Interaction of an opportunistic fungus Purpureocillium lilacinum with human macrophages and dendritic cells.

INTRODUCTION: Purpureocillium lilacinum is emerging as a causal agent of hyalohyphomycosis that is refractory to antifungal drugs; however, the pathogenic mechanisms underlying P. lilacinum infection are not understood. In this study, we investigated the interaction of P. lilacinum conidia with human macrophages and dendritic cells in vitro. METHODS: Spores of a P. lilacinum clinical isolate were obtained by chill-heat shock. Mononuclear cells were isolated from eight healthy individuals. Monocytes were separated by cold aggregation and differentiated into macrophages by incubation for 7 to 10 days at 37°C or into dendritic cells by the addition of the cytokines human granulocyte-macrophage colony stimulating factor and interleukin-4. Conidial suspension was added to the human cells at 1:1, 2:1, and 5:1 (conidia:cells) ratios for 1h, 6h, and 24h, and the infection was evaluated by Giemsa staining and light microscopy. RESULTS: After 1h interaction, P. lilacinum conidia were internalized by human cells and after 6h contact, some conidia became inflated. After 24h interaction, the conidia produced germ tubes and hyphae, leading to the disruption of macrophage and dendritic cell membranes. The infection rate analyzed after 6h incubation of P. lilacinum conidia with cells at 2:1 and 1:1 ratios was 76.5% and 25.5%, respectively, for macrophages and 54.3% and 19.5%, respectively, for cultured dendritic cells. CONCLUSIONS: P. lilacinum conidia are capable of infecting and destroying both macrophages and dendritic cells, clearly demonstrating the ability of this pathogenic fungus to invade human phagocytic cells.

Interaction of an opportunistic fungus Purpureocillium lilacinum with human macrophages and dendritic cells

Introduction Purpureocillium lilacinum is emerging as a causal agent of hyalohyphomycosis that is refractory to antifungal drugs; however, the pathogenic mechanisms underlying P. lilacinum infection are not understood. In this study, we investigated the interaction of P. lilacinum conidia with human macrophages and dendritic cells in vitro. Methods Spores of a P. lilacinum clinical isolate were obtained by chill-heat shock. Mononuclear cells were isolated from eight healthy individuals. Monocytes were separated by cold aggregation and differentiated into macrophages by incubation for 7 to 10 days at 37°C or into dendritic cells by the addition of the cytokines human granulocyte-macrophage colony stimulating factor and interleukin-4. Conidial suspension was added to the human cells at 1:1, 2:1, and 5:1 (conidia:cells) ratios for 1h, 6h, and 24h, and the infection was evaluated by Giemsa staining and light microscopy. Results After 1h interaction, P. lilacinum conidia were internalized by human cells and after 6h contact, some conidia became inflated. After 24h interaction, the conidia produced germ tubes and hyphae, leading to the disruption of macrophage and dendritic cell membranes. The infection rate analyzed after 6h incubation of P. lilacinum conidia with cells at 2:1 and 1:1 ratios was 76.5% and 25.5%, respectively, for macrophages and 54.3% and 19.5%, respectively, for cultured dendritic cells. Conclusions P. lilacinum conidia are capable of infecting and destroying both macrophages and dendritic cells, clearly demonstrating the ability of this pathogenic fungus to invade human phagocytic cells. .

Evaluation of Leishmania (Leishmania) chagasi strains isolated from dogs originating from two visceral leishmaniasis-endemic areas in Brazil using multilocus enzyme electrophoresis / Avaliação de amostras de Leishmania (Leishmania) chagasi isoladas de cães oriundos de duas áreas endêmicas de leishmaniose visceral no Brasil através da eletroforese de isoenzimas

INTRODUCTION: Domestic dogs are the most important reservoir in the peridomestic transmission cycle of Leishmania (Leishmania) chagasi. The genetic variability of subpopulations of this parasite circulating in dogs has not been thoroughly analyzed in Brazil, even though this knowledge has important implications in the clinical-epidemiological context. METHODS: The objective of this study was to evaluate and compare the phenotypic variability of 153 L. chagasi strains isolated from dogs originating from the municipalities of Rio de Janeiro (n = 57) and Belo Horizonte (n = 96), where the disease is endemic. Strains isolated only from intact skin were selected and analyzed by multilocus enzyme electrophoresis using nine enzyme systems (6PG, GPI, NH1 and NH2, G6P, PGM, MDH, ME, and IDHNADP). RESULTS: The electrophoretic profile was identical for all isolates analyzed and was the same as that of the L. chagasi reference strain (MHOM/BR/74/PP75). Phenetic analysis showed a similarity index of one for all strains, with the isolates sharing 100 percent of the characteristics analyzed. CONCLUSIONS: The results demonstrate that the L. chagasi populations circulating in dogs from Rio de Janeiro and Belo Horizonte belong to a single zymodeme.

INTRODUÇÃO: Cães domésticos são considerados os reservatórios mais importantes no ciclo peridoméstico de transmissão de Leishmania (Leishmania) chagasi. No entanto, a variabilidade genética de sub-populações que circulam neste hospedeiro é ainda pouco explorada no Brasil, sendo tal conhecimento de grande importância no contexto clínico-epidemiológico. MÉTODOS: O objetivo deste estudo foi avaliar e comparar a variabilidade fenotípica de 153 amostras de L. chagasi isoladas de cães oriundos dos municípios do Rio de Janeiro (n = 57) e Belo Horizonte (n = 96), onde a doença é endêmica. Foram selecionadas somente amostras isoladas de pele íntegra e analisadas por eletroforese de isoenzimas (MLEE) empregando nove sistemas enzimáticos (6PG, GPI, NH1 e NH2, G6P, PGM, MDH, ME, IDHNADP). RESULTADOS: Todas as amostras analisadas apresentaram perfil eletroforético idêntico entre si e com a amostra de L. chagasi utilizada como referência neste estudo (MHOM/BR/74/PP75). A análise fenética demonstrou índice de similaridade igual a um para todas as amostras, revelando um compartilhamento de 100 por cento dos caracteres avaliados. CONCLUSÕES: A partir desses resultados, podemos inferir que as populações de L. chagasi que estão circulando nos cães do Rio de Janeiro e Belo Horizonte podem ser agrupadas em um único zimodema.

Evaluation of Leishmania (Leishmania) chagasi strains isolated from dogs originating from two visceral leishmaniasis-endemic areas in Brazil using multilocus enzyme electrophoresis.

INTRODUCTION: Domestic dogs are the most important reservoir in the peridomestic transmission cycle of Leishmania (Leishmania) chagasi. The genetic variability of subpopulations of this parasite circulating in dogs has not been thoroughly analyzed in Brazil, even though this knowledge has important implications in the clinical-epidemiological context. METHODS: The objective of this study was to evaluate and compare the phenotypic variability of 153 L. chagasi strains isolated from dogs originating from the municipalities of Rio de Janeiro (n = 57) and Belo Horizonte (n = 96), where the disease is endemic. Strains isolated only from intact skin were selected and analyzed by multilocus enzyme electrophoresis using nine enzyme systems (6PG, GPI, NH1 and NH2, G6P, PGM, MDH, ME, and IDHNADP). RESULTS: The electrophoretic profile was identical for all isolates analyzed and was the same as that of the L. chagasi reference strain (MHOM/BR/74/PP75). Phenetic analysis showed a similarity index of one for all strains, with the isolates sharing 100% of the characteristics analyzed. CONCLUSIONS: The results demonstrate that the L. chagasi populations circulating in dogs from Rio de Janeiro and Belo Horizonte belong to a single zymodeme.

Antiophidian sera sterility control: topics in perspective / Controle de esterilidade de soros antiofídicos: temas em perspectiva

The aim of this work is to review the most important topics about the antiophidic sera sterility, including obtaining methods, sterilization procedures and clean room control using Vital Brazil Institute (VBI) as an example. Bibliographical research was performed through Medline, Lilacs, PubMed, ISI and the Fundação Oswaldo Cruz - RJ and VBI Libraries, from 1960 to 2009. The antiophidic sera for human use are immunobiologic products produced in Brazil by three national laboratories, including VBI. Due to the parenteral use, these products should be sterile and pyrogen-free, which demands the microbiological control during the whole fabrication process. The sterility and pyrogen tests are important steps to ensure the quality and safety of these immunobiological products. Thus, these tests are target for continue evaluation and improvement. The most interfering aspects in the consistency and analytical patterns include the proper method selection, sampling, culture conditions and validation criteria. As the national and international legal requirements are cautious with the assays validation and approval of sterile parenteral products; the intrinsic limitations for established assays still require more investigation aiming the continue improvement of the microorganism and contaminants detection methods and optimization of the analysis extent.

O objetivo deste trabalho é revisar os tópicos mais relevantes para o controle da esterilidade de soros antiofídicos, abordando-se métodos de obtenção, procedimentos de esterilização e o controle de áreas limpas utilizando como exemplo os procedimentos adotados pelo Instituto Vital Brazil (IVB). Um levantamento bibliográfico foi realizado no Medline, ISI, Biblioteca da Fundação Oswaldo Cruz-RJ e IVB, no período de 1960 a 2009. Os soros antiofídicos para uso humano são produtos imunobiológicos fabricados no Brasil por três laboratórios nacionais, dentre eles o IVB. Por serem de administração parenteral, devem ser obrigatoriamente estéreis e apirogênicos, exigindo controle microbiológico durante todo o processo de fabricação. O teste de esterilidade e apirogenia são importantes instrumentos para garantir a qualidade e segurança microbiológica desses produtos, sendo alvo de avaliações constantes para seus aprimoramentos. Os aspectos que mais interferem na sua consistência e valor analítico incluem a escolha adequada do método, amostragem, condições de cultivo e critérios de validação. À medida que os requisitos legais nacionais e internacionais se mostram rigorosos na validação de ensaios e aprovação de produtos estéreis parenterais, limitações intrínsecas ao ensaio padronizado requisitam mais investigações, objetivando o aprimoramento contínuo nos métodos de detecção de microorganisms, contaminantes e otimização do tempo total de análise.

Staphylococcus aureus: visitando uma cepa de importância hospitalar / Staphylococcus aureus: visiting a strain of clinical importance

O Staphylococcus aureus é uma bactéria do grupo dos cocos gram-positivos que faz parte da microbiota humana, mas que pode provocar doenças que vão desde uma infecção simples, como espinhas e furúnculos, até as mais graves, como pneumonia, meningite, endocardite, síndrome do choque tóxico e septicemia, entre outras. Essa bactéria foi uma das primeiras a serem controladas com a descoberta dos antibióticos, mas, devido a sua enorme capacidade de adaptação e resistência, tornou-se uma das espécies de maior importância no quadro das infecções hospitalares e comunitárias. Neste artigo faremos uma revisão sobre esse agente infeccioso e as bases dos mecanismos das patologias por ele provocadas, de forma a ressaltar a necessidade de mantê-lo como alvo para o desenho de novos antibióticos.

Staphylococcus aureus is a bacterium from Gram-positive coccus group, which is part of the human microbiota. It may cause diseases that may vary from simple infections (i.e., pimples and furuncles) to severe infections, such as pneumonia, meningitis, endocarditis, toxic shock syndrome and septicemia, among others. This bacterium was one of the first bacteria affected by antibacterial agents, however, its ability of adaptation and resistance turns it into an important hospital and communitarian pathogenic species of great concern. In this article we will discuss some important points related to the S. aureus and the pathologies related to it to reinforce it as a target for the design of new antibiotics.

Plaquetas: ainda um alvo terapêutico / Platelets: still a therapeutical target

As plaquetas são fragmentos citoplasmáticos anucleados presentes no sangue e produzidos na medula óssea a partir dos megacariócitos. O objetivo deste trabalho é rever as bases mecanísticas e moleculares das plaquetas, revelando sua participação em síndromes importantes e na trombose arterial, além de seu potencial como alvo terapêutico para o desenho de novos agentes antitrombóticos.

Platelets are cytoplasmatic fragments from bone marrow megakaryocytes present on blood. The aim of this work is to review the basis of platelet mechanisms, their participation in syndromes and in arterial thrombosis, and its potential as a target for design of antithrombotic agents.

Tourette: por dentro da síndrome / Tourette: within the syndrome

A síndrome de Tourette (ST) é uma patologia de comprometimento psicossocial que acarreta alterações significativas na vida dos seus portadores e respectivos familiares. Este artigo aborda diversos aspectos relacionados a esta doença, incluindo etiologia, epidemiologia, aspectos neurobiológicos, quadro clínico, diagnóstico, patologias associadas e tratamento (clássico e alternativo). Neste trabalho, ainda comparamos a ST com outras doenças, envolvendo tiques e mencionamos as associações de apoio aos pacientes portadores de ST, que auxiliam no tratamento e na socialização do paciente afetado.

Snake Venom: Any Clue for Antibiotics and CAM?

Lately several naturally occurring peptides presenting antimicrobial activity have been described in the literature. However, snake venoms, which are an enormous source of peptides, have not been fully explored for searching such molecules. The aim of this work is to review the basis of antimicrobial mechanisms revealing snake venom as a feasible source for searching an antibiotic prototype. Therefore, it includes (i) a description of the constituents of the snake venoms involved in their main biological effects during the envenomation process; (ii) examples of snake venom molecules of commercial use; (iii) mechanisms of action of known antibiotics; and (iv) how the microorganisms can be resistant to antibiotics. This review also shows that snake venoms are not totally unexplored sources for antibiotics and complementary and alternative medicine (CAM).

Role of the undergraduate student research assistant in the new millennium.

In this study, we analyze the contribution of the undergraduate student who participates in the process of generating scientific data and developing a research project using Brazilian research as an example. Historically, undergraduate students have performed the critical role of research assistants in developing countries. This aspect has been underappreciated as a means of generating scientific data in Brazilian research facilities. Brazilian educational institutions are facing major age-related generational changes among the science faculty within the next 5-10 yr. A lack of adequate support for graduate students leads to a concern that undergraduates will not be interested in choosing research assistant programs and, subsequently, academic research careers. To remedy this situation it is important to focus on ways to encourage new research careers and enhance university-industry collaborations.

Solving an ethical issue involved in experimentation with animals in a brazilian teaching laboratory*.

Changes are occurring within Brazilian institutes of higher education; currently several universities are reviewing their course offerings and teaching approaches to determine if they meet the needs of today's undergraduate students. When changes are made to the curriculum of experimental courses, there should be an understood guarantee that all efforts to avoid ethical and biosafety issues have been diligently considered. Ethical considerations lead us to create an alternative experimental session to be conducted that eliminated the use of rats, the conventional in vivo model employed for learning metabolism of glycogen in our university. To avoid possible biosafety issues, we prepared an alternative sample to simulate human urine, which we called guarurine. Using our new method, it is possible to verify positive results imitating a diabetic and starving people samples for detection of glucose and ketone. The alternative tool described herein is not only particularly suited to bypass the ethics of using animals for teaching, but also permits the discussion of significant aspects of pathological and physiological situations such as diabetics and starvation in a simple, safe, and interesting way.

Atividades funcionais de monócitos na lepra / ?

Este trabalho consiste de estudos de ativação de monócitos na lepra, em resposta a diferentes estímulos. Para investigar a presença ou não desta ativação, utilizamos parâmetros distintos tais como: detecção de quimioluminescência (CL); atividade pro-coagulante (PCA); produção de tumor necrosis factor (TNF); apresentação antigênica e linpoproliferação. Investigamos ainda, se a ativação de monócitos isolados de pacientes lepromatosos (LL) se expressava do mesmo modo nestes pacientes, em diferentes condições clínicas (pacientes não tratados; pacientes em tratamento; pacientes em episódios reacionais - eritema nodosum leprosum (ENL) e reação reversa (RR); bem como no estado pós reacional). Em resumo, nossos resultados mostraram que: 1. Em relação a produção de radicais livres, somente monócitos isolados e pacientes LL apresentando episódios reacionais (ENL e RR) respondem ao estímulo com forbol-miristato-acetato (PMA) em maior grau de monócitos isolados de indivíduos normais. E interessante ainda notar que, monócitos isolados destes pacientes também apresentam maior atividade basal de resposta quimioluminescente. 2. Mycobacterium leprae (M. leprae) é incapaz de induzir respostas quimioluminescente em monócitos isolados de todos os grupos estudados e, suprime a resposta ao PMA. 3. A supressão de resposta quimioluminescente ao PMA induzida pelo M. leprae, não é devido, provalvelmente, a componentes estruturais da classe peptidoglicano presente em micobactérias, desde que muramil-dipeptidio (MDP), um análogo sistético ao peptidoglicano, não altera a resposta quimioluminescente ao PMA.