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Souza-Silva F.,Laboratorio Of Biologia Molecular E Doencas Endemicas | Pereira B.A.S.,Laboratorio Of Biologia Molecular E Doencas Endemicas | Finkelstein L.C.,Laboratorio Of Imunoparasitologia | Zucolotto V.,University of Sao Paulo | And 2 more authors.
Journal of Molecular Recognition | Year: 2014

Peptides from the COOH-terminal extension of cysteine proteinase B from Leishmania (Leishmania) amazonensis (cyspep) can modulate immune responses in vertebrate hosts. With this hypothesis as base, we used the online analysis tool SYFPEITHI to predict seven epitopes from this region with potential to bind H2 proteins. We performed proliferation tests and quantified reactive T lymphocytes applying a cytometry analysis, using samples from draining lymph node of lesions from L. (L.) amazonensis-infected mice. To define reactivity of T cells, we used complexes of DimerX (H2 Db:Ig and H2 Ld:Ig) and the putative epitopes. Additionally, we applied surface plasmon resonance to verify real time interactions between the putative epitopes and DimerX proteins. Five peptides induced blastogenesis in BALB/c cells, while only two presented the same property in C57BL/6 mouse cells. In addition, our data indicate the existence of CD8+ T lymphocyte populations able to recognize each tested peptide in both murine strains. We observed an overlapping of results between the peptides that induced lymphocyte proliferation and those capable of binding to the DimerX in the surface plasmon resonance assays thus indicating that using these recombinant proteins in biosensing analyses is a promising tool to study real time molecular interactions in the context of major histocompatibility complex epitopes. The data gathered in this study reinforce the hypothesis that cyspep-derived peptides are important factors in the murine host infection by L. (L.) amazonensis. Copyright © 2014 John Wiley & Sons, Ltd.


Lima-Camara T.N.,Programa de Computacao Cientifica PROCC | Lima-Camara T.N.,Instituto Oswaldo Cruz | Lima-Camara T.N.,University of Sao Paulo | Lima J.B.P.,Instituto Oswaldo Cruz | And 6 more authors.
Parasites and Vectors | Year: 2014

Background: Dengue is an arbovirus disease transmitted by two Aedes mosquitoes: Ae. aegypti and Ae. albopictus. Virgin females of these two species generally show a bimodal and diurnal pattern of activity, with early morning and late afternoon peaks. Although some studies on the flight activity of virgin, inseminated and blood-fed Ae. aegypti females have been carried out under laboratory conditions, little is known about the effects of such physiological states on the locomotor activity of Ae. albopictus and Ae. aegypti females. The aim of this study was to analyze, under laboratory conditions, the effects of insemination and blood-feeding on the locomotor activity of Ae. albopictus and Ae. aegypti females under LD 12:12, at 25°C. Methods. Both Ae. albopictus and Ae. aegypti females were obtained from established laboratory colonies. Control groups were represented by virgin/unfed Ae. albopictus and Ae. aegypti females. Experiments were conducted under laboratory conditions, using an activity monitor that registers individual activity every thirty minutes. Results: Virgin/unfed Ae. albopictus and Ae. aegypti females showed a diurnal and bimodal pattern of locomotor activity, with peaks at early morning and late afternoon. Insemination and blood-feeding significantly decreased the locomotor activity of Ae. aegypti females, but inseminated/blood-fed Ae. aegypti and Ae. albopictus females showed a similar significant decrease on the locomotor activity compared to virgin/unfed females. Conclusions: This study is the first demonstration of the effects of insemination and blood-feeding on the locomotor activity of Ae. albopictus and Ae. aegypti females under artificial conditions. Data suggest that Ae. albopictus and Ae. aegypti females respond in different ways to physiological status changes and such divergence between these two dengue vectors, associated with several ecological differences, could be related to the greater dengue vectorial capacity of Ae. aegypti in Americas in comparison to Ae. albopictus. © 2014 Lima-Camara et al.; licensee BioMed Central Ltd.


PubMed | Nucleo Operacional Sentinela de Mosquitos Vetores, Federal University of Acre, Instituto Oswaldo Cruz Fiocruz, Programa de Computacao Cientifica PROCC and 2 more.
Type: | Journal: Malaria journal | Year: 2015

In the past decade fish farming has become an important economic activity in the Occidental Brazilian Amazon, where the number of new fish farms is rapidly increasing. One of the primary concerns with this phenomenon is the contribution of fishponds to the maintenance and increase of the anopheline mosquito population, and the subsequent increase in human malaria burden. This study reports the results of a 2-year anopheline abundance survey in fishponds and natural water bodies in a malaria-endemic area in northwest Brazil. The objective of this study was to investigate the contribution of natural water bodies (rivers, streams, creeks, ponds, and puddles) and artificial fishponds as breeding sites for Anopheles spp. in Mncio Lima, Acre and to investigate the effect of limnological and environmental variables on Anopheles spp. larval abundance.Natural water bodies and fishponds were sampled at eight different times over 2 years (early, mid and late rainy season, dry season) in the Amazonian town of Mncio Lima, Acre. Anopheline larvae were collected with an entomological dipper, and physical, chemical and ecological characteristics of each water body were measured. Management practices of fishpond owners were ascertained with a systematic questionnaire.Fishponds were four times more infested with anopheline larvae than natural water bodies. Electrical conductivity and the distance to the nearest house were both significant inverse predictors of larval abundance in natural water bodies. The density of larvae in fishponds raised with increasing border vegetation. Fishponds owned by different farmers varied in the extent of anopheline larval infestation but ponds owned by the same individual had similar infestation patterns over time. Commercial fishponds were 1.7-times more infested with anopheline larvae compared to fishponds for family use.These results suggest that fishponds are important breeding sites for anopheline larvae, and that adequate management activities, such as removal of border vegetation could reduce the abundance of mosquito larvae, most importantly Anopheles darlingi.


Xavier-Carvalho C.,Instituto Oswaldo Cruz | Gibson G.,Escola Nacional de Saude Publica ENSP | Brasil P.,Institute Pesquisa Clinica Evandro Chagas IPEC FIOCRUZ | Ferreira R.X.,Hospital Universitario Antonio Pedro | And 8 more authors.
Infection, Genetics and Evolution | Year: 2013

Dengue is an arthropod-borne emerging viral disease with high morbidity and mortality risk in tropical countries like Brazil. Clinical manifestations are vast, ranging from asymptomatic to most severe forms of dengue such as shock. Previous data have shown that host genetics play a role in disease susceptibility and severity. Herein, we have tested the association of single nucleotide polymorphisms (SNPs) at TNF, IL10, MIF, DCSIGN, CLEC5A, NOD2, CCR5 and MRC1 as candidate genes using a matched case-control study design including 88 severe children cases of dengue patients and 335 healthy unrelated subjects that was also separated in IgG+ and IgG- controls. We demonstrated that the TT genotype of CLEC5A SNP (rs1285933 C>T) is associated with dengue severity (OR=2.25; p=0.03) and that GG genotype of -336G>A DCSIGN (CD209) SNP is associated with protection to severe dengue (OR=0.12; p=0.04). Both comparisons were borderline significant when cases were compared with IgG+ controls subgroup. Nevertheless, genotype-phenotype correlation was also assessed using serum levels of TNF from infected patients at the onset of dengue fever, and CT/TT carriers in CLEC5A secreted higher levels of TNF than CC individuals in 5-7days of infection. No significant difference was observed in TNF levels between genotypes GG versus AG/AA at DCSIGN promoter. Next, we performed a meta-analysis retrieving results from the literature for -336G>A DCSIGN and -308G>A TNF SNPs demonstrating that the consensus estimates of these SNPs indicated no association with dengue severity (when compared to Dengue fever) in the overall analysis. But, a subgroup analysis in the -336G>A DCSIGN, the G allele was associated with severe dengue susceptibility in Asians (ORallele=2.77; p=0.0001; ORcarriers=2.99; p=0.0001) and protection in Brazilians (ORallele=0.66; p=0.013). In summary, our results suggest that genetic variations at CLEC5A increase the risk and regulate TNF secretion in dengue severity among Brazilians. Also, combined data of the literature suggest population-specific effect of the -336 DCSIGN SNP more prominent in Asians and in a different direction than Brazilians. © 2013 Elsevier B.V.


PubMed | Federal University of Ouro Preto, São Paulo State University, Programa Pos Graduacao em Epidemiologia em Saude Publica, Federal University of Minas Gerais and Programa de Computacao Cientifica PROCC
Type: Journal Article | Journal: International journal of environmental research and public health | Year: 2016

The prevention and control of dengue are great public health challenges for many countries, particularly since 2015, as other arboviruses have been observed to interact significantly with dengue virus. Different approaches and methodologies have been proposed and discussed by the research community. An important tool widely used is modeling and simulation, which help us to understand epidemic dynamics and create scenarios to support planning and decision making processes. With this aim, we proposed and developed DengueME, a collaborative open source platform to simulate dengue disease and its vectors dynamics. It supports compartmental and individual-based models, implemented over a GIS database, that represent Aedes aegypti population dynamics, human demography, human mobility, urban landscape and dengue transmission mediated by human and mosquito encounters. A user-friendly graphical interface was developed to facilitate model configuration and data input, and a library of models was developed to support teaching-learning activities. DengueME was applied in study cases and evaluated by specialists. Other improvements will be made in future work, to enhance its extensibility and usability.


Carlevaro C.M.,CONICET | Martins-Da-Silva J.H.,Programa de Computacao Cientifica PROCC | Savino W.,Laboratorio Of Pesquisas Sobre O Timo | Caffarena E.R.,Programa de Computacao Cientifica PROCC
Journal of Theoretical and Computational Chemistry | Year: 2013

In the last years, the development of small molecule antagonists of VLA-4 for the treatment of diseases, where cell trafficking and activation are important, has increased considerably. Among them, the MK-0617 ligand has proven to be a highly potent and orally active α4β1 antagonist. However, the binding mode of this ligand in the integrin binding site remains unknown. Herein we report a thermodynamic analysis of the interaction between MK-0617 (and one of its isomers) and the VLA-4 protein using molecular docking and the free energy perturbation calculations, based on a comparative model of the α4β1 receptor. Initial complex coordinates were taken from molecular docking assays and submitted to alchemical transformations. Free energy of binding ΔΔG values, derived from experimental IC50 values, were taken as a parameter for determining the most likely binding mode. In addition, molecular dynamics simulations of these ligands within the α4β1 binding site were carried out to elucidate the binding energy profile and identify the most significant residues. Our results indicate that MK-0617 fits within the binding site in a stretched conformation, pointing the carboxylate group towards the MIDAS ion. We observe that, despite the fact that the main contribution to the energetic binding process is due to the electrostatic ion contribution, the nonpolar contribution is not negligible. Additionally, a network of hydrogen bonds participate in stabilizing the ligand-receptor interaction. © 2013 World Scientific Publishing Company.


Ferreira R.A.X.,Federal University of Fluminense | de Oliveira S.A.,Federal University of Fluminense | Gandini M.,Instituto Oswaldo Cruz | Ferreira L.C.,Federal University of Fluminense | And 5 more authors.
Acta Tropica | Year: 2015

Dengue fever is usually a benign acute viral infection transmitted by arthropods but may evolve to severe clinical manifestations such as coagulation and/or hemodynamic disorders, caused mainly by an increase of vascular permeability. Deregulated circulating immunological factors have been associated with severity. In Brazil severe cases appeared in children only recently and we evaluated the profile of cytokine/chemokine kinetics in 134 hospitalized young patients during the epidemic in Rio de Janeiro in 2008. Inflammatory cytokines TNF and IFNγ were found elevated during the acute phase in children as well as the anti-inflammatory IL10 and chemokines MIF and CXCL10/IP10, all last three persisting longer during the recovery phase. Severe disease fitting the dengue hemorrhagic fever pattern (WHO, 1997) was associated with higher IL10 and CXCL10/IP10 circulating levels (peak levels at seven days with P< 0.01 and P< 0.001 respectively as compared to DF). These factors were higher in patients pulmonary effusion or ascites (P< 0.05 for IL10 and P< 0.01 for CXCL10/IP10). Both factors were also associated with liver changes such as AST increase correlated with CXCL10/IP10 (r=0.4300 with P< 0.0001) and patients presenting painful hepatomegaly showed higher circulating levels of IL10 (P< 0.01, at 7-9 days) and of CXCL10/IP10 (P< 0.05, 4-6 days and P< 0.001, 7-9 days) when compared to patients without apparent liver alterations. Most cases presented a history of prior infection (93%). This is the first study demonstrating cytokine and chemokine association with severity during dengue fever in Brazilian children. IL10 and CXCL10/IP10 play a role in the disease severity associated with induction of vascular leakage and a novel association with changes in liver dysfunction. © 2015 Elsevier B.V.


Santos D.A.,Programa De Computacao Cientifica PROCC | de Souza Costa M.G.,Programa De Computacao Cientifica PROCC | Alves C.R.,Instituto Oswaldo Cruz | Caffarena E.R.,Programa De Computacao Cientifica PROCC
Journal of Molecular Graphics and Modelling | Year: 2016

Cysteine proteinase B (CPB) is a significant virulence factor for Leishmania infections. Upon processing from its zymogen form, it happens a release of the immunomodulatory CPB C-terminal extension (cyspep) into the cytoplasm of the macrophage. Epitopes derived from this fragment were shown to influence the proportion of lymphocytes CD8+ upon infection, favoring the parasite escaping from the host́s immune system. At present, there is no available structural data of cyspep, which impairs a proper understanding of its biological functions. Here, we attempted to build molecular models for this fragment and subsequently evaluate their stabilities in aqueous solution from molecular dynamics simulations analysis. Characterization of our models obtained with distinct techniques (comparative modeling, threading, and ab initio) indicates a prevalence of β−sheets in agreement with consensus secondary structure predictions. Simulation data supported this finding since the formation of new strands, along with a rapid disruption of helical content, were observed. Overall, this study provides a rationalization of epitope mapping data and an improved understanding of cyspep antigenicity. © 2016 Elsevier Inc.


PubMed | Instituto Oswaldo Cruz and Programa De Computacao Cientifica PROCC
Type: | Journal: Journal of molecular graphics & modelling | Year: 2016

Cysteine proteinase B (CPB) is a significant virulence factor for Leishmania infections. Upon processing from its zymogen form, it happens a release of the immunomodulatory CPB C-terminal extension (cyspep) into the cytoplasm of the macrophage. Epitopes derived from this fragment were shown to influence the proportion of lymphocytes CD8+ upon infection, favoring the parasite escaping from the hosts immune system. At present, there is no available structural data of cyspep, which impairs a proper understanding of its biological functions. Here, we attempted to build molecular models for this fragment and subsequently evaluate their stabilities in aqueous solution from molecular dynamics simulations analysis. Characterization of our models obtained with distinct techniques (comparative modeling, threading, and ab initio) indicates a prevalence of -sheets in agreement with consensus secondary structure predictions. Simulation data supported this finding since the formation of new strands, along with a rapid disruption of helical content, were observed. Overall, this study provides a rationalization of epitope mapping data and an improved understanding of cyspep antigenicity.


PubMed | Instituto Oswaldo Cruz and Programa De Computacao Cientifica PROCC
Type: Journal Article | Journal: Proteins | Year: 2016

New strategies to control Leishmania disease demand an extensive knowledge about several aspects of infection including the understanding of its molecular events. In murine models, cysteine proteinase B from Leishmania amazonensis promotes regulation of immune response, and fragments from its C-terminus extension (cyspep) can play a decisive role in the host-parasite interaction. The interaction between cyspep-derived peptides and major histocompatibility complex (MHC) proteins is a crucial factor in Leishmania infections. Seven cyspep-derived peptides, previously identified as capable of interacting with H-2 (murine) MHC class I proteins, were studied in this work. We established a protocol to simulate the unbinding of these peptides from the cleft of H-2 receptors. From the simulations, we estimated the corresponding free energy of dissociation (Gd ) and described the molecular events that occur during the exit of peptides from the cleft. To test the reliability of this method, we first applied it to a calibration set of four crystallographic MHC/peptide complexes. Next, we explored the unbinding of the seven complexes mentioned above. Results were consistent with Gd values obtained from surface plasmon resonance (SPR) experiments. We also identified some of the primary interactions between peptides and H-2 receptors, and we detected three regions of influence for the interaction. This pattern was systematically observed for the peptides and helped determine a minimum distance for the real interaction between peptides and H-2 proteins occurring at 25 .

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