vaccines, the immunogen is a single protein or a small assembly of epitopes from antigenic proteins. Since most immune responses against protein and peptide antigens are T-cell dependent, the molecular target of such vaccines is to generate at least 50-100 complexes between MHC molecule and the antigenic peptide per antigen-presenting cell, sensitizing a T cell population of appropriate clonal size and effector characteristics. Thus, the immunobiology of antigen recognition by T cells must be taken into account when designing new generation peptide- or gene-based vaccines. Since T cell recognition is MHC-restricted, and given the wide polymorphism of the different MHC molecules, distinct epitopes may be recognized by different individuals in the population. Therefore, the issue of whether immunization will be effective in inducing a protective immune response, covering the entire target population, becomes an important question. Many pathogens have evolved molecular mechanisms to escape recognition by the immune system by variation of antigenic protein sequences. In this short review, we will discuss the several concepts related to selection of amino acid sequences to be included in DNA and peptide vaccines.

DNA vaccines for viral diseases

DNA plasmids encoding foreign proteins may be used as immunogens by direct intramuscular injection alone, or with various adjuvants and excipients, or by delivery of DNA-coated gold particles to the epidermis through biolistic immunization. Antibody, helper T lymphocyte, and cytotoxic T lymphocyte (CTL) responses have been induced in laboratory and domesticated animals by these methods. In a number of animal models, immune responses induced by DNA vaccination have been shown to be protective against challenge with various infectious agents. Immunization by injection of plasmids encoding foreign proteins has been used successfully as a research tool. This review summarizes the types of DNA vaccine vectors in common use, the immune responses and protective responses that have been obtained in animal models, the safety considerations pertinent to the evaluation of DNA vaccines in humans and the very limited information that is available from early clinical studies.

Reactivity of anti-thyroid antibodies to thyroglobulin tryptic fragments: comparison of autoimmune and non-autoimmune thyroid diseases

Studies concerning the antigenicity of thyroglobulin fragments allow the characterization of the epitopes but do not consider the role of heavier antigenic fragments that could result in vivo from the action of endoproteases. Here we assess the relative importance of the fragments obtained from thyroglobulin by limited proteolysis with trypsin and compare by immunoblotting their reactivity to serum from patients with autoimmune (Graves' disease and Hashimoto's thyroiditis) and non-autoimmune (subacute thyroiditis) disease. The results showed no difference in frequency of recognition of any peptide by sera from patients with autoimmune thyroiditis. In contrast, sera from patients with subacute thyroiditis reacted more frequently with a peptide of 80 kDa. These results suggest the presence of antibody subpopulations directed at fragments produced in vivo by enzymatic cleavage of thyroglobulin. This fragment and antibodies to it may represent markers for subacute thyroiditis.

Two related thrombin-like enzymes present in Bothrops atrox venom

This article describes the presence of two new forms of a thrombin-like enzyme, both with apparent molecular masses of 38 kDa, in Bothrops atrox venom. Both share the ability to cleave fibrinogen into fibrin and to digest casein. Both present identical Km on the substrate BApNA. Their N-terminal amino acid sequences are identical for 26 residues, sharing 80% homology with batroxobin and flavoxobin. Two groups of monoclonal antibodies (mAbs) raised against the purified enzyme forms recognized different epitopes of the putative corresponding enzymes present in B. atrox crude venom. On Western blotting analysis of B. atrox crude venom, mAbs 5DB2C8, 5AA10 and 5CF11, but not mAbs 6CC5 and 6AD2-G5, revealed two or more protein bands ranging from 25 to 38 kDa. By immunoprecipitation assays, the 6AD2-G5 mAb was able to precipitate protein bands of 36-38 kDa from B. atrox, B. leucurus, B. pradoi, B. moojeni, B. jararaca and B. neuwiedii crude venoms. Fibrinogen-clotting activity was inhibited when the same venom specimens were pre-incubated with mAb 6AD2-G5, except for B. jararaca and B. neuwiedii.

The pathogenesis of tropical spastic paraparesis/human T-cell leukemia type I-associated myelopathy

Tropical spastic paraparesis/human T-cell leukemia type I-associated myelopathy (TSP/HAM) is caused by a human T-cell leukemia virus type I (HTLV-I) after a long incubation period. TSP/HAM is characterized by a chronic progressive paraparesis with sphincter disturbances, no/mild sensory loss, the absence of spinal cord compression and seropositivity for HTLV-I antibodies. The pathogenesis of this entity is not completely known and involves a multivariable phenomenon of immune system activation against the presence of HTLV-I antigens, leading to an inflammatory process and demyelination, mainly in the thoracic spinal cord. The current hypothesis about the pathogenesis of TSP/HAM is: 1) presence of HTLV-I antigens in the lumbar spinal cord, noted by an increased DNA HTLV-I load; 2) CTL either with their lytic functions or release/production of soluble factors, such as CC-chemokines, cytokines, and adhesion molecules; 3) the presence of Tax gene expression that activates T-cell proliferation or induces an inflammatory process in the spinal cord; 4) the presence of B cells with neutralizing antibody production, or complement activation by an immune complex phenomenon, and 5) lower IL-2 and IFN-gamma production and increased IL-10, indicating drive to a cytokine type 2 pattern in the TSP/HAM subjects and the existence of a genetic background such as some HLA haplotypes. All of these factors should be implicated in TSP/HAM and further studies are necessary to investigate their role in the development of TSP/HAM.

Production and characterization of monoclonal antibodies to Brazilian isolates of bovine viral diarrhea virus

Three Brazilian isolates of bovine viral diarrhea virus (BVDV), antigenically distinct from the standard North American isolates, were selected to immunize BALB/c mice in order to obtain hybridoma cells secreting anti-BVDV monoclonal antibodies (mAbs). Two hybridoma clones secreting mAbs, reacting specifically with BVDV-infected cells (mAbs 3.1C4 and 6.F11), were selected after five fusions and screening of 1001 hypoxanthine-aminopterin-thymidine-resistant clones. These mAbs reacted in an indirect fluorescent antibody (IFA) assay with all 39 South and North American BVDV field isolates and reference strains available in our laboratory, yet failed to recognize other pestiviruses, namely the hog cholera virus. The mAbs reacted at dilutions up to 1:25,600 (ascitic fluid) and 1:100 (hybridoma culture supernatant) in IFA and immunoperoxidase (IPX) staining of BVDV-infected cells but only mAb 3.1C4 neutralized virus infectivity. Furthermore, both mAbs failed to recognize BVDV proteins by IPX in formalin-fixed paraffin-embedded tissues and following SDS-PAGE and immunoblot analysis of virus-infected cells, suggesting they are probably directed to conformational-type epitopes. The protein specificity of these mAbs was then determined by IFA staining of CV-1 cells transiently expressing each of the BVDV proteins: mAb 3.1C4 reacted with the structural protein E2/gp53 and mAb 6.F11 reacted with the structural protein E1/gp25. Both mAbs were shown to be of the IgG2a isotype. To our knowledge, these are the first mAbs produced against South American BVDV isolates and will certainly be useful for research and diagnostic purposes.

N-terminal amino acids of bovine alpha interferons are relevant for the neutralization of their antiviral activity

The structure-function relationship of interferons (IFNs) has been studied by epitope mapping. Epitopes of bovine IFNs, however, are practically unknown, despite their importance in virus infections and in the maternal recognition of pregnancy. It has been shown that recombinant bovine (rBo)IFN-alphaC and rBoIFN-alpha1 differ only in 12 amino acids and that the F12 monoclonal antibody (mAb) binds to a linear sequence of residues 10 to 34. We show here that the antiviral activities of these two IFNs were neutralized by the F12 mAb to different extents using two tests. In residual activity tests the antiviral activity dropped by more than 99% with rBoIFN-alphaC and by 84% with rBoIFN-alpha1. In checkerboard antibody titrations, the F12 mAb titer was 12,000 with rBoIFN-alphaC and only 600 with rBoIFN-alpha1. Since these IFNs differ in their amino acid sequence at positions 11, 16 and 19 of the amino terminus, only these amino acids could account for the different neutralization titers, and they should participate in antibody binding. According to the three-dimensional structure described for human and murine IFNs, these amino acids are located in the alpha helix A; amino acids 16 and 19 of the bovine IFNs would be expected to be exposed and could bind to the antibody directly. The amino acid at position 11 forms a hydrogen bond in human IFNs-alpha and it is possible that, in bovine IFNs-alpha, the F12 mAb, binding near position 11, would disturb this hydrogen bond, resulting in the difference in the extent of neutralization observed.