Characterization of proteinases from the midgut of Rhipicephalus (Boophilus) microplus involved in the generation of antimicrobial peptides

Background: Hemoglobin is a rich source of biologically active peptides, some of which are potent antimicrobials (hemocidins). A few hemocidins have been purified from the midgut contents of ticks. Nonetheless, how antimicrobials are generated in the tick midgut and their role in immunity is still poorly understood. Here we report, for the first time, the contribution of two midgut proteinases to the generation of hemocidins. Results: An aspartic proteinase, designated BmAP, was isolated from the midgut of Rhipicephalus (Boophilus) microplus using three chromatographic steps. Reverse transcription-quantitative polymerase chain reaction revealed that BmAP is restricted to the midgut. The other enzyme is a previously characterized midgut cathepsin L-like cysteine proteinase designated BmCL1. Substrate specificities of native BmAP and recombinant BmCL1 were mapped using a synthetic combinatorial peptide library and bovine hemoglobin. BmCL1 preferred substrates containing non-polar residues at P2 subsite and polar residues at P1, whereas BmAP hydrolysed substrates containing non-polar amino acids at P1 and P1`. Conclusions: BmAP and BmCL1 generate hemocidins from hemoglobin alpha and beta chains in vitro. We postulate that hemocidins may be important for the control of tick pathogens and midgut flora.

TICK INNATE IMMUNITY

Ticks are blood feeding parasites transmitting a wide variety of pathogens to their vertebrate hosts. The vector competence of ticks is tightly linked with their immune system. Despite its importance, our knowledge of tick innate immunity is still inadequate and the limited number of sufficiently characterized immune molecules and cellular reactions are dispersed across numerous tick species. The phagocytosis of microbes by tick hemocytes seems to be coupled with a primitive complement-like system, which possibly involves self/nonself recognition by fibrinogen-related lectins and the action of thioester-containing proteins. Ticks do not seem to possess a pro-phenoloxidase system leading to melanization and also coagulation of tick hemolymph has not been experimentally proven. They are capable of defending themselves against microbial infection with a variety of antimicrobial peptides comprising lysozymes, defensins and molecules not found in other invertebrates. Virtually nothing is known about the signaling cascades involved in the regulation of tick antimicrobial immune responses. Midgut immunity is apparently the decisive factor of tick vector competence. The gut content is a hostile environment for ingested microbes, which is mainly due to the antimicrobial activity of hemoglobin fragments generated by the digestion of the host blood as well as other antimicrobial peptides. Reactive oxygen species possibly also play an important role in the tick-pathogen interaction. The recent release of the Ixodes scapularis genome and the feasibility of RNA interference in ticks promise imminent and substantial progress in tick innate immunity research.

Exogenous insulin stimulates glycogen accumulation in Rhipicephalus (Boophilus) microplus embryo cell line BME26 via PI3K/AKT pathway

Ticks are obligatory blood-feeding arthropods and important vectors of both human and animal disease agents. Besides its metabolic role, insulin signaling pathway (ISP) is widely described as crucial for vertebrate and invertebrate embryogenesis, development and cell survival. In such cascade, Phosphatidylinositol 3-OH Kinase (PI3K) is hierarchically located upstream Protein Kinase B (PKB). To study the insulin-triggered pathway and its possible roles during embryogenesis we used a culture of embryonic Rhipicephalus microplus cells (BME26). Exogenous insulin elevated cell glycogen content in the absence of fetal calf serum (FCS) when compared to cells without treatment. Moreover, in the presence of PI3K inhibitors (Wortmannin or LY294002) these effects were blocked. We observed an increase in the relative expression level of PI3K`s regulatory subunit (p85), as determined by qRT-PCR. In the presence of PI3K inhibitors these effects on transcription were also reversed. Additionally, treatment with Wortmannin increased the expression level of the insulin-regulated downstream target glycogen synthase kinase 3 beta (GSK3 beta). The p85 subunit showed elevated transcription levels in ovaries from fully engorged females, but was differentially expressed during tick embryogenesis. These results strongly suggest the presence of an insulin responsive machinery in BME26 cells, and its correlation with carbohydrate/glycogen metabolism also during embryogenesis. (C) 2009 Published by Elsevier Inc.

Propagation of a Brazilian isolate of Anaplasma marginale with appendage in a tick cell line (BME26) derived from Rhipicephalus (Boophilus) microplus

Anaplasma marginale is a tick-borne pathogen of cattle responsible for the disease anaplasmosis. Data suggest that Rhipicephalus (Boophilus) microplus and R. annulatus may be the major tick vectors of A. marginale in tropical and subtropical regions of the world. In this work we demonstrated the first infection and propagation of a Brazilian isolate of A. marginale (UFMG1) in the BME26 cell line derived originally from embryos of R. (Boophilus) microplus. The establishment of A. marginale infection in a cell line derived from R. (Boophilus) microplus is relevant for studying the A. marginale/tick interface. (C) 2008 Elsevier B.V. All rights reserved.

Cathepsin L-like genes of Trypanosoma vivax from Africa and South America - characterization, relationships and diagnostic implications

We characterized sequences from genes encoding cathepsin L-like (CatL-like) cysteine proteases from African and South American isolates of Trypanosoma vivax and T. vivax-like organisms, and evaluated their suitability as genetic markers for population structure analysis and diagnosis. Phylogenetic analysis of sequences corresponding to CatL-like catalytic domains revealed substantial polymorphism, and clades of sequences (TviCatL1-9) were separated by large genetic distances. TviCatL1-4 sequences were from cattle isolates from West Africa (Nigeria and Burkina Faso) and South America (Brazil and Venezuela), which belonged to the same T. vivax genotype. T. vivax-like genotypes from East Africa showed divergent sequences, including TviCatL5-7 for isolates from Mozambique and TviCatL8-9 for an isolate from Kenya. Phylogenetic analysis of CatL-like gene data supported the relationships among trypanosome species reflected in the phylogenies based on the analysis of small subunit (SSU) of ribosomal RNA gene sequence data. The discovery of different CatL-like sequences for each genotype, defined previously by ribosomal DNA data, indicate that these sequences provide useful targets for epidemiological and population genetic studies. Regions in CatL-like sequences shared by all T. vivax genotypes but not by other trypanosomes allowed the establishment of a specific and sensitive diagnostic PCR for epidemiological studies in South America and Africa. (C) 2008 Elsevier Ltd. All rights reserved.

The combined effect of copper and low pH on antioxidant defenses and biochemical parameters in neotropical fish pacu, Piaractus mesopotamicus (Holmberg, 1887)

Copper sulfate is widely used in aquaculture. Exposure to this compound can be harmful to fish, resulting in oxidative metabolism alterations and gill tissue damage. Pacu, Piaractus mesopotamicus, (wt = 43.4 +/- A 3.35 g) were distributed in experimental tanks (n = 10; 180 l) and exposed for 48 h to control (without copper addition), 0.4Cu (0.4 mg l(-1)), 0CupH (without copper addition, pH = 5.0) and 0.4CupH (0.4 mg l(-1), pH = 5.0). In liver and red muscle, the superoxide dismutase (SOD) was responsive to the increases in the aquatic copper. The plasmatic intermediary metabolites and hematological variables in the fish of group 0.4Cu were similar to those of the control group. Conversely, the exposure to 0.4CupH caused an increase in the plasmatic lactate, number of red blood cells (RBC) and hemoglobin (Hb). Plasmatic copper concentration [Cu(p)] increased in group 0.4Cu and 0.4CupH, which is higher in group 0.4CupH, suggests an effect of water pH on the absorbed copper. Exposure to 0.4Cu and 0.4CupH resulted in a reduction in the Na(+)/K(+)-ATPase activity and an increase in metallothionein (MT) in the gills. Exposure to 0CupH caused a decrease in glucose and pyruvate concentrations and an increase in RBC, Hb, and the branchial Na(+)/K(+)-ATPase activity. These responses suggest that the fish triggered mechanisms to revert the blood acidosis, save energy and increase the oxygen uptake. MT was an effective biomarker, responding to copper in different pHs and dissolved oxygen. Combined-factors caused more significant disturbance in the biomarkers than single-factors.

Selection of an ivermectin-resistant strain of Rhipicephalus microplus (Acari: Ixodidae) in Brazil

Resistance to ivermectin (IVM) in field Populations of Rhipicephalus microplus of Brazil has been observed since 2001 In this work, four selection methods (infestations with: (I) IVM-treated larvae, (2) larvae from IVM-treated adult female ticks, (3) larvae from IVM-treated adult female ticks on an IVM-treated host, and (4) larvae obtained from W-treated females that produced eggs with a high eclosion rate) were used oil a field population with an initial ivermectin (IVM) resistance ratio at LC50 (RR50) of 1 37 with the objective to obtain experimentally a highly-resistant strain After ten generations, using these methods combined, the final RR50 was 8 06 This work shows for the first time that it was possible to increase IVM resistance in R. microplus in laboratory conditions. The establishment of a drug resistant R microplus strain is a fundamental first step for further research into the mechanisms of ivermectin-resistance in R. microplus and potentially methods to control this resistance (C) 2009 Elsevier B V All rights reserved

Rhipicephalus (Boophilus) microplus (Acari: Ixodidae) resistance to fipronil in Uruguay evaluated by in vitro bioassays

Rhipicephalus (Boophilus) microplus obtained from four local populations in Uruguay (2007-2008) were subjected to various bioassay techniques to determine the presence of fipronil resistance within the country. Resistance ratios (RRs) obtained by larval immersion test varied between 3.3 and 3635 for tick populations subjected to treatment with fipronil for the last 3-7 years. The highest RR was observed in the population which received fewer treatments. Using discriminating concentration (8 ppm) for larval immersion test, all field strains were correctly diagnosed as fipronil-resistant. This study presents the first diagnoses of cattle tick resistance to fipronil in Uruguay`s field populations. It also highlights the importance of the possible conflict between programs to control agricultural pests and cattle ticks. The findings provide valuable information for selection and adoption of new control alternatives to manage drug resistance exhibited by cattle ticks. (C) 2009 Elsevier BM. All rights reserved.

Variação, contato e mudança linguística em Moçambique e Cabo Verde : A concordância variável de número em sintagmas nominais do português

This study investigates variable noun phrase number agreement (VNA) in two second language varieties of Portuguese, spoken in Maputo, Mozambique and in Mindelo, Cape Verde. Quantitative VARBRUL analysis is carried out based on recordings made in Maputo and Mindelo 2007 and 2008. Previous quantitative studies on VNA in varieties of Brazilian Portuguese (Guy, 1981; Lopes, 2001; Andrade, 2003) as well as on VNA in first and second language varieties of Portuguese from São Tomé (Baxter, 2004; Figueiredo, 2008, 2010) indicate contact between Portuguese and African languages as the main origin of this phenomenon. VNA in Brazilian Portuguese is, however, interpreted by Scherre (1988) and Naro & Scherre (1993, 2007) as the result of language internal drift. Varieties of Portuguese from Mozambique and Cape Verde are particularly interesting to contrast in order to investigate influences from African languages on VNA, as in Mozambique Bantu languages are first languages of the vast majority of Portuguese speakers, whereas in Cape Verde, practically all Portuguese speakers are first language speakers of Cape Verdean Creole, whose substrates are West African, and not Bantu, languages. Comparison is also made with previous studies from Brazil and São Tomé. The results of this study comment previously postulated explanations for VNA in Portuguese in various ways. The analysis of the variables onset age and age stratum indicates that VNA in the analyzed varieties is a phenomenon linked to the acquisition of Portuguese as a second language and/or language contact rather than the result of internal drift. The fact that all the compared varieties tend to mark plural on pre-head components contradicts Bantu transfer as an explanation for this pattern, and raises the need to also consider more general explanations based on language contact. The basic structural similarity between the compared varieties suggests the existence of a grammatical restructuring continuum.

Kangaroo Mother Care in a Mozambican perinatal ward : a clinical case study

Kangaroo mother care (KMC) was first introduced in Mozambique in 1984. The aim of this study was to describe Mozambican mothers’ experiences of going through admission, passing from an intensive care ward to a nursery ward with their premature baby, undergoing KMC training before early discharge. A clinical case study was conducted, involving naturalistic observations and a face-to-face interview with 41 mothers participating to complete a questionnaire. Descriptive statistics and manifest content analysis were used in this study. The results show that the mothers were of low socio-economic standing and felt that they did not have enough information on KMC. The hierarchical organization within the hospital setting as well as communalistic behaviours influenced the mothers’ support of KMC, including information, communication, relationships and actions. The conclusion is that there is an important challenge for trained neonatal nurses to improve the guidelines for KMC and to empower mothers and their families to adopt KMC.

Projeto para criação do Centro de Pesquisa e Documentação de História Oral e Ciências Sociais (Beira, Moçambique)

A presente dissertação tem como objetivo central propor a criação do Centro de Pesquisa e Documentação de História Oral e Ciencias Sociais na cidade da Beira, em Moçambique. Pretende-se, com o Centro, promover projetos que possibilitem a constituição de acervos de caráter histórico e documental, contribuindo, assim, para o desenvolvimento da pesquisa acadêmica e para a valorização da história e da cultura Moçambicanas. O Centro terá como principal ferramenta metodológica de pesquisa a história oral. O acervo a ser constituído no contexto de distintos projetos resultará, portanto, de entrevistas gravadas com atores de diferentes extratos sociais, todos eles participantes diretos de eventos relevantes da história recente de Moçambique. Essas entrevistas deverão estar disponíveis, no Centro, juntamente com outros tipos de fontes, coletadas e organizadas com o objetivo de constituir um centro de referência sobre a história contemporanea do país. Toda a atividade do Centro será acompanhada de projetos de qualificação de equipes. Neste sentido, o Centro terá de lidar com dois importantes campos: a pesquisa, produção, preservação e difusão de fontes para a história contemporânea; e o apoio didático-pedagógico aos diversos níveis do processo de aprendizagem.

O papel das relações interpessoais na condução do ProSavana-JBM (Japão-Brasil-Moçambique): um estudo de caso: programa de Cooperação Triangular para o Desenvolvimento Agrícola da Savana Tropical de Moçambique

O objetivo deste estudo de caso foi o de identificar o papel das relações interpessoais na condução do ProSAVANA-JBM (JAPÃO-BRASIL-MOÇAMBIQUE), Programa de Cooperação Triangular para o Desenvolvimento Agrícola da Savana Tropical de Moçambique. Os objetivos intermediários permitem também investigar quais outros fatores podem ter sido determinantes na condução do ProSAVANA. Este programa além de representar uma parceria internacional entre organizações de três países Brasil, Japão e Moçambique, possui impactos de ordem social, econômica e ambiental. A análise dos dados demonstra que as relações interpessoais exerceram a maior centralidade entre os demais fatores determinantes à condução do Programa, e também corrobora para o desenho de um modelo de rede de relações interpessoais relacionada ao ProSAVANA. As limitações do estudo referem-se à dificuldade deste estudo ser replicado ou generalizado cientificamente, considerando que não seria possível emitir avaliações sobre determinado comportamento, a partir de um estudo de caso único. Finalmente, o propósito deste estudo é oferecer instrumentos teóricos e empíricos que permitam aprofundar a reflexão sobre o papel das relações interpessoais, além de contribuir para as lacunas existentes no campo da gestão empresarial e pública.

A cooperação trilateral brasileira em Moçambique: um estudo de caso comparado: o ProALIMENTOS e o ProSAVANA

A arquitetura do sistema de ajuda internacional passa por um processo de transformação, no qual as barreiras Norte e Sul tornam-se cada vez mais indefinidas, e a cooperação trilateral, que une as práticas opostas da cooperação Norte-Sul com a cooperação Sul-Sul, emerge como uma nova modalidade da cooperação técnica brasileira. Com o objetivo de compreender esse cenário, esta tese almeja identificar e contrastar as motivações e as práticas dos agentes da cooperação trilateral brasileira por meio de um estudo de caso comparado de dois projetos, desenvolvidos pela Agência Brasileira de Cooperação (ABC), no setor agrícola em Moçambique: o ProALIMENTOS, parceria entre a United States Agency for International Development (USAID- -Brasil) e ABC, e o ProSAVANA, parceria entre Japan International Cooperation Agency (JICA) e ABC. Para isso, a pesquisa parte dos pressupostos da actor-oriented approach para estabelecer uma análise multinível, que cria desta forma um elo entre o agente e a estrutura, a prática e a política, com um olhar sobre o contexto macro, meso e micro. A utilização da metodologia qualitativa aplicada a essa investigação combinou a técnica de participação observante com a técnica de análise documental, acrescentando à análise 59 entrevistas semiestruturadas, realizadas principalmente entre os meses de março e junho de 2013, em Moçambique. Os resultados da pesquisa indicam que, no caso do ProALIMENTOS, há ganhos em complementaridade e troca de conhecimento para as três contrapartes, porém há a sobreposição das práticas Norte-Sul de cooperação para o desenvolvimento internacional sobre as práticas da cooperação Sul-Sul. Enquanto que, no caso do ProSAVANA, não há qualquer ganho de complementariedade, uma vez que o Programa enfrenta desafios internos e externos. A falta de harmonização e coordenação técnica no âmbito interno intensificam o embate externo com os representantes da sociedade civil ao gerar constantes falhas de comunicação, o que coloca em xeque a própria continuidade do ProSAVANA. Por último, a pesquisa mostra que é necessário um maior comprometimento do governo brasileiro nos projetos de Cooperação Trilateral, uma vez que os resultados desses projetos podem impactar e afetar a credibilidade do Brasil como um novo prestador de ajuda internacional.

O Estado e a construção da ordem arquivística em Moçambique (1975-2010)

Esta tese tem como objetivo compreender o Arquivo Histórico de Moçambique (AHM) como lugar de informação arquivística e de ação do Estado em Moçambique, analisando o processo histórico de sua configuração, tendo em conta as implicações desse processo no cenário arquivístico nacional e relação com o projeto pós-colonial de nação, particularmente entre 1975 e 2010. Com base na visão do Estado ampliado em Gramsci e na teoria do Estado como relação em Poulantzas a tese mapeia a dimensão teórica do Estado, cujo poder baseia-se em informação. Esta abordagem, baseada na concepção teórica do Estado como campo de informação, consolida um quadro conceitual fundamental para o entendimento do Estado moçambicano e seu processo histórico de construção. A mesma abordagem conduziu o estudo de caráter histórico na análise dos processos de constituição e disponibilização de arquivos públicos, constituídos no quadro da configuração do AHM dentro do processo histórico de construção do Estado neste país em suas várias redes de interações, envolvendo diversos atores sociais, seja no universo político-administrativo ou arquivístico em si. Constatou-se uma mudança na trajetória do AHM, redefinindo o processo histórico de construção da memória e da identidade nacional através dos arquivos, dentro do processo de construção de uma ordem arquivística politicamente aceite, iniciado em 1975 e consolidado nos anos 2000.

Brazil & Lusphone Africa: a study of history, international relations, & international trade

This report was inspired by a personal motivation to acquire more in depth knowledge about Brazil and Lusophone (Portuguese speaking) African nations and how they interact with each other in relation to their common colonial histories, cultures, and on matters of international relations, international development, and international trade. The countries selected for purpose and focus of this report are Brazil, Angola, and Mozambique; reference will also be made with respect to other Lusophone African countries such as Cabo Verde, Guinea-Bissau, and São Tomé e Príncipe. Some of the research methodologies used to gather information about Brazil, Angola, Mozambique, and other Lusophone African nations in relation to their respective histories, international relations, international trade relations, and roles in the global economy as emerging market nations.