Hitchhiking Trypanosoma cruzi minicircle DNA affects gene expression in human host cells via LINE-1 retrotransposon

The horizontal transfer of Trypanosoma cruzi mitochondrial minicircle DNA to the genomes of naturally infected humans may play an important role in the pathogenesis of Chagas disease. Minicircle integrations within LINE-1 elements create the potential for foreign DNA mobility within the host genome via the machinery associated with this retrotransposon. Here we document integration of minicircle DNA fragments in clonal human macrophage cell lines and their mobilization over time. The movement of an integration event in a clonal transfected cell line was tracked at three months and three years post-infection. The minicircle sequence integrated into a LINE-1 retrotransposon; one such foreign fragment subsequently relocated to another genomic location in association with associated LINE-1 elements. The p15 locus was altered at three years as a direct effect of minicircle/LINE-1 acquisition, resulting in elimination of p15 mRNA. Here we show for the first time a molecular pathology stemming from mobilization of a kDNA/LINE-1 mutation. These genomic changes and detected transcript variations are consistent with our hypothesis that minicircle integration is a causal component of parasite-independent, autoimmune-driven lesions seen in the heart and other target tissues associated with Chagas disease.

Attaining the minimum target of resolution WHA 54.19 for schistosomiasis control in the Rainforest Zone of the state of Pernambuco, Northeastern Brazil

Resolution 19 of the 54th World Health Assembly (WHA-54.19) urged member nations to promote preventive measures, ensure treatment and mobilize resources for control of schistosomiasis and soil-transmitted helminthiases (STH). The minimum target is to attend 75% of all school-age children at risk by year 2010. The Brazilian Ministry of Health (MoH) recommends biennial surveys of whole communities and treatment of the positives through the Schistosomiasis Control Program within the Unified Health System (PCE-SUS). However, by 2004 the PCE-SUS had covered only 8.4% of the 1.2 million residents in the Rainforest Zone of Pernambuco (ZMP). Six of the 43 municipalities still remained unattended. Only three of the municipalities already surveyed reached coverage of 25% or more. At least 154 thousand children in the 7-14 years old range have to be examined (and treated if positive) within the next five years to attend the minimum target of the WHA 54.19 for the ZMP. To make this target feasible, it is suggested that from 2006 to 2010 the PCE-SUS actions should be complemented with school-based diagnosis and treatment, involving health and educational organs as well as community associations to include both children in schools and non-enrolled school-age children.

Improving the production of the denatured recombinant N-terminal domain of rhoptry-associated protein 2 from a Plasmodium falciparum target in the pathology of anemia in falciparum malaria

Rhoptry-associated protein 2 (RAP2) is known to be discharged from rhoptry onto the membrane surface of infected and uninfected erythrocytes (UEs) ex vivo and in vitro and this information provides new insights into the understanding of the pathology of severe anemia in falciparum malaria. In this study, a hexahistidine-tagged recombinant protein corresponding to residues 5-190 of the N-terminal of Plasmodium falciparum RAP2 (rN-RAP2) was produced using a new method of solubilization and purification. Expression was induced with D-lactose, a less expensive alternative inducer to the more common isopropyl-²-D-thio-galactopyranosidase. The recombinant protein was purified using two types of commercially-available affinity columns, iminodiacetic and nitrilotriacetic. rN-RAP2 had immunogenic potential, since it induced high titers of anti-RAP2 antibodies in mice. These antibodies recognized full-length RAP2 prepared from Triton X-100 extracts from two strains of P. falciparum. In fact, the antibody recognized a 29-kDa product of RAP2 cleavage as well as 82 and 70-kDa products of RAP1 cleavage. These results indicate that the two antigens share sequence epitopes. Our expressed protein fragment was shown to contain a functional epitope that is also present in rhoptry-derived ring surface protein 2 which attaches to the surface of both infected and UEs and erythroid precursor cells in the bone marrow of malaria patients. Serum from malaria patients who developed anemia during infection recognized rN-RAP2, suggesting that this protein fragment may be important for epidemiological studies investigating whether immune responses to RAP2 exacerbate hemolysis in falciparum malaria patients.

A new approach for potential drug target discovery through in silico metabolic pathway analysis using Trypanosoma cruzi genome information

The current drug options for the treatment of chronic Chagas disease have not been sufficient and high hopes have been placed on the use of genomic data from the human parasite Trypanosoma cruzi to identify new drug targets and develop appropriate treatments for both acute and chronic Chagas disease. However, the lack of a complete assembly of the genomic sequence and the presence of many predicted proteins with unknown or unsure functions has hampered our complete view of the parasite's metabolic pathways. Moreover, pinpointing new drug targets has proven to be more complex than anticipated and has revealed large holes in our understanding of metabolic pathways and their integrated regulation, not only for this parasite, but for many other similar pathogens. Using an in silicocomparative study on pathway annotation and searching for analogous and specific enzymes, we have been able to predict a considerable number of additional enzymatic functions in T. cruzi. Here we focus on the energetic pathways, such as glycolysis, the pentose phosphate shunt, the Krebs cycle and lipid metabolism. We point out many enzymes that are analogous to those of the human host, which could be potential new therapeutic targets.

Search for a platelet-activating factor receptor in the Trypanosoma cruzi proteome: a potential target for Chagas disease chemotherapy

Chagas disease (CD) causes the highest burden of parasitic diseases in the Western Hemisphere and is therefore a priority for drug research and development. Platelet-activating factor (PAF) causes the CD parasite Trypanosoma cruzi to differentiate, which suggests that the parasite may express PAF receptors. Here, we explored the T. cruzi proteome for PAF receptor-like proteins. From a total of 23,000 protein sequences, we identified 29 hypothetical proteins that are predicted to have seven transmembrane domains (TMDs), which is the main characteristic of the G protein-coupled receptors (GPCRs), including the PAF receptor. The TMDs of these sequences were independently aligned with domains from 25 animal PAF receptors and the sequences were analysed for conserved residues. The conservation score mean values for the TMDs of the hypothetical proteins ranged from 31.7-44.1%, which suggests that if the putative T. cruzi PAF receptor is among the sequences identified, the TMDs are not highly conserved. These results suggest that T. cruzi contains several GPCR-like proteins and that one of these GPCRs may be a PAF receptor. Future studies may further validate the PAF receptor as a target for CD chemotherapy.

Evolution of insecticide resistance in non-target black flies (Diptera: Simuliidae) from Argentina

Black flies, a non-target species of the insecticides used in fruit production, represent a severe medical and veterinary problem. Large increases in the level of resistance to the pyrethroids fenvalerate (more than 355-fold) and deltamethrin (162-fold) and a small increase in resistance to the organophosphate azinphos methyl (2-fold) were observed between 1996-2008 in black fly larvae under insecticide pressure. Eventually, no change or a slight variation in insecticide resistance was followed by a subsequent increase in resistance. The evolution of pesticide resistance in a field population is a complex and stepwise process that is influenced by several factors, the most significant of which is the insecticide selection pressure, such as the dose and frequency of application. The variation in insecticide susceptibility within a black fly population in the productive area may be related to changes in fruit-pest control. The frequency of individuals with esterase activities higher than the maximum value determined in the susceptible population increased consistently over the sampling period. However, the insecticide resistance was not attributed to glutathione S-transferase activity. In conclusion, esterase activity in black flies from the productive area is one mechanism underlying the high levels of resistance to pyrethroids, which have been recently used infrequently. These enzymes may be reselected by currently used pesticides and enhance the resistance to these insecticides.

The zinc finger protein TcZFP2 binds target mRNAs enriched during Trypanosoma cruzi metacyclogenesis

Trypanosomes are parasitic protozoa in which gene expression is primarily controlled through the regulation of mRNA stability and translation. This post-transcriptional control is mediated by various families of RNA-binding proteins, including those with zinc finger CCCH motifs. CCCH zinc finger proteins have been shown to be essential to differentiation events in trypanosomatid parasites. Here, we functionally characterise TcZFP2 as a predicted post-transcriptional regulator of differentiation in Trypanosoma cruzi. This protein was detected in cell culture-derived amastigotes and trypomastigotes, but it was present in smaller amounts in metacyclic trypomastigote forms of T. cruzi. We use an optimised recombinant RNA immunopreciptation followed by microarray analysis assay to identify TcZFP2 target mRNAs. We further demonstrate that TcZFP2 binds an A-rich sequence in which the adenosine residue repeats are essential for high-affinity recognition. An analysis of the expression profiles of the genes encoding the TcZFP2-associated mRNAs throughout the parasite life cycle by microarray hybridisation showed that most of the associated mRNAs were upregulated in the metacyclic trypomastigote forms, also suggesting a role for TcZFP2 in metacyclic trypomastigote differentiation. Knockdown of the orthologous Trypanosoma brucei protein levels showed ZFP2 to be a positive regulator of specific target mRNA abundance.

The mazEF toxin-antitoxin system as a novel antibacterial target in Acinetobacter baumannii

Although analysis of toxin-antitoxin (TA) systems can be instructive, to date, there is no information on the prevalence and identity of TA systems based on a large panel of Acinetobacter baumannii clinical isolates. The aim of the current study was to screen for functional TA systems among clinical isolates of A. baumannii and to identify the systems’ locations. For this purpose, we screened 85 A. baumannii isolates collected from different clinical sources for the presence of the mazEF, relBE and higBA TA genes. The results revealed that the genes coding for the mazEF TA system were commonly present in all clinical isolates of A. baumannii. Reverse transcriptase-polymerase chain reaction analysis showed that transcripts were produced in the clinical isolates. Our findings showed that TA genes are prevalent, harboured by chromosomes and transcribed within A. baumannii. Hence, activation of the toxin proteins in the mazEF TA system should be investigated further as an effective antibacterial strategy against this bacterium.

Vascular dysfunction as a target for adjuvant therapy in cerebral malaria

Cerebral malaria (CM) is a life-threatening complication of Plasmodium falciparum malaria that continues to be a major global health problem. Brain vascular dysfunction is a main factor underlying the pathogenesis of CM and can be a target for the development of adjuvant therapies for the disease. Vascular occlusion by parasitised red blood cells and vasoconstriction/vascular dysfunction results in impaired cerebral blood flow, ischaemia, hypoxia, acidosis and death. In this review, we discuss the mechanisms of vascular dysfunction in CM and the roles of low nitric oxide bioavailability, high levels of endothelin-1 and dysfunction of the angiopoietin-Tie2 axis. We also discuss the usefulness and relevance of the murine experimental model of CM by Plasmodium berghei ANKA to identify mechanisms of disease and to screen potential therapeutic interventions.

The Duffy binding protein as a key target for a Plasmodium vivax vaccine: lessons from the Brazilian Amazon

Plasmodium vivax infects human erythrocytes through a major pathway that requires interaction between an apical parasite protein, the Duffy binding protein (PvDBP) and its receptor on reticulocytes, the Duffy antigen/receptor for chemokines (DARC). The importance of the interaction between PvDBP (region II, DBPII) and DARC to P. vivax infection has motivated our malaria research group at Oswaldo Cruz Foundation (state of Minas Gerais, Brazil) to conduct a number of immunoepidemiological studies to characterise the naturally acquired immunity to PvDBP in populations living in the Amazon rainforest. In this review, we provide an update on the immunology and molecular epidemiology of PvDBP in the Brazilian Amazon - an area of markedly unstable malaria transmission - and compare it with data from other parts of Latin America, as well as Asia and Oceania.

"I came from there (but no one warned me!)": reflections on people management policies that target intranational interculturality

The purpose of this article is to analyze how professionals who decided to risk intramobility in their careers and who were hired by organizations from the industrial complex of Camaçari (Bahia) perceive the development of people management strategies that target intranational interculturality within these organizations. To achieve this, besides a review and theoretical analysis of the concepts of mobility, intercultural management and people management in organizations, 13 professionals were interviewed who had moved from different Brazilian states to work in companies in this particular complex. The results indicate that generally organizations ignore intercultural aspects, which is reflected in a gap in people management strategies. One indication of this refers to the lack of initial support, which generates problems for the individuals who arrive with great expectations in their new workplace. Furthermore, there is evidence that the financial help provided is insufficient and it becomes necessary to pay attention also to subjective aspects that involve relocation and the consequent international interculturality. Finally, it is believed that expansion of the focus of studies on intercultural management, with a look at intranational aspects, makes it possible to learn theoretical and practical lessons because of the experiences of the players who underwent the process, since intercultural management and people management can generate comparative advantages for organizations.

Modes of acquisition of health literacy skills in informal learning contexts

In this article we try to analyze the learning processes of health literacy skills in informal contexts. We intend to broaden the understanding of the learning process beyond the formal contexts, thus contributing to the elucidation of health professionals on how individuals acquire and manage their knowledge in health matters. Given our goal, we use an analytic corpus constituted by one hundred autobiographical narratives written between 2006 and 2011, in educational contexts but with recognized potential for use in different scientific fields, including health. The results reveal the existence of three different types of modes of learning health literacy skills in informal context: : i) learning that takes place in action, in achieving daily tasks; ii) learning processes that result from problem solving; iii) learning that occurs in an unplanned manner, resulting from accidental circumstances and, in some cases, devoid of intentionality.

Does Bt maize cultivation affect the non-target insect community in the agro ecosystem?

ABSTRACT The cultivation of genetically modified crops in Brazil has led to the need to assess the impacts of this technology on non-target species. Under field conditions, the potential effect on insect biodiversity was evaluated by comparing a homogeneous corn field with conventional and transgenic maize, expressing different Bt proteins in seven counties of Minas Gerais, Brazil. The richness pattern of non-target insect species, secondary pests and natural enemies were observed. The results do not support the hypothesis that Bt protein affects insect biodiversity. The richness and diversity data of insects studied were dependent on the location and other factors, such as the use of insecticides, which may be a major factor where they are used.

Seeds with high molybdenum concentration improved growth and nitrogen acquisition of rhizobium-inoculated and nitrogen-fertilized common bean plants

Seeds of common bean (Phaseolus vulgaris) with high molybdenum (Mo) concentration can supply Mo plant demands, but to date no studies have concomitantly evaluated the effects of Mo-enriched seeds on plants inoculated with rhizobia or treated with N fertilizer. This work evaluated the effects of seed Mo on growth and N acquisition of bean plants fertilized either by symbiotic N or mineral N, by measuring the activities of nitrogenase and nitrate reductase and the contribution of biological N2 fixation at different growth stages. Seeds enriched or not with Mo were sown with two N sources (inoculated with rhizobia or fertilized with N), in pots with 10 kg of soil. In experiment 1, an additional treatment consisted of Mo-enriched seeds with Mo applied to the soil. In experiment 2, the contribution of N2 fixation was estimated by 15N isotope dilution. Common bean plants grown from seeds with high Mo concentration flowered one day earlier. Seeds with high Mo concentration increased the leaf area, shoot mass and N accumulation, with both N sources. The absence of effects of Mo application to the soil indicated that Mo contents of Mo-enriched seeds were sufficient for plant growth. Seeds enriched with Mo increased nitrogenase activity at the vegetative stage of inoculated plants, and nitrate reductase activity at late growth stages with both N sources. The contribution of N2 fixation was 17 and 61 % in plants originating from low- or high-Mo seeds, respectively. The results demonstrate the benefits of sowing Mo-enriched seeds on growth and N nutrition of bean plants inoculated with rhizobia or fertilized with mineral N fertilizer.

Phosphorus acquisition and internal utilization efficiency in tropical maize genotypes

The objective of this work was to determine the relative importance of phosphorus acquisition efficiency (PAE - plant P uptake per soil available P), and phosphorus internal utilization efficiency (PUTIL - grain yield per P uptake) in the P use efficiency (PUE - grain yield per soil available P), on 28 tropical maize genotypes evaluated at three low P and two high P environments. PAE was almost two times more important than PUTIL to explain the variability observed in PUE, at low P environments, and three times more important at high P environments. These results indicate that maize breeding programs, to increase PUE in these environments, should use selection index with higher weights for PAE than for PUTIL. The correlation between these two traits showed no significance at low or at high P environments, which indicates that selection in one of these traits would not affect the other. The main component of PUTIL was P quotient of utilization (grain yield per grain P) and not the P harvest index (grain P per P uptake). Selection to reduce grain P concentration should increase the quotient of utilization and consequently increase PUTIL.