Search

BackgroundTraditional medicinal plants and other remedies are one of the many potential sources of new antimalarial drugs, there being an increasing interest in studying their potential for the treatment of malaria and other illnesses. The Cabo Verde archipelago harbors a rich native plant diversity, most of which are used with medicinal purposes. MethodsThe present study investigated the in vitro antiplasmodial activities of four native plants from Cabo Verde. Aqueous and ethanolic extracts of these plants were tested in vitro against both chloroquine-sensitive (3D7) and chloroquine-resistant (Dd2) P. falciparum strains using a SYBR Green detection method, and their cytotoxicity was evaluated in Caco and PLP2 cells by using a sulforhodamine B colorimetric assay. ResultsAn ethanolic extract of Artemisia gorgonum and an aqueous extract of Tamarix senegalensis exhibited high antiplasmodial activity (EC50 < 5 μg/mL) without cytotoxicity. Extracts of Lavandula rotundifolia and Sideroxylon marginatum exhibited moderate activity, with EC50 values ranging from 10-30 μg/mL. The A. gorgonum ethanolic extract showed activity toward early ring stages, and parasites treated with the T. senegalensis aqueous extract progressed to early trophozoite stage but did not develop further to the late trophozoite or schizont stages.ConclusionsAntimalarial activities as well as the lack of toxicity of the extracts found in the present study support the claim by traditional practitioners for the use of the plants against malaria and suggests their ethnopharmacological usefulness as future antimalarials.

Increasing reports of multidrug-resistant malaria parasites urge the discovery of new effective drugs with different chemical scaffolds. Protein kinases play a key role in many cellular processes such as signal transduction and cell division, making them interesting targets in many diseases. Protein kinase 7 (PK7) is an orphan kinase from the Plasmodium genus, essential for the sporogonic cycle of these parasites. Here, we applied a robust and integrative artificial intelligence-assisted virtual-screening (VS) approach using shape-based and machine learning models to identify new potential PK7 inhibitors with in vitro antiplasmodial activity. Eight virtual hits were experimentally evaluated, and compound LabMol-167 inhibited ookinete conversion of Plasmodium berghei and blood stages of Plasmodium falciparum at nanomolar concentrations with low cytotoxicity in mammalian cells. As PK7 does not have an essential role in the Plasmodium blood stage and our virtual screening strategy aimed for both PK7 and blood-stage inhibition, we conducted an in silico target fishing approach and propose that this compound might also inhibit P. falciparum PK5, acting as a possible dual-target inhibitor. Finally, docking studies of LabMol-167 with P. falciparum PK7 and PK5 proteins highlighted key interactions for further hit-to lead optimization.

Antimalarial drugs with novel modes of action and wide therapeutic potential are needed to pave the way for malaria eradication. Violacein is a natural compound known for its biological activity against cancer cells and several pathogens, including the malaria parasite, Plasmodium falciparum (Pf). Herein, using chemical genomic profiling (CGP), we found that violacein affects protein homeostasis. Mechanistically, violacein binds Pf chaperones, PfHsp90 and PfHsp70-1, compromising the latter's ATPase and chaperone activities. Additionally, violacein-treated parasites exhibited increased protein unfolding and proteasomal degradation. The uncoupling of the parasite stress response reflects the multistage growth inhibitory effect promoted by violacein. Despite evidence of proteotoxic stress, violacein did not inhibit global protein synthesis via UPR activation - a process that is highly dependent on chaperones, in agreement with the notion of a violacein-induced proteostasis collapse. Our data highlight the importance of a functioning chaperone-proteasome system for parasite development and differentiation. Thus, a violacein-like small molecule might provide a good scaffold for development of a novel probe for examining the molecular chaperone network and/or antiplasmodial drug design. publishersversion published

Funding Information: We acknowledge the participants in the study, without whom this research could not have been done. For laboratorial support the authors wish to show their appreciation to Gabriel Barbosa de Abreu (in memorian). To Norman Ratcliffe for the English revision of this manuscript. Part of this work is described in a Master?s Dissertation by PS, conducted at the Applied Microbiology and Parasitology Post-graduation Program, Federal Fluminense University. Publisher Copyright: © Copyright © 2021 Salles, Perce-da-Silva, Rossi, Raposo, Ramirez Ramirez, Pereira Bastos, Pratt-Riccio, Cassiano, Baptista, Cardoso, Banic and Machado. Genetic variability was linked with individual responses to treatment and susceptibility to malaria by Plasmodium vivax. Polymorphisms in the CYP2D6 gene may modulate enzyme level and activity, thereby affecting individual responses to pharmacological treatment. The aim of the study was to investigate whether or not CYP2D6 single nucleotide polymorphisms rs1065852, rs38920-97, rs16947 and rs28371725 are unequally distributed in malaria by Plasmodium vivax individuals from the Brazilian Amazon region. The blood samples were collected from 220 unrelated Plasmodium vivax patients from five different endemic areas. Genotyping was performed using SNaPshot® and real-time polymerase chain reaction methods. In all five areas, the rs1065852 (CYP2D6*10, C.100C > T), rs3892097 (CYP2D6*4, 1846C > T) and rs16947 (CYP2D6*2, C.2850G > A), as a homozygous genotype, showed the lowest frequencies. The rs28371725 (CYP2D6*41, 2988G > A) homozygous genotype was not detected, while the allele A was found in a single patient from Macapá region. No deviations from Hardy-Weinberg equilibrium were found, although a borderline p-value was observed (p = 0.048) for the SNP rs3892097 in Goianésia do Pará, Pará state. No significant associations were detected in these frequencies among the five studied areas. For the SNP rs3892097, a higher frequency was observed for the C/T heterozygous genotype in the Plácido de Castro and Macapá, Acre and Amapá states, respectively. The distribution of the CYP2D6 alleles investigated in the different areas of the Brazilian Amazon is not homogeneous. Further investigations are necessary in order to determine which alleles might be informative to assure optimal drug dosing recommendations based on experimental pharmacogenetics. publishersversion published

Aim: Computer-aided drug design approaches were applied to identify chalcones with antiplasmodial activity. Methodology: The virtual screening was performed as follows: structural standardization of in-house database of chalcones; identification of potential Plasmodium falciparum protein targets for the chalcones; homology modeling of the predicted P. falciparum targets; molecular docking studies; and in vitro experimental validation. Results: Using these models, we prioritized 16 chalcones with potential antiplasmodial activity, for further experimental evaluation. Among them, LabMol-86 and LabMol-87 showed potent in vitro antiplasmodial activity against P. falciparum, while LabMol-63 and LabMol-73 were potent inhibitors of Plasmodium berghei progression into mosquito stages. Conclusion: Our results encourage the exploration of chalcones in hit-to-lead optimization studies for tackling malaria.

Plasmodium falciparum dihydroorotate dehydrogenase (PfDHODH) has been clinically validated as a target for antimalarial drug discovery, as a triazolopyrimidine class inhibitor (DSM265) is currently undergoing clinical development. Here, we have identified new hydroxyazole scaffold-based PfDHODH inhibitors belonging to two different chemical series. The first series was designed by a scaffold hopping strategy that exploits the use of hydroxylated azoles. Within this series, the hydroxythiadiazole 3 was identified as the best selective PfDHODH inhibitor (IC50 12.0 μM). The second series was designed by modulating four different positions of the hydroxypyrazole scaffold. In particular, hydroxypyrazoles 7e and 7f were shown to be active in the low μM range (IC50 2.8 and 5.3 μM, respectively). All three compounds, 3, 7e and 7f showed clear selectivity over human DHODH (IC50 > 200 μM), low cytotoxicity, and retained micromolar activity in P. falciparum-infected erythrocytes. The crystallographic structures of PfDHODH in complex with compounds 3 and 7e proved their binding mode, supplying essential data for future optimization of these scaffolds.

Malaria is one of the most impacting public health problems in tropical and subtropical areas of the globe, with approximately 200 million cases worldwide annually. In the absence of an effective vaccine, rapid treatment is vital for effective malaria control. However, parasite resistance to currently available drugs underscores the urgent need for identifying new antimalarial therapies with new mechanisms of action. Among potential drug targets for developing new antimalarial candidates, protein kinases are attractive. These enzymes catalyze the phosphorylation of several proteins, thereby regulating a variety of cellular processes and playing crucial roles in the development of all stages of the malaria parasite life cycle. Moreover, the large phylogenetic distance between Plasmodium species and its human host is reflected in marked differences in structure and function of malaria protein kinases between the homologs of both species, indicating that selectivity can be attained. In this review, we describe the functions of the different types of Plasmodium kinases and highlight the main recent advances in the discovery of kinase inhibitors as potential new antimalarial drug candidates.

The discovery and development of a new drug is a complex, time consuming and costly process that typically takes over 10 years and costs around 1 billion dollars from bench to market. This scenario makes the discovery of novel drugs targeting neglected tropical diseases (NTDs), which afflict in particular people in low-income countries, prohibitive. Despite the intensive use of High-Throughput Screening (HTS) in the past decades, the speed with which new drugs come to the market has remained constant, generating doubts about the efficacy of this approach. Here we review a few of the yeast-based high-throughput approaches that can work synergistically with parasite-based, in vitro, or in silico methods to identify and optimize novel antiparasitic compounds. These yeast-based methods range from HTP screens to identify novel hits against promising parasite kinase targets to the identification of potential antiparasitic kinase inhibitors extracted from databases of yeast chemical genetic screens.

Widespread resistance against antimalarial drugs thwarts current efforts for controlling the disease and urges the discovery of new effective treatments. Drug repositioning is increasingly becoming an attractive strategy since it can reduce costs, risks, and time-to-market. Herein, we have used this strategy to identify novel antimalarial hits. We used a comparative in silico chemogenomics approach to select Plasmodium falciparum and Plasmodium vivax proteins as potential drug targets and analyzed them using a computer-assisted drug repositioning pipeline to identify approved drugs with potential antimalarial activity., Widespread resistance against antimalarial drugs thwarts current efforts for controlling the disease and urges the discovery of new effective treatments. Drug repositioning is increasingly becoming an attractive strategy since it can reduce costs, risks, and time-to-market. Herein, we have used this strategy to identify novel antimalarial hits. We used a comparative in silico chemogenomics approach to select Plasmodium falciparum and Plasmodium vivax proteins as potential drug targets and analyzed them using a computer-assisted drug repositioning pipeline to identify approved drugs with potential antimalarial activity. Among the seven drugs identified as promising antimalarial candidates, the anthracycline epirubicin was selected for further experimental validation. Epirubicin was shown to be potent in vitro against sensitive and multidrug-resistant P. falciparum strains and P. vivax field isolates in the nanomolar range, as well as being effective against an in vivo murine model of Plasmodium yoelii. Transmission-blocking activity was observed for epirubicin in vitro and in vivo. Finally, using yeast-based haploinsufficiency chemical genomic profiling, we aimed to get insights into the mechanism of action of epirubicin. Beyond the target predicted in silico (a DNA gyrase in the apicoplast), functional assays suggested a GlcNac-1-P-transferase (GPT) enzyme as a potential target. Docking calculations predicted the binding mode of epirubicin with DNA gyrase and GPT proteins. Epirubicin is originally an antitumoral agent and presents associated toxicity. However, its antiplasmodial activity against not only P. falciparum but also P. vivax in different stages of the parasite life cycle supports the use of this drug as a scaffold for hit-to-lead optimization in malaria drug discovery.

Widespread resistance against antimalarial drugs thwarts current efforts for controlling the disease and urges the discovery of new effective treatments. Drug repositioning is increasingly becoming an attractive strategy since it can reduce costs, risks, and time-to-market. Herein, we have used this strategy to identify novel antimalarial hits. We used a comparative in silico chemogenomics approach to select Plasmodium falciparum and Plasmodium vivax proteins as potential drug targets and analyzed them using a computer-assisted drug repositioning pipeline to identify approved drugs with potential antimalarial activity. Among the seven drugs identified as promising antimalarial candidates, the anthracycline epirubicin was selected for further experimental validation. Epirubicin was shown to be potent in vitro against sensitive and multidrug-resistant P. falciparum strains and P. vivax field isolates in the nanomolar range, as well as being effective against an in vivo murine model of Plasmodium yoelii Transmission-blocking activity was observed for epirubicin in vitro and in vivo Finally, using yeast-based haploinsufficiency chemical genomic profiling, we aimed to get insights into the mechanism of action of epirubicin. Beyond the target predicted in silico (a DNA gyrase in the apicoplast), functional assays suggested a GlcNac-1-P-transferase (GPT) enzyme as a potential target. Docking calculations predicted the binding mode of epirubicin with DNA gyrase and GPT proteins. Epirubicin is originally an antitumoral agent and presents associated toxicity. However, its antiplasmodial activity against not only P. falciparum but also P. vivax in different stages of the parasite life cycle supports the use of this drug as a scaffold for hit-to-lead optimization in malaria drug discovery.

Conselho Nacional para o Desenvolvimento Cient?fico e Tecnol?gico (CNPq) ; Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior, Brazil (CAPES ? DS and PDSE) ? Finance Code 001 S?o Paulo State University. Graduate Program in Biosciences. S?o Jos? do Rio Preto, SP, Brazil. Lisboa University. Tropical Medicine and Hygiene Institut. Global Health and Tropical Medicine. Lisboa, Portugal. Federal University of Sergipe. Department of Biology. Aracaj?, SE, Brazil. Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Laborat?rio de Imunogen?tica da Mal?ria Ananindeua, PA, Brasil. Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Laborat?rio de Microscopia Eletr?nica. Ananindeua, PA, Brasil / Universidade Federal do Par?. Bel?m, PA, Brasil. Fluminense Federal University. Center of Microorganisms Investigation. Niter?i, RJ, Brazil. Fluminense Federal University. Center of Microorganisms Investigation. Niter?i, RJ, Brazil. Fluminense Federal University. Center of Microorganisms Investigation. Niter?i, RJ, Brazil. Funda??o Oswaldo Cruz. Oswaldo Cruz Institute. Laboratory of Malaria Research. Rio de Janeiro, RJ, Brazil. S?o Paulo State University. Graduate Program in Biosciences. S?o Jos? do Rio Preto, SP, Brazil. Funda??o Oswaldo Cruz. Oswaldo Cruz Institute. Laboratory of Immunoparasitology. Rio de Janeiro, RJ, Brazil. S?o Paulo State University. Graduate Program in Biosciences. S?o Jos? do Rio Preto, SP, Brazil / Fluminense Federal University. Center of Microorganisms Investigation. Niter?i, RJ, Brazil. BACKGROUND: It is well established that infection by Plasmodium vivax is a result of host-parasite interactions. In the present study, association with the IL1/IL2 cytokine profiles, anticircumsporozoite protein antibody levels and parasitic loads was evaluated in individuals naturally infected with P. vivax in an endemic area of the Brazilian Amazon. METHODS: Molecular diagnosis of P. vivax and variants was performed using the PCR-RFLP method and IL1B -511C>T, IL2 -330T>G and IL2+114T>G polymorphisms were identified using PCR-RFLP and allele-specific PCR. IL-1? and IL-2 cytokine levels were detected by flow cytometry and circumsporozoite protein (CSP) antibodies were measured by ELISA. RESULTS: Three variants of P. vivax CSP were identified and VK247 was found to be the most frequent. However, the prevalence and magnitude of IgG antibodies were higher for the VK210 variant. Furthermore, the antibody response to the CSP variants was not associated with the presence of the variant in the infection. Significant differences were observed between the single nucleotide polymorphism (SNP) -511T>C in the IL1B gene and levels of antibodies to the VK247 and P. vivax-like variants, but there were no associations between SNPs in IL1 and IL2 genes and their plasma products. CONCLUSIONS: Individuals with the rs16944 CC genotype in the IL1? gene have higher antibody levels to the CSP of P. vivax of VK247 and P. vivax-like variants.

Malaria is a tropical infectious disease that affects over 219 million people worldwide. Due to the constant emergence of parasitic resistance to the current antimalarial drugs, the discovery of new antimalarial drugs is a global health priority. Multi-target drug discovery is a promising and innovative strategy for drug discovery and it is currently regarded as one of the best strategies to face drug resistance. Aiming to identify new multi-target antimalarial drug candidates, we developed an integrative computational approach to select multi-kinase inhibitors for Plasmodium falciparum calcium-dependent protein kinases 1 and 4 (CDPK1 and CDPK4) and protein kinase 6 (PK6). For this purpose, we developed and validated shape-based and machine learning models to prioritize compounds for experimental evaluation. Then, we applied the best models for virtual screening of a large commercial database of drug-like molecules. Ten computational hits were experimentally evaluated against asexual blood stages of both sensitive and multi-drug resistant P. falciparum strains. Among them, LabMol-171, LabMol-172, and LabMol-181 showed potent antiplasmodial activity at nanomolar concentrations (EC50 ≤ 700 nM) and selectivity indices >15 folds. In addition, LabMol-171 and LabMol-181 showed good in vitro inhibition of P. berghei ookinete formation and therefore represent promising transmission-blocking scaffolds. Finally, docking studies with protein kinases CDPK1, CDPK4, and PK6 showed structural insights for further hit-to-lead optimization studies.

BACKGROUND Despite treatment with effective antimalarial drugs, the mortality rate is still high in severe cases of the disease, highlighting the need to find adjunct therapies that can inhibit the adhesion of Plasmodium falciparum-infected erythrocytes (Pf-iEs). OBJECTIVES In this context, we evaluated a new heparan sulfate (HS) from Nodipecten nodosus for antimalarial activity and inhibition of P. falciparum cytoadhesion and rosetting. METHODS Parasite inhibition was measured by SYBR green using a cytometer. HS was assessed in rosetting and cytoadhesion assays under static and flow conditions using Chinese hamster ovary (CHO) and human lymphatic endothelial cell (HLEC) cells expressing intercellular adhesion molecule-1 (ICAM1) and chondroitin sulfate A (CSA), respectively. FINDINGS This HS inhibited merozoite invasion similar to heparin. Moreover, mollusk HS decreased cytoadherence of P. falciparum to CSA and ICAM-1 on the surface of endothelial cells under static and flow conditions. In addition, this glycan efficiently disrupted rosettes. CONCLUSIONS These findings support a potential use for mollusk HS as adjunct therapy for severe malaria.

This work was supported by Funda??o de Amparo ? Pesquisa do Estado de S?o Paulo (FAPESP) Grant 2012/16525-2, and fellowships grants 2015/02808-0 (to NB), 2013/25807-4 (to JAL), 2016/12855-9 (to JLS-F), 2015/20774-6 (to GCC), 2013/20509-5 (to CB). Evandro Chagas core grant (to RLDM). MUF, MVGL and FTMC are Conselho Nacional do Desenvolvimento Cient?fico e Tecnol?gico (CNPq) research fellows. University of Campinas-UNICAMP. Department of Genetics, Evolution, Microbiology and Immunology. Laboratory of Tropical Diseases Prof. Dr. Luiz Jacintho da Silva. Campinas, SP, Brazil. University of Campinas-UNICAMP. Department of Genetics, Evolution, Microbiology and Immunology. Laboratory of Tropical Diseases Prof. Dr. Luiz Jacintho da Silva. Campinas, SP, Brazil. Funda??o Oswaldo Cruz. Instituto Carlos Chagas. Curitiba, PR, Brazil. Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Laborat?rio de Ensaios Cl?nicos e Imunogen?tica em Mal?ria. Ananindeua, PA, Brasil. University of Campinas-UNICAMP. Department of Genetics, Evolution, Microbiology and Immunology. Laboratory of Tropical Diseases Prof. Dr. Luiz Jacintho da Silva. Campinas, SP, Brazil. University of Campinas-UNICAMP. Department of Genetics, Evolution, Microbiology and Immunology. Laboratory of Tropical Diseases Prof. Dr. Luiz Jacintho da Silva. Campinas, SP, Brazil. Funda??o Oswaldo Cruz. Instituto Le?nidas & Maria Deane. Manaus, AM, Brazil / Funda??o de Medicina Tropical Dr. Heitor Vieira Dourado. Manaus, AM, Brazil. University of Campinas-UNICAMP. Department of Genetics, Evolution, Microbiology and Immunology. Laboratory of Tropical Diseases Prof. Dr. Luiz Jacintho da Silva. Campinas, SP, Brazil. University of Campinas-UNICAMP. Department of Genetics, Evolution, Microbiology and Immunology. Laboratory of Tropical Diseases Prof. Dr. Luiz Jacintho da Silva. Campinas, SP, Brazil. University of S?o Paulo. School of Pharmaceutical Sciences. S?o Paulo, SP, Brazil. Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Laborat?rio de Ensaios Cl?nicos e Imunogen?tica em Mal?ria. Ananindeua, PA, Brasil. Funda??o Oswaldo Cruz. Instituto Le?nidas & Maria Deane. Manaus, AM, Brazil / Funda??o de Medicina Tropical Dr. Heitor Vieira Dourado. Manaus, AM, Brazil. University of S?o Paulo. Institute of Biomedical Sciences. Department of Parasitology. S?o Paulo, SP, Brazil. Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Laborat?rio de Ensaios Cl?nicos e Imunogen?tica em Mal?ria. Ananindeua, PA, Brasil. Funda??o Oswaldo Cruz. Instituto Carlos Chagas. Curitiba, PR, Brazil. University of Campinas-UNICAMP. Department of Genetics, Evolution, Microbiology and Immunology. Laboratory of Tropical Diseases Prof. Dr. Luiz Jacintho da Silva. Campinas, SP, Brazil. BACKGROUND: The genetic diversity of malaria antigens often results in allele variant-specific immunity, imposing a great challenge to vaccine development. Rhoptry Neck Protein 2 (PvRON2) is a blood-stage antigen that plays a key role during the erythrocyte invasion of Plasmodium vivax. This study investigates the genetic diversity of PvRON2 and the naturally acquired immune response to P. vivax isolates. RESULTS: Here, the genetic diversity of PvRON21828-2080 and the naturally acquired humoral immune response against PvRON21828-2080 in infected and non-infected individuals from a vivax malaria endemic area in Brazil was reported. The diversity analysis of PvRON21828-2080 revealed that the protein is conserved in isolates in Brazil and worldwide. A total of 18 (19%) patients had IgG antibodies to PvRON21828-2080. Additionally, the analysis of the antibody response in individuals who were not acutely infected with malaria, but had been infected with malaria in the past indicated that 32 patients (33%) exhibited an IgG immune response against PvRON2. CONCLUSIONS: PvRON2 was conserved among the studied isolates. The presence of naturally acquired antibodies to this protein in the absence of the disease suggests that PvRON2 induces a long-term antibody response. These results indicate that PvRON2 is a potential malaria vaccine candidate.

Made available in DSpace on 2018-11-26T15:28:15Z (GMT). No. of bitstreams: 0 Previous issue date: 2015-11-01 Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Co-stimulatory molecules are essential in the orchestration of immune response and polymorphisms in their genes are associated with various diseases. However, in the case of variable allele frequencies among continental populations, this variation can lead to biases in genetic studies conducted in admixed populations such as those from Brazil. The aim of this study was to evaluate the influence of genomic ancestry on distributions of co-stimulatory genes polymorphisms in an admixed Brazilian population. A total of 273 individuals from the north of Brazil participated in this study. Nine single nucleotide polymorphisms in 7 genes (CD28, CTLA4, ICOS, CD86, CD40, CD40L and BLYS) were determined by polymerase chain reaction-restriction fragment length polymorphism. We also investigated 48 insertion/deletion ancestry markers to characterize individual African, European and Amerindian ancestry proportions in the samples. The analysis showed that the main contribution was European (43.9%) but also a significant contribution of African (31.6%) and Amerindian (24.5%) ancestry. ICOS, CD40L and CD86 polymorphisms were associated with genomic ancestry. However there were no significant differences in the proportions of ancestry for the other SNPs and haplotypes studied. Our findings reinforce the need to apply AIMs in genetic association studies involving these polymorphisms in the Brazilian population. (c) 2015 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved. Univ Estadual Paulista, Inst Biociencias Letras & Ciencias Exatas, Sao Jose Do Rio Preto, Brazil Fac Med Sao Jose do Rio Preto, Ctr Invest Microrganismos, Sao Jose Do Rio Preto, Brazil Fed Univ Para, Inst Ciencias Biol, BR-66059 Belem, Para, Brazil Univ Fed Sergipe, Dept Biol, Aracaju, Brazil Fed Univ Para, Lab Genet Humana & Med, BR-66059 Belem, Para, Brazil Secretaria Vigilancia Saude, Inst Evandro Chagas, Lab Pesquisas Basicas Malaria, Ananindeua, Brazil Univ Estadual Paulista, Inst Biociencias Letras & Ciencias Exatas, Sao Jose Do Rio Preto, Brazil CNPq: 471605/2011-5

The Plasmodium vivax Cell-traversal protein for ookinetes and sporozoites (PvCelTOS) plays an important role in the traversal of host cells. Although essential to PvCelTOS progress as a vaccine candidate, its genetic diversity remains uncharted. Therefore, we investigated the PvCelTOS genetic polymorphism in 119 field isolates from five different regions of Brazilian Amazon (Manaus, Novo Repartimento, Porto Velho, Plácido de Castro and Oiapoque). Moreover, we also evaluated the potential impact of non-synonymous mutations found in the predicted structure and epitopes of PvCelTOS. The field isolates showed high similarity (99.3% of bp) with the reference Sal-1 strain, presenting only four Single-Nucleotide Polymorphisms (SNP) at positions 24A, 28A, 109A and 352C. The frequency of synonymous C109A (82%) was higher than all others (p, Author summary Cell-traversal protein for ookinetes and sporozoites (CelTOS) presents a pivotal role in the cell traversal of host cells in mosquito and vertebrate hosts. For this reason, it has been considered a potential novel alternative for a vaccine against malaria caused by P. falciparum. However, little is known about its orthologous P. vivax CelTOS. Although the genetic diversity of this protein could be a limiting factor for acquisition of immunity and present implications for an effective vaccine development, it has never been explored. Thus, considering that the epidemiology of malaria in Brazil presents variable transmission rates and the knowledge on the genetic polymorphism of PvCelTOS remains unknown, we aimed to identify the pvceltos gene in isolates from five different regions of the Brazilian Amazon and to study the potential impacts of the genetic diversity of PvCelTOS in protein structures and predicted epitopes. Our findings indicate that PvCelTOS is an extremely conserved protein, presenting only four SNPs in the entire sequences of field isolates from Brazilian Amazon. The two non-synonymous mutations found in our field isolates presented no significant effect on the protein structure and a very low impact on potential T and B-cell epitopes indicated by our epitope prediction. Collectively, our data suggest that the small need to avoid the immune recognition by the human host and its importance on the parasite’s survival and transmission reflects a very conservative profile of pvceltos gene in field samples from Brazil and other endemic areas worldwide.

Background Plasmodium vivax is the most prevalent malaria species in Brazil. The parasite-host coevolutionary process can be viewed as an 'arms race', in which adaptive genetic changes in one are eventually matched by alterations in the other. Methods Following the candidate gene approach we analyzed the CD40, CD40L and BLYS genes that participate in B-cell co-stimulation, for associations with P. vivax malaria. The study sample included 97 patients and 103 controls. We extracted DNA using the extraction and purification commercial kit and identified the following SNPs: -1C > T in the CD40 gene, -726T > C in the CD40L gene and the -871C > T in the BLyS gene using PCR-RFLP. We analyzed the genotype and allele frequencies by direct counting. We also compared the observed with the expected genotype frequencies using the Hardy-Weinberg equilibrium. Results The allele and genotype frequencies for these SNPs did not differ statistically between patient and control groups. Gene-gene interactions were not observed between the CD40 and BLYS and between the CD40L and BLYS genes. Overall, the genes were in Hardy-Weinberg equilibrium. Significant differences were not observed among the frequencies of antibody responses against P. vivax sporozoite and erythrocytic antigens and the CD40 and BLYS genotypes. Conclusions The results of this study show that, although the investigated CD40, CD40L and BLYS alleles differ functionally, this variation does not alter the functionality of the molecules in a way that would interfere in susceptibility to the disease. The variants of these genes may influence the clinical course rather than simply increase or decrease susceptibility.

Made available in DSpace on 2018-11-26T16:48:24Z (GMT). No. of bitstreams: 0 Previous issue date: 2016-07-19 Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Para State Research Support Foundation (Fundacao de Amparo a Pesquisa do Estado do Para) CAPPES Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Background: The immune response against Plasmodium vivax immunogenic epitopes is regulated by pro-and anti-inflammatory cytokines that determine antibody levels and class switching. Cytokine gene polymorphisms may be responsible for changes in the humoral immune response against malaria. The aim of this study was to evaluate whether polymorphisms in the TNFA, IFNG and IL10 genes would alter the levels of anti-PvAMA1, PvDBP and -PvMSP-1(19) IgG antibodies in patients with vivax malaria. Methods: Samples from 90 vivax malaria-infected and 51 uninfected subjects from an endemic area of the Brazilian Amazon were genotyped using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) to identify polymorphisms of the genes TNFA (-1031T > C, -308G > A, -238G > A), IFNG (+874T > A) and IL10 (-819C > T, -592C > A). The levels of total IgG against PvAMA1, PvDBP and -PvMSP-1(19) were determined using an enzyme-linked immunosorbent assay (ELISA). Associations between the polymorphisms and the antibody response were assessed by means of logistic regression models. Results: No significant differences were found in the levels of IgG antibodies against the PvAMA-1, PvDBP or PvMSP-1(19) proteins in relation to the studied polymorphisms. Conclusions: Although no associations were found among the evaluated genotypes and alleles and anti-merozoite IgG class P. vivax antibody levels, this study helps elucidate the immunogenic profile involved in the humoral immune response in malaria. Sao Jose Rio Preto Med Sch, Dept Dermatol Infect & Parasit Dis, Sao Jose Do Rio Preto, SP, Brazil Sao Paulo State Univ, Dept Biol, Sao Jose Do Rio Preto, SP, Brazil Univ Fed Sergipe, Dept Biol, Lab Mol Genet & Biotechnol, Sao Cristovao, SE, Brazil Univ Fed Para, UFPA, Inst Biol Sci, Lab Microbiol & Immunol, Belem, Para, Brazil Univ Estadual Campinas, Lab Trop Dis, Dept Genet Evolut & Bioagents, Campinas, SP, Brazil Evandro Chagas Inst, Sect Parasitol, Lab Basic Res Malaria, Belem, PA, Brazil Sao Paulo State Univ, Dept Biol, Sao Jose Do Rio Preto, SP, Brazil CNPq: 472135/2012

S?o Paulo State University. Department of Biology. S?o Jos? do Rio Preto, SP, Brazil / S?o Jos? do Rio Preto Medical School. Department of Skin. Infectious and Parasitic Diseases. S?o Jos? do Rio Preto, SP, Brazil. S?o Jos? do Rio Preto Medical School. Department of Skin. Infectious and Parasitic Diseases. S?o Jos? do Rio Preto, SP, Brazil. S?o Jos? do Rio Preto Medical School. Department of Skin. Infectious and Parasitic Diseases. S?o Jos? do Rio Preto, SP, Brazil / S?o Paulo State University. Department of Biology. S?o Jos? do Rio Preto, SP, Brazil. Federal University of Sergipe. Department of Biology. S?o Crist?v?o, SE, Brazil. Oswaldo Cruz Foundation. Le?nidas and Maria Deane Institute. Manaus, AM, Brazil. Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Laborat?rio de Pesquisa B?sica de Mal?ria. Bel?m, PA, Brasil. Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Laborat?rio de Pesquisa B?sica de Mal?ria. Bel?m, PA, Brasil. Oswaldo Cruz Foundation. Le?nidas and Maria Deane Institute. Manaus, AM, Brazil. S?o Paulo State University. Department of Biology. S?o Jos? do Rio Preto, SP, Brazil / S?o Jos? do Rio Preto Medical School. Department of Skin. Infectious and Parasitic Diseases. S?o Jos? do Rio Preto, SP, Brazil / Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Laborat?rio de Pesquisa B?sica de Mal?ria. Bel?m, PA, Brasil. Background: Humoral immune responses against proteins of asexual blood-stage malaria parasites have been associated with clinical immunity. However, variations in the antibody-driven responses may be associated with a genetic component of the human host. The objective of the present study was to evaluate the influence of co-stimulatory molecule gene polymorphisms of the immune system on the magnitude of the humoral immune response against a Plasmodium vivax vaccine candidate antigen. Methods: Polymorphisms in the CD28, CTLA4, ICOS, CD40, CD86 and BLYS genes of 178 subjects infected with P. vivax in an endemic area of the Brazilian Amazon were genotyped by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). The levels of IgM, total IgG and IgG subclasses specific for ICB2-5, i.e., the N-terminal portion of P. vivax merozoite surface protein 1 (PvMSP-1), were determined by enzyme-linked immuno assay. The associations between the polymorphisms and the antibody response were assessed by means of logistic regression models. Results: After correcting for multiple testing, the IgG1 levels were significantly higher in individuals recessive for the single nucleotide polymorphism rs3116496 in CD28 (p = 0.00004). Furthermore, the interaction between CD28 rs35593994 and BLYS rs9514828 had an influence on the IgM levels (p = 0.0009). Conclusions: The results of the present study support the hypothesis that polymorphisms in the genes of co-stimulatory components of the immune system can contribute to a natural antibody-driven response against P. vivax antigens.

S?o Paulo State University. Department of Biology. S?o Jos? do Rio Preto, SP, Brazil. College of Medicine of S?o Jos? do Rio Preto. Department of Dermatologic, Infectious, and Parasitic Diseases. S?o Jos? do Rio Preto, SP, Brazil. S?o Paulo State University. Department of Biology. S?o Jos? do Rio Preto, SP, Brazil. Federal University of Sergipe. Department of Biology. Aracaju, SE, Brazil. Federal University of Par?. Institute of Biological Sciences. Laborat?rio of Microbiology and Immunology. Bel?m, PA, Brazil. Oswaldo Cruz Foundation. Laboratory of Malaria, Ren? Rachou Research Center. Belo Horizonte, MG, Brazil. Oswaldo Cruz Foundation. Laboratory of Malaria, Ren? Rachou Research Center. Belo Horizonte, MG, Brazil. University of S?o Paulo. Faculty of Pharmaceutical Sciences. Department of Clinical and Toxicological Analyses. S?o Paulo, SP, Brazil. Federal University of Par?. Laboratory of Human and Medical Genetics. Bel?m, PA, Brazil. Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Laborat?rio de Pesquisa B?sica em Mal?ria. Ananindeua, PA, Brasil. Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Laborat?rio de Pesquisa B?sica em Mal?ria. Ananindeua, PA, Brasil / S?o Paulo State University. Department of Biology. S?o Jos? do Rio Preto, SP, Brazil. The development of an effective immune response can help decrease mortality from malaria and its clinical symptoms. However, this mechanism is complex and has significant inter-individual variation, most likely owing to the genetic contribution of the human host. Therefore, this study aimed to investigate the influence of polymorphisms in genes involved in the costimulation of B-lymphocytes in the naturally acquired humoral immune response against proteins of the asexual stage of Plasmodium vivax. A total of 319 individuals living in an area of malaria transmission in the Brazilian Amazon were genotyped for four SNPs in the genes CD40, CD40L, BLYS and CD86. In addition, IgG antibodies against P. vivax apical membrane antigen 1 (PvAMA-1), Duffy binding protein (PvDBP) and merozoite surface protein 1 (PvMSP-119) were detected by ELISA. The SNP BLYS -871CT was associated with the frequency of IgG responders to PvAMA-1 and PvMSP-119. The SNP CD40 -1CT was associated with the IgG response against PvDBP, whereas IgG antibody titers against PvMSP-119 were influenced by the polymorphism CD86 +1057GA. These data may help to elucidate the immunological aspects of vivax malaria and consequently assist in the design of malaria vaccines.

Universidade Federal do Par?. Bel?m, PA, Brasil. Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Ananindeua, PA, Brasil. Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Ananindeua, PA, Brasil. Universidade Estadual Paulista. Botucatu, SP, Brasil. Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Ananindeua, PA, Brasil. Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Ananindeua, PA, Brasil. Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Ananindeua, PA, Brasil / Universidade Estadual Paulista. Botucatu, SP, Brasil. Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Ananindeua, PA, Brasil / Universidade Federal do Par?. Bel?m, PA, Brasil. O objetivo deste estudo foi investigar a frequ?ncia de gen?tipos da prote?na circunsporozo?ta (CSP) em sangue humano e sua correla??o com a parasitemia, bem como avaliar a presen?a desses gen?tipos em Anopheles no Munic?pio de Goian?sia do Par?, uma ?rea end?mica do sudeste do Estado do Par?, Brasil, de 2012 a 2013. Amostras de sangue foram coletadas de 118 pacientes com Plasmodium vivax e 369 mosquitos anofelinos. O gene da CSP foi genotipado usando-se a rea??o em cadeia da polimerase/polimorfismo de comprimento de fragmento de restri??o, e a infectividade dos anofelinos foi determinada pelo ELISA. A parasitemia variou de 5-70.000 parasitas/mm?, e os tr?s gen?tipos (VK210, VK247 e P. vivax-like) foram detectados tanto em infec??es simples quanto em mistas. Nenhuma amostra apresentou infec??o mista com todos os tr?s gen?tipos. O gen?tipo mais frequente foi o VK210, seguido pelo VK247 e o ?ltimo associado com os valores mais altos de parasitemia (p < 0,0001). Entre os mosquitos identificados, somente 11 esp?cimes foram infectados; de sete esp?cimes Anopheles darlingi, quatro foram infectados por Plasmodium falciparum, dois por VK210 e um por VK247. Os tr?s Anopheles albitarsis foram infectados por VK247 e um Anopheles nuneztovari por VK210. O gen?tipo VK210 continua sendo o mais prevalente no sudeste do Par?; entretanto, novas evid?ncias indicam a adapta??o do VK247. Os esp?cimes An. darlingi, An. albitarsis e An. nuneztovari desempenham um importante papel na transmiss?o dos gen?tipos CSP na ?rea de estudo. Essa descoberta pode ser um problema de sa?de p?blica devido ? possibilidade de ressurgimento de epidemias de mal?ria por P. vivax em comunidades suscet?veis. The objective of this study was therefore to investigate the frequency of circumsporozoite protein (CSP) genotypes in human blood and their correlation with parasitemia, as well as to evaluate the presence of these genotypes in Anopheles in the Municipality of Goian?sia do Par?, an endemic area of southeastern Par? State, Brazil from 2012-2013. Blood samples were collected from 118 patients with Plasmodium vivax and 369 anopheline mosquitoes. The CSP gene was genotyped using the polymerase chain reaction/restriction fragment length polymorphism technique, and the infectivity of the anophelines was determined using ELISA. Parasitemia ranged from 5-70,000 parasites/mm?, and the three genotypes (VK210, VK247, and P. vivax-like) were detected both in single and mixed infections. No sample exhibited mixed infection with all three genotypes. The most frequent genotype was VK210 followed by VK247 and the latter associated with the highest parasitemia values (p < 0.0001). Among the identified mosquitoes, only 11 specimens were infected; of the seven Anopheles darlingi specimens four were infected with Plasmodium falciparum, two with VK210, and one with VK247. The three Anopheles albitarsis specimens were infected with VK247, and one Anopheles nuneztovari specimen was infected with VK210. The VK210 genotype continues to be the most prevalent in southeastern Par?; however, a new evidence shows the adaptation of VK247. The species An. darlingi, An. albitarsis, and An. nuneztovari play an important role in the transmission of CSP genotypes in the study area. This finding may be a public health concern due to the possibility of resurgence of P. vivax malaria epidemics in susceptible communities.

The work reported in this manuscript was funded by the National Scientific and Technological Development in the Research Program for Health System Unique (CNPq PPSUS Public Notice No. 00077/2008/ MCT/CNPq/SETEC, Process No. 35.000.162/2009), and Government of the State of Amap?, and also, Central Public Health Laboratory (LACEN) of Amap? Secretaria de Estado da Sa?de do Amap?. Laborat?rio Central. Macap?, AP, Brazil / Universidade Federal do Par?. Bel?m, PA, Brazil. Universidade Federal do Par?. Bel?m, PA, Brazil. Universidade Estadual Paulista. Instituto de Bioci?ncias, Letras e Ci?ncias Exatas. S?o Jos? do Rio Preto, SP, Brazil. Secretaria de Estado da Sa?de do Amap?. Macap?, AP, Brazil Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Ananindeua, PA, Brasil / Universidade Federal do Par?. Bel?m, PA, Brazil / Universidade Estadual Paulista. Instituto de Bioci?ncias, Letras e Ci?ncias Exatas. S?o Jos? do Rio Preto, SP, Brazil. Universidade Federal do Par?. Bel?m, PA, Brazil. Institut Pasteur de la Guyane. Centre National de R?f?rence du Paludisme. Laboratoire de Parasitologie. Cayenne, French Guiana. Institut Pasteur de la Guyane. Centre National de R?f?rence du Paludisme. Laboratoire de Parasitologie. Cayenne, French Guiana. Universit? des Antilles et de la Guyane. Cayenne, French Guiana. Malaria is a major health problem for people who live on the border between Brazil and French Guiana. Here we discuss Plasmodium vivax distribution pattern in the town of Oiapoque, Amap? State using the circumsporozoite (CS) gene as a marker. Ninety-one peripheral blood samples from P. vivax patients have been studied. Of these, 64 individuals were from the municipality of Oiapoque (Amap? State, Brazil) and 27 patients from French Guiana (August to December 2011). DNA extraction was performed, and a fragment of the P. vivax CS gene was subsequently analyzed using PCR/RFLP. The VK210 genotype was the most common in both countries (48.36% in Brazil and 14.28% in French Guiana), followed by the P. vivax-like (1.10% in both Brazil and French Guiana) and VK247 (1.10% only in Brazil) in single infections. We were able to detect all three CS genotypes simultaneously in mixed infections. There were no statistically significant differences either regarding infection site or parasitaemia among individuals with different genotypes. These results suggest that the same genotypes circulating in French Guiana are found in the municipality of Oiapoque in Brazil. These findings suggest that there may be a dispersion of parasitic populations occurring between the two countries. Most likely, this distribution is associated with prolonged and/or more complex transmission patterns of these genotypes in Brazil, bordering French Guiana.

Interleukin 2 (IL-2) is a monomeric glycoprotein that is primarily produced by activated CD4+ T cells, CD8+ T cells and dendritic cells. It is characterized as a proinflammatory cytokine that is secreted by Th1 cells. IL-2 plays a central role in the activation of regulatory T cells to produce the cytokines tumor necrosis factor alpha (TNF-α) and interferon gamma (IFN-γ). IL-2 may also enhance the cytolytic activity of natural killer cells, thereby ensuring their significance in the control of the immune response, and effectively participate in the pathogenesis of several pathological conditions, such as cancer and metabolic, infectious, autoimmune and inflammatory diseases. We emphasize the importance of studies of IL-2 and discuss perspectives resulting from our increasing understanding of genetic diversity and its role in the immune response.

Made available in DSpace on 2018-11-26T17:13:49Z (GMT). No. of bitstreams: 0 Previous issue date: 2016-01-01 Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Polymorphisms in cytokine genes can alter the production of these proteins and consequently affect the immune response. The trihybrid heterogeneity of the Brazilian population is characterized as a condition for the use of ancestry informative markers. The objective of this study was to evaluate the frequency of -1031T>C, -308G>A and -238G>A TNFA, +874 A>T IFNG and -819C>T, and -592C>A IL10 gene polymorphisms and their association with malaria vivax and genomic ancestry. Samples from 90 vivax malaria-infected individuals and 51 noninfected individuals from northern Brazil were evaluated. Genotyping was carried out by using ASO-PCR or PCR/RFLP. The genomic ancestry of the individuals was classified using 48 insertion/deletion polymorphism biallelic markers. There were no differences in the proportions of African, European, and Native American ancestry between men and women. No significant association was observed for the allele and genotype frequencies of the 6 SNPs between malaria-infected and noninfected individuals. However, there was a trend toward decreasing the frequency of individuals carrying the TNF-308A allele with the increasing proportion of European ancestry. No ethnic-specific SNPs were identified, and there was no allelic or genotype association with susceptibility or resistance to vivax malaria. Understanding the genomic mechanisms by which ancestry influences this association is critical and requires further study. Coll Med Sao Jose do Rio Preto, Dept Dermatol Infect & Parasit Dis, Sao Jose Do Rio Preto, SP, Brazil UNIRP, Univ Ctr Rio Preto, Sao Jose Do Rio Preto, SP, Brazil Univ Estadual Campinas, Dept Genet Evolut & Bioagents, Lab Trop Dis Prof Luiz Jacintho da Silva, Campinas, SP, Brazil Sao Paulo State Univ, Dept Biol, Sao Jose Do Rio Preto, SP, Brazil Fed Univ Para, Lab Human & Med Genet, Belem, PA, Brazil Evandro Chagas Inst, Sect Parasitol, Lab Basic Res Malaria, Belem, PA, Brazil Sao Paulo State Univ, Dept Biol, Sao Jose Do Rio Preto, SP, Brazil CNPq: 472135/2012-0

Made available in DSpace on 2015-04-27T11:55:53Z (GMT). No. of bitstreams: 0 Previous issue date: 2010Bitstream added on 2015-04-27T12:08:27Z : No. of bitstreams: 1 ISSN2176-6223-2010-01-04-51-55-pt.pdf: 492582 bytes, checksum: 0cac8184e90166467328b352ef211bb3 (MD5)Bitstream added on 2015-04-27T12:08:28Z : No. of bitstreams: 2 ISSN2176-6223-2010-01-04-51-55-pt.pdf: 492582 bytes, checksum: 0cac8184e90166467328b352ef211bb3 (MD5) ISSN2176-6223-2010-01-04-51-55-es.pdf: 494680 bytes, checksum: 25f6c23325eeb284531767af45a9119c (MD5)Bitstream added on 2015-04-27T12:08:28Z : No. of bitstreams: 3 ISSN2176-6223-2010-01-04-51-55-pt.pdf: 492582 bytes, checksum: 0cac8184e90166467328b352ef211bb3 (MD5) ISSN2176-6223-2010-01-04-51-55-es.pdf: 494680 bytes, checksum: 25f6c23325eeb284531767af45a9119c (MD5) ISSN2176-6223-2010-01-04-51-55-en.pdf: 492026 bytes, checksum: f9d7f82627e4aafae345fab86dc5fc8f (MD5) Este estudo avaliou a frequência de diferentes alelos HLA-DRB1 em indivíduos infectados por Plasmodium vivax e em doadores de sangue saudáveis provenientes de áreas endêmicas de malária do Brasil. Foi realizada uma genotipagem de baixa resolução dos alelos HLA-DRB1 em 73 pacientes com malária e em 29 doadores de sangue saudáveis. Os alelos mais frequentes em indivíduos do norte do Brasil foram HLA-DRB1 *04, *08, *07 e *13. A frequência de HLA-DRB1 *07 foi maior nos indivíduos infectados com malária do que no grupo controle, o que reforça a hipótese de que esse alelo desempenha um papel importante na suscetibilidade à malária. Esta pesquisa fornece novas informações sobre um fator potencial de suscetibilidade à malária por P. vivax em uma população brasileira naturalmente exposta à doença. We evaluated the frequency of different HLA-DRB1 alleles in Plasmodium vivax-infected individuals and in healthy blood donors from malaria endemic areas of Brazil. Low-resolution human leukocyte antigen-DRB1 genotyping was performed for 73 malaria patients and 29 healthy blood donors. The most frequent alleles in individuals from northern Brazil were human leukocyte antigen-DRB1*04, *08, *07 and *13. The frequency of human leukocyte antigen-DRB1*07 was higher in malaria-infected individuals than in the control group, which reinforces the theory that this allele plays an important role in susceptibility to malaria. This study offers new information about a potential susceptibility factor for P. vivax malaria in a Brazilian population that is naturally exposed to malaria. Este estudio evaluó la frecuencia de diferentes alelos HLA-DRB1 en individuos infectados por Plasmodium vivax y en donantes de sangre saludables provenientes de áreas endémicas de malaria de Brasil. Se realizó un genotipado de baja resolución de los alelos HLA-DRB1 en 73 pacientes con malaria y en 29 donantes de sangre saludables. Los alelos más frecuentes en individuos del norte de Brasil fueron HLA-DRB1*04, *08, *07 y *13. La frecuencia de HLA-DRB1*07 fue más grande en individuos infectados con malaria que en el grupo control, lo que refuerza la hipótesis de que ese alelo desempeña un papel importante en la susceptibilidad a la malaria. Esta investigación suministra nuevas informaciones sobre un factor potencial de susceptibilidad a la malaria por P. vivax en una población brasileña naturalmente expuesta a la enfermedad. Universidade Estadual Paulista Júlio de Mesquita Filho, Departamento de Biologia, Instituto de Biociências Letras e Ciências Exatas de São José do Rio Preto, Sao Jose do Rio Preto, Rua Cristóvão Colombo, 2265, Jardim Nazareth, CEP 15054-000, SP, Brasil Universidade Estadual Paulista Júlio de Mesquita Filho, Departamento de Biologia, Instituto de Biociências Letras e Ciências Exatas de São José do Rio Preto, Sao Jose do Rio Preto, Rua Cristóvão Colombo, 2265, Jardim Nazareth, CEP 15054-000, SP, Brasil

Background Several studies have recently demonstrated that the immune responses against malaria is governed by different factors, including the genetic components of the host. The IL-4 gene appears to be a strong candidate factor because of its role in the regulation of the Th2 response. The present study investigated the role of IL-4 polymorphisms in the development of IgG antibodies against PvAMA-1 and the IL-4 levels in individuals infected with Plasmodium vivax in a malaria endemic area in the Brazilian Amazon. Methods The study sample included 83 patients who were diagnosed with P. vivax infection using thick smear and confirmed by nested-PCR. The IL-4 −590 C>T and IL-4 −33 C>T polymorphisms were genotyped by PCR–RFLP, and the intron 3 VNTR was genotyped by PCR. A standardised ELISA protocol was used to measure the total IgG against PvAMA-1. The cytokine/chemokine levels were measured using a Milliplex multiplex assay (Millipore). All of the subjects were genotyped with 48 ancestry informative markers to determine the proportions of African, European and Amerindian ancestry using STRUCTURE software. Results Of the 83 patients, 60 (73%) produced IgG antibodies against PvAMA-1. A significant decrease in the percentage of respondents was observed among the primo-infected individuals. No significant differences were observed in the frequencies of genotypes and haplotypes among individuals who were positive or negative for IgG antibodies against PvAMA-1. Furthermore, no significant correlation was observed between the IL-4 polymorphisms, antibody levels, IL-4 levels, and parasitemia. Conclusions This study indicated that the polymorphisms identified in the IL-4 gene are not likely to play a role in the regulation of the antibody response against PvAMA-1 and IL-4 production in vivax malaria.

Made available in DSpace on 2015-04-27T11:55:52Z (GMT). No. of bitstreams: 0 Previous issue date: 2013Bitstream added on 2015-04-27T12:08:32Z : No. of bitstreams: 1 ISSN2036-7481-2013-04-01-26-31.pdf: 651293 bytes, checksum: c1ac1e4545c616387c1901b3605fe9ed (MD5) In the period from July 2009 to October 2010, fecal samples from 61 animals and 154 humans from the municipality of Aracatuba (São Paulo State, Brazil) were studied. Fecal samples from animals were collected in the Municipal Animal Shelter and the Veterinary Hospital of the Universidade Estadual Paulista. Human fecal specimens were collected in playschools in the outskirts of the city by the private network of clinical analysis laboratories of the municipal. Diagnosis was done by optical microscopy using the Faust and Hoffmann, Pons and Janer techniques. The genotypes of Giardia intestinalis were characterized by PCR-RFLP and confirmed by sequencing the ß-giardin gene. Human specimens were positive in 25.3% (39/154) of the cases with 26.8% (36/134) of the specimens from children and 15% (3/20) from adults being positive. The frequency of G. intestinalis among the animals was 23.0% (14/61). A total of 32 isolates of G. intestinalis obtained from human feces and six from dogs and cats were characteristic of the A genotype (AI and AII/AIII). The results of this study in respect to frequency of giardiasis are similar to reported in most studies in Brazil. The prevalence observed in animal populations conforms to worldwide infection rates. G. intestinalis genotypes considered zoonotic were detected in both pets and humans from the city of Aractuba, suggesting a possible zoonotic transmission of the parasite in the northwestern region of São Paulo State. The absence of these genotypes in farm animals may imply that they are not involved in the chain of transmission to humans in this region. Universidade Estadual Paulista Júlio de Mesquita Filho, Departamento de Biologia, Instituto de Biociências Letras e Ciências Exatas de São José do Rio Preto, São José do Rio Preto, Rua Cristóvão Colombo, 2265, Jardim Nazaré, CEP 15054000, SP, Brasil Universidade Estadual Paulista Júlio de Mesquita Filho, Departamento de Clínica, Cirurgia e Reprodução Animal, Faculdade de Odontologia de Araçatuba Universidade Estadual Paulista Júlio de Mesquita Filho, Departamento de Biologia, Instituto de Biociências Letras e Ciências Exatas de São José do Rio Preto

Mechanisms involved in severe P. vivax malaria remain unclear. Parasite polymorphisms, parasite load and host cytokine profile may influence the course of infection. In this study, we investigated the influence of circumsporozoite protein (CSP) polymorphisms on parasite load and cytokine profile in patients with vivax malaria. A cross-sectional study was carried out in three cities: São Luís, Cedral and Buriticupu, Maranhão state, Brazil, areas of high prevalence of P. vivax. Interleukin (IL)-2, IL-4, IL-10, IL-6, IL-17, tumor necrosis factor alpha (TNF-α, interferon gamma (IFN-γ and transforming growth factor beta (TGF-β were quantified in blood plasma of patients and in supernatants from peripheral blood mononuclear cell (PBMC) cultures. Furthermore, the levels of cytokines and parasite load were correlated with VK210, VK247 and P. vivax-like CSP variants. Patients infected with P. vivax showed increased IL-10 and IL-6 levels, which correlated with the parasite load, however, in multiple comparisons, only IL-10 kept this association. A regulatory cytokine profile prevailed in plasma, while an inflammatory profile prevailed in PBMC culture supernatants and these patterns were related to CSP polymorphisms. VK247 infected patients showed higher parasitaemia and IL-6 concentrations, which were not associated to IL-10 anti-inflammatory effect. By contrast, in VK210 patients, these two cytokines showed a strong positive correlation and the parasite load was lower. Patients with the VK210 variant showed a regulatory cytokine profile in plasma, while those infected with the VK247 variant have a predominantly inflammatory cytokine profile and higher parasite loads, which altogether may result in more complications in infection. In conclusion, we propose that CSP polymorphisms is associated to the increase of non-regulated inflammatory immune responses, which in turn may be associated with the outcome of infection., Author Summary Recent evidences have associated P. vivax infections with clinical complications, previously only attributed to P. falciparum malaria. The interaction between host and parasite may contribute to severity of the disease, however, the specific contribution of each factor remains unclear. Previous studies have shown that polymorphisms in Plasmodium vivax CSP may interfere in systemic reactions, response to drug treatments, leading to different symptoms as well as humoral responses. In this study, we investigate whether these polymorphisms could influence the parasite load and cytokine profile, which altogether may influence the malaria outcome. In this sense, studies have shown that subjects with high parasitic loads, responding with production of pro-inflammatory cytokines develop a more severe disease. The present data indicate that VK247 variant are associated with significant higher parasite loads and pro-inflammatory cytokine profile compared to VK210 variant. In this regard, this study demonstrates that P. vivax CSP polymorphisms have systemic effects in the host immune response, and the investigation of immunogenicity of parasite proteins may provide evidences for a better understanding of this infection.

Made available in DSpace on 2013-09-27T14:52:56Z (GMT). No. of bitstreams: 1 WOS000300132800014.pdf: 212581 bytes, checksum: 6535572dbec9f900d22c06ab647009a4 (MD5) Previous issue date: 2012-02-01 Made available in DSpace on 2013-09-30T19:24:38Z (GMT). No. of bitstreams: 1 WOS000300132800014.pdf: 212581 bytes, checksum: 6535572dbec9f900d22c06ab647009a4 (MD5) Previous issue date: 2012-02-01 Submitted by Vitor Silverio Rodrigues (vitorsrodrigues@reitoria.unesp.br) on 2014-05-20T15:34:42Z No. of bitstreams: 1 WOS000300132800014.pdf: 212581 bytes, checksum: 6535572dbec9f900d22c06ab647009a4 (MD5) Made available in DSpace on 2014-05-20T15:34:42Z (GMT). No. of bitstreams: 1 WOS000300132800014.pdf: 212581 bytes, checksum: 6535572dbec9f900d22c06ab647009a4 (MD5) Previous issue date: 2012-02-01 Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) COLCIENCIAS Ministry of Social Protection National Institute of Allergy and Infectious Diseases (NIAID) We evaluated the influence of allelic frequency of the human leukocyte antigen (HLA) -DRB1 on the acquisition of antibody response against malaria sporozoite and merozoite peptides in patients with Plasmodium vivax malaria acquired in endemic areas of Brazil. IgG antibodies were detected by enzyme-linked immunosorbent assay against four peptides of circumsporozoite protein (CSP) (amino, carboxyl, and VK210 and VK247 repeats) and peptides of merozoite surface protein 1 (MSP-1), apical membrane antigen 1 (AMA-1), and Duffy-binding protein (DBP). We found an association between HLA-DR3 and HLA-DR5 alleles and lack of antibody response to CSP amino terminal, as well as an association between HILA-DR3 and the highest antibody response to MSP1 (Pv200L). In conclusion, we suggest a potential regulatory role of the H1A-DRB1 alleles in the production of antibodies to a conserved region of P. vivax CSP and MSP1 in Brazilian population exposed to malaria. (C) 2011 Elsevier B.V. All rights reserved. Univ Fed Sergipe, Dept Biociencias, BR-49500000 Itabaiana, SE, Brazil Faculdade de Medicina de São José do Rio Preto (FAMERP), Dept Doencas Dermatol Infecciosas & Parasitarias, Ctr Invest Microrganismos, BR-15090000 São Paulo, Brazil Univ Fed Triangulo Mineiro, Inst Ciencias Exatas Nat & Educ, BR-38025180 Uberaba, MC, Brazil Univ Estadual Paulista, BR-15054000 São Paulo, Brazil Nucleo Estadual Amapa CRDT, Minist Saude, BR-68900430 Macapa, Amapa, Brazil Secretaria Vigilancia Saude, Inst Evandro Chagas, Lab Pesquisas Basicas Malaria, BR-6703000 Ananindeua, Para, Brazil Univ São Paulo, Fac Ciencias Farmaceut, BR-05508900 São Paulo, Brazil Fundação Oswaldo Cruz, Ctr Pesquisas Rene Rachou, Lab Malaria, BR-30190002 Belo Horizonte, MG, Brazil Univ Valle, Fac Salud, Inst Inmunol, Cali, Colombia Univ Fed Fluminense, Inst Biomed, Dept Microbiol & Parasitol, BR-24210130 Rio de Janeiro, Brazil Faculdade de Medicina de São José do Rio Preto (FAMERP), Dept Epidemiol, BR-15090000 São Paulo, Brazil Faculdade de Medicina de São José do Rio Preto (FAMERP), Dept Biol Mol, Lab Imunohematol, BR-15090000 São Paulo, Brazil Univ Estadual Paulista, BR-15054000 São Paulo, Brazil FAPESP: 02/09546-1 FAPESP: 06/09546-1 CNPq: 302353/03-8 CNPq: 410405/2006-0 COLCIENCIAS: 1106-04-16489) Ministry of Social Protection: 2304-04-19524 Ministry of Social Protection: 253-2005 National Institute of Allergy and Infectious Diseases (NIAID): 49486/TMRC

Frequency and levels of IgG antibodies to an N-terminal fragment of the Plasmodium vivax MSP-1 (Pv200L) protein, in individuals naturally exposed to malaria in four endemic areas of Brazil, were evaluated by enzyme-linked immunosorbent assay. Plasma samples of 261 P. vivax-infected individuals from communities of Macapa, Novo Repartimento, Porto Velho, and Placido de Castro in the Amazonian region with different malaria transmission intensities. A high mean number of studied individuals (89.3%) presented with antibodies to the Pv200L that correlated with the number of previous malaria infections; there were significant differences in the frequency of the responders (71.9–98.7) and in the antibody levels (1:200–1:51,200) among the four study areas. Results of this study provide evidence that Pv200L is a naturally immunogenic fragment of the PvMSP-1 and is associated with the degree of exposure to parasites. The fine specificity of antibodies to Pv200L is currently being assessed.

Made available in DSpace on 2013-08-28T14:08:24Z (GMT). No. of bitstreams: 1 WOS000280170500002.pdf: 996199 bytes, checksum: 9083a09051da6c2f8c0223ae75897080 (MD5) Made available in DSpace on 2013-09-30T18:42:06Z (GMT). No. of bitstreams: 2 WOS000280170500002.pdf: 996199 bytes, checksum: 9083a09051da6c2f8c0223ae75897080 (MD5) WOS000280170500002.pdf.txt: 35215 bytes, checksum: bc2641f7411a84598733b43bf7ecafd7 (MD5) Previous issue date: 2010-06-23 Submitted by Vitor Silverio Rodrigues (vitorsrodrigues@reitoria.unesp.br) on 2014-05-20T14:01:14Z No. of bitstreams: 2 WOS000280170500002.pdf: 996199 bytes, checksum: 9083a09051da6c2f8c0223ae75897080 (MD5) WOS000280170500002.pdf.txt: 35215 bytes, checksum: bc2641f7411a84598733b43bf7ecafd7 (MD5) Made available in DSpace on 2014-05-20T14:01:14Z (GMT). No. of bitstreams: 2 WOS000280170500002.pdf: 996199 bytes, checksum: 9083a09051da6c2f8c0223ae75897080 (MD5) WOS000280170500002.pdf.txt: 35215 bytes, checksum: bc2641f7411a84598733b43bf7ecafd7 (MD5) Previous issue date: 2010-06-23 Background: Plasmodium vivax circumsporozoite variants have been identified in several geographical areas. The real implication of the genetic variation in this region of the P. vivax genome has been questioned for a long time. Although previous studies have observed significant association between VK210 and the Duffy blood group, we present here that evidences of this variation are limited to the CSP central portion.Methods: The phylogenetic analyses were accomplished starting from the amplification of conserved domains of 18 SSU RNAr and Cyt B. The antibodies responses against the CSP peptides, MSP-1, AMA-1 and DBP were detected by ELISA, in plasma samples of individuals infected with two P. vivax CS genotypes: VK210 and P. vivax-like.Results: These analyses of the two markers demonstrate high similarity among the P. vivax CS genotypes and surprisingly showed diversity equal to zero between VK210 and P. vivax-like, positioning these CS genotypes in the same clade. A high frequency IgG antibody against the N- and C-terminal regions of the P. vivax CSP was found as compared to the immune response to the R- and V-repetitive regions (p = 0.0005, Fisher's Exact test). This difference was more pronounced when the P. vivax-like variant was present in the infection (p = 0.003, Fisher's Exact test). A high frequency of antibody response against MSP-1 and AMA-1 peptides was observed for all P. vivax CS genotypes in comparison to the same frequency for DBP.Conclusions: This results target that the differences among the P. vivax CS variants are restrict to the central repeated region of the protein, mostly nucleotide variation with important serological consequences. Univ Estadual Paulista, Dept Biol, Sao Jose do Rio Preto, SP, Brazil Fac SEAMA, Macapa, Amapa, Brazil Univ São Paulo, Fac Ciencias Farmaceut, Dept Anal Clin & Toxicol, BR-05508 São Paulo, Brazil Fundação Oswaldo Cruz, Ctr Pesquisas Rene Rachou, Belo Horizonte, MG, Brazil Fed Univ Para, Inst Ciencias Biol, Ananindeua, Para, Brazil MS SVS, Inst Evandro Chagas, Ananindeua, Para, Brazil Univ Valle, Inst Imunol, Cali, Colombia Fac Med Sao Jose do Rio Preto, Dept Doencas Dermatol Infecciosas & Parasitarias, Ctr Invest Microrg, Sao Jose Dos Campos, Brazil Univ Estadual Paulista, Dept Biol, Sao Jose do Rio Preto, SP, Brazil

The identification of Plasmodium species in Anopheles mosquitoes is an integral component of malaria control programs. We developed a new assay to identify Plasmodium falciparum, Plasmodium malariae, and Plasmodium vivax variants. Specific primers were designed to hybridize to CS gene-specific regions. Polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) were used to distinguish the P. vivax variants VK210, VK247, and P. vivax-like. The new PCR-RFLP assay revealed good agreement when compared with a nested PCR using artificially infected Anopheles mosquitoes. This sensitive PCR-RFLP method can be useful when detection of Plasmodium species and P. vivax variants is required and may be employed to improve the understanding of malaria transmission dynamics by Anopheles species.

Made available in DSpace on 2022-04-29T08:47:52Z (GMT). No. of bitstreams: 0 Previous issue date: 2009-07-01 The circumsporozoite protein (CSP) of the Plasmodium vivax infective sporozoite is considered to be a major target for the development of recombinant malaria vaccines. The Duffy blood group molecule acts as the red blood cell receptor for P. vivax. We review the frequency of P. vivax CSP variants and report their association with the Duffy blood group genotypes from Brazilian Amazon patients carrying P. vivax malaria. Peripheral blood samples were collected from 155 P. vivax-infected individuals from five Brazilian malaria-endemic areas. The P. vivax CSP variants and the Duffy blood group genotypes were assessed using PCR/RFLP. In single infections, the VK210 variant was the commonest followed by the P. vivax-like variant. The typing of P. vivax indicated that the frequency of variants among the study areas was significantly different from one to another. This is the first detection of the VK247 and P. vivax-like variant in single infections in endemic areas of Brazil. Association of the CSP P. vivax variants with the heterozygous Duffy blood group system genotype was significant for VK210 single infection. These observations provide additional data on the Plasmodium-host interactions concerning the Duffy blood group and P. vivax capability of causing human malaria. © 2008 Royal Society of Tropical Medicine and Hygiene. University of São Paulo State Júlio Mesquita Filho, Rua Cristovao Colombo 2265, 15054-000 Sao Jose do Rio Preto Sao Paulo State Faculty of Medicine of São José do Rio Preto, Ave. Brigadeiro Faria Lima 5416, Vila Sao Pedro, Sao Jose do Rio Preto, São Paulo State Mato Grosso Federal University Department of Medical Clinic, Rua Luiz Phellipe Pereira Leite, 78048-902 Cuiaba Mato Grosso State SEAMA Faculty, Macapá, Amapá State Evandro Chagas Institute MS/SVS, Rodovia BR, -316 km 7 s/n, Levilandia 67030-000 Ananindeua, Para State Faculty of Medicine of São José Rio Preto Foundation, Avenida Brigadeiro Faria Lima 5416, 15090-000 Sao Jose do Rio Preto Sao Paulo State University of São Paulo State Júlio Mesquita Filho, Rua Cristovao Colombo 2265, 15054-000 Sao Jose do Rio Preto Sao Paulo State

BACKGROUND The central repetitive region (CRR) of the Plasmodium vivax circumsporozoite surface protein (CSP) is composed of a repetitive sequence that is characterised by three variants: VK210, VK247 and P. vivax-like. The most important challenge in the treatment of P. vivax infection is the possibility of differential response based on the parasite genotype. OBJECTIVES To characterise the CSP variants in P. vivax isolates from individuals residing in a malaria-endemic region in Brazil and to profile these variants based on sensitivity to chloroquine and mefloquine. METHODS The CSP variants were determined by sequencing and the sensitivity of the P. vivax isolates to chloroquine and mefloquine was determined by Deli-test. FINDINGS Although five different allele sizes were amplified, the sequencing results showed that all of the isolates belonged to the VK210 variant. However, we observed substantial genetic diversity in the CRR, resulting in the identification of 10 different VK210 subtypes. The frequency of isolates that were resistant to chloroquine and mefloquine was 11.8 and 23.8%, respectively. However, we did not observe any difference in the frequency of the resistant isolates belonging to the VK210 subtypes. MAIN CONCLUSION The VK210 variant is the most frequently observed in the studied region and there is significant genetic variability in the CRR of the P. vivax CSP. Moreover, the antimalarial drug sensitivity profiles of the isolates does not seem to be related to the VK210 subtypes.

Submitted by Vitor Silverio Rodrigues (vitorsrodrigues@reitoria.unesp.br) on 2014-05-27T11:25:22Z No. of bitstreams: 0Bitstream added on 2014-05-27T14:31:01Z : No. of bitstreams: 1 2-s2.0-84855452304.pdf: 229414 bytes, checksum: ea25b3e0865094e29f65eef7b07467d0 (MD5) Made available in DSpace on 2014-05-27T11:25:22Z (GMT). No. of bitstreams: 0 Previous issue date: 2010-12-01 We investigated the ABO genotypes and heterogeneity of the O alleles in Plasmodium falciparum-infected and non-infected individuals from the Brazilian Amazon region. Sample collection took place from May 2003 to August 2005, from P. falciparum malaria patients from four endemic regions of the Brazilian Amazon. The control group consisted of donors from four blood banks in the same areas. DNA was extracted using the Easy-DNA(TM) extraction kit. ABO genotyping was performed using PCR/RFLP. There was a high frequency of ABO*O01O01. ABO*AO01 was the second most frequent genotype, and the third most frequent genotype was ABO*BO01. There were low frequencies of the ABO*O01O02, ABO*AA, ABO*AB, ABO*BB, and ABO*O02O02 genotypes. We analyzed the alleles of the O phenotype; the O(1variant) allele was the most frequent, both in malaria and non-malaria groups; consequently, the homozygous genotype O(1)(v)O(1)(v) was the most frequently observed. There was no evidence of the homozygous O(2) allele. Significant differences were not detected in the frequency of individuals with the various alleles in the comparison of the malaria patients and the general population (blood donors).