Análise da expressão heteróloga de uma serino protease (RnTrypsin) e de um inibidor de serino protease (RnKazalPI) salivares de Rhodnius neglectus, vetor da Doença de Chagas

Dissertação (mestrado)—Universidade de Brasília, Faculdade de Medicina, Programa de Pós-Graduação em Ciências Médicas, 2018.O triatomíneo Rhodnius neglectus (Hemiptera: Reduviidae) é um vetor potencial do protozoário Trypanosoma cruzi, agente etiológico da doença de Chagas. O R. neglectus apresenta em sua saliva moléculas importantes que neutralizam os mecanismos hemostáticos do hospedeiro, auxiliando na alimentação sanguínea e, de forma indireta, na transmissão do protozoário. Assim, a saliva desse inseto hematófago se torna um relevante alvo de pesquisas devido à presença de moléculas com elevado potencial farmacológico, como por exemplo serino proteases e inibidores de serino protease. Este estudo teve por objetivo a expressão heteróloga da serino protease do tipo tripsina (RnTrypsin) e do inibidor de serino protease tipo Kazal (RnKazalPI) salivares de R. neglectus. As sequências nucleotídicas das proteínas foram obtidas a partir do transcriptoma de R. neglectus, clonadas no vetor pET100/D-TOPO e expressas em bactérias Escherichia coli da linhagem BL21(DE3), BL21(DE3)pLysS e Rosetta(DE3)pLysS. A serino protease do tipo tripsina (RnTrypsin) recombinante foi observada somente na fração insolúvel após a lise das bactérias, sendo então purificada por cromatografia de afinidade e usada para a produção de anticorpos policlonais em camundongos. Após a otimização do protocolo de expressão, o inibidor de serino protease tipo Kazal (RnKazalPI) foi expresso na fração solúvel do lisado bacteriano. Como perspectiva, é pretendido utilizar o sistema de expressão de proteínas recombinantes em células de inseto/baculovírus para expressar a serino protease do tipo tripsina (RnTrypsin) recombinante na fração solúvel, e também determinar a caracterização do inibidor de serino protease tipo Kazal (RnKazalPI) recombinante solúvel, buscando contribuir na compreensão das interações entre vetor e hospedeiro e potenciais atividades farmacológicas.Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq); Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) e Fundação de Apoio à Pesquisa do Distrito Federal (FAP/DF).The triatomine Rhodnius neglectus (Hemiptera: Reduviidae) is a potential vector of the protozoan Trypanosoma cruzi, etiologic agent of Chagas' disease. R. neglectus presents in its saliva important molecules that neutralize the haemostatic mechanisms of the host, aiding in the blood supply and, indirectly, in the transmission of the protozoan. Thus, the saliva of this hematophagous insect becomes a relevant research target due to the presence of molecules with high pharmacological potential, for example serine proteases and serine protease inhibitors. This study aimed at the heterologous expression of the trypsin-like serine protease (RnTrypsin) and the Kazal type serine protease inhibitor (RnKazalPI) of R. neglectus. The nucleotide sequences of the proteins were obtained from the transcriptome of R. neglectus, cloned in the vector pET100/D-TOPO and expressed in Escherichia coli bacteria of line BL21(DE3), BL(DE3)pLysS and Rosetta(DE3)pLysS. The recombinant trypsin-like serine protease (RnTrypsin) was observed only in the insoluble fraction after bacterial lysis, and was then purified by affinity chromatography and used for the production of polyclonal antibodies in mice. After optimization of the expression protocol, the Kazal type serine protease inhibitor (RnKazalPI) was expressed in the soluble fraction of the bacterial lysate. As a perspective, it is intended to use the recombinant protein expression system in insect/baculovirus cells to express the recombinant trypsin-like serine protease (RnTrypsin) in the soluble fraction, and also to determine the characterization of the recombinant Kazal type serine protease inhibitor (RnKazalPI), seeking to contribute to the understanding of the interactions between vector and host and potential pharmacological activities

ASPECTOS FACILITADORES E DIFICULTADORES NO ABANDONO DO TABAGISMO ENTRE PESSOAS COM DIABETES MELLITUS TIPO 2

RESUMO Objetivo: apreender comportamentos e percepções de pacientes com diabetes mellitus tipo 2, tabagistas e ex-tabagistas sobre o tabagismo. Metodologia: estudo descritivo, de abordagem qualitativa, realizado em um município do noroeste do Paraná. Participaram 23 pacientes com diabetes mellitus tipo 2 (13 tabagistas e 10 ex-tabagistas). A coleta de dados ocorreu entre janeiro e março de 2017 por meio de entrevistas domiciliares. A análise de conteúdo, modalidade temática, foi empregada como referencial metodológico analítico. Resultados: identificou-se que ter percepção positiva sobre o abandono do vício; apresentar doenças respiratórias e/ou agravos do diabetes mellitus; ter receio de sofrer com complicações no futuro; e receber apoio profissional e familiar influenciava positivamente para a cessação do tabagismo. Contudo, a ambígua relação de domínio e dependência sobre o tabaco; o reconhecimento de que o tabagismo não é prejudicial às pessoas com diabetes mellitus; e a ausência de apoio familiar dificultavam o abandono. Conclusão: profissionais de saúde necessitam elaborar estratégias de intervenção que acolham os tabagistas e favoreçam a aquisição de mais conhecimentos sobre os malefícios do tabaco para o diabetes mellitus tipo 2, sendo que a inclusão das famílias nesse processo parece oportuna

Triatomines: Trypanosomatids, Bacteria, and Viruses Potential Vectors? [Review]

Triatominae bugs are the vectors of Chagas disease, a major concern to public health especially in Latin America, where vector-borne Chagas disease has undergone resurgence due mainly to diminished triatomine control in many endemic municipalities. Although the majority of Triatominae species occurs in the Americas, species belonging to the genus Linshcosteus occur in India, and species belonging to the Triatoma rubrofasciata complex have been also identified in Africa, the Middle East, South-East Asia, and in the Western Pacific. Not all of Triatominae species have been found to be infected with Trypanosoma cruzi, but the possibility of establishing vector transmission to areas where Chagas disease was previously non-endemic has increased with global population mobility. Additionally, the worldwide distribution of triatomines is concerning, as they are able to enter in contact and harbor other pathogens, leading us to wonder if they would have competence and capacity to transmit them to humans during the bite or after successful blood feeding, spreading other infectious diseases. In this review, we searched the literature for infectious agents transmitted to humans by Triatominae. There are reports suggesting that triatomines may be competent vectors for pathogens such as Serratia marcescens, Bartonella, and Mycobacterium leprae, and that triatomine infection with other microrganisms may interfere with triatomine-T. cruzi interactions, altering their competence and possibly their capacity to transmit Chagas disease

Triatomines : trypanosomatids, bacteria, and viruses potential vectors?

Triatominae bugs are the vectors of Chagas disease, a major concern to public health especially in Latin America, where vector-borne Chagas disease has undergone resurgence due mainly to diminished triatomine control in many endemic municipalities. Although the majority of Triatominae species occurs in the Americas, species belonging to the genus Linshcosteus occur in India, and species belonging to the Triatoma rubrofasciata complex have been also identified in Africa, the Middle East, South-East Asia, and in the Western Pacific. Not all of Triatominae species have been found to be infected with Trypanosoma cruzi, but the possibility of establishing vector transmission to areas where Chagas disease was previously non-endemic has increased with global population mobility. Additionally, the worldwide distribution of triatomines is concerning, as they are able to enter in contact and harbor other pathogens, leading us to wonder if they would have competence and capacity to transmit them to humans during the bite or after successful blood feeding, spreading other infectious diseases. In this review, we searched the literature for infectious agents transmitted to humans by Triatominae. There are reports suggesting that triatominesmay be competent vectors for pathogens such as Serratiamarcescens, Bartonella, andMycobacteriumleprae, and that triatomine infection with other microrganisms may interfere with triatomine-T. cruzi interactions, altering their competence and possibly their capacity to transmit Chagas disease

Cobertura do Sistema de Vigilância Alimentar e Nutricional (SISVAN), estado nutricional de idosos e sua relação com desigualdades sociais no Brasil, 2008-2019: estudo ecológico de série temporal

Objective: To analyze the temporal trend in the coverage of the Food and Nutrition Surveillance System (SISVAN) and the nutritional status of the elderly, correlating them with indicators of social inequality. Methods: Ecological study using SISVAN records of the population aged ≥60 years. Correlation analyzes were performed between indicators of social inequality and the rate of increase in nutritional status and analyzes of absolute and relative inequalities to obtain the angular inequality index and the concentration index. Results: 11,587,933 records were identified. National coverage evolved from 0.1% in 2008 to 2.9% in 2019, with a statistically significant upward trend. A moderate inverse correlation was found with the annual increase rate of overweight for HDI and per capita GDP. Conclusion: There was a trend of growth in SISVAN coverage. The increase in overweight was associated with social inequality.Objetivo: Analizar la tendencia temporal de cobertura del Sistema de Vigilancia Alimentaria y Nutricional (SISVAN), y estado nutricional de adultos mayores, correlacionándolos con indicadores de desigualdad social. Métodos: Estudio ecológico mediante registros SISVAN de la población ≥60 años. Se realizaron análisis de correlación entre indicadores de desigualdad social y la tasa de incremento del estado nutricional y análisis de desigualdades absolutas y relativas para obtener el índice de desigualdad angular y el índice de concentración. Resultados: Se identificaron 11.587.933 registros. La cobertura nacional evolucionó del 0,1% en 2008 al 2,9% en 2019, con una tendencia ascendente estadísticamente significativa. Se encontró una correlación inversa moderada con la tasa de incremento anual de sobrepeso para IDH y PIB per cápita. Conclusión: Hubo una tendencia de crecimiento en la cobertura del SISVAN. El aumento del sobrepeso se asoció con la desigualdad social.Objetivo: analisar a tendência temporal da cobertura do Sistema de Vigilância Alimentar e Nutricional (SISVAN) e do estado nutricional de idosos, e sua correlação com indicadores de desigualdade social no Brasil, no período 2008-2019. Métodos: estudo ecológico, sobre registros do SISVAN relativos à população na idade de 60 anos ou mais; analisaram-se a tendência temporal da cobertura e a correlação entre indicadores de desigualdade social e taxa de incremento do estado nutricional; os índices angular e de concentração foram utilizados para medir desigualdades absolutas e relativas. Resultados: foram identificados 11.587.933 registros de idosos; a cobertura nacional evoluiu de 0,1% (2008) para 2,9% (2019), com tendência de aumento estatisticamente significativa; foi encontrada correlação inversa moderada com taxa de incremento anual de sobrepeso, para índice de desenvolvimento humano e produto interno bruto per capita. Conclusão: houve tendência de crescimento da cobertura do SISVAN; o aumento de sobrepeso esteve associado à desigualdade social

Microcephaly in Pernambuco State, Brazil: epidemiological characteristics and evaluation of the diagnostic accuracy of cutoff points for reporting suspected cases.

The increase in the number of reported cases of microcephaly in Pernambuco State, and Northeast Brazil, characterized an epidemic that led the Brazilian Ministry of Health to declare a national public health emergency. The Brazilian Ministry of Health initially defined suspected cases as newborns with gestational age (GA) ≥ 37 weeks and head circumference (HC) ≤ 33cm, but in December 2015 this cutoff was lowered to 32cm. The current study aimed to estimate the accuracy, sensitivity, and specificity of different cutoff points for HC, using ROC curves, with the Fenton and Intergrowth (2014) curves as the gold standard. The study described cases reported in Pernambuco from August 8 to November 28, 2015, according to sex and GA categories. The Fenton and Intergrowth methods provide HC growth curves according to GA and sex, and microcephaly is defined as a newborn with HC below the 3rd percentile in these distributions. Of the 684 reported cases, 599 were term or post-term neonates. For these, the analyses with ROC curves show that according to the Fenton criterion the cutoff point with the largest area under the ROC curve, with sensitivity greater than specificity, is 32cm for both sexes. Using the Intergrowth method and following the same criteria, the cutoff points are 32cm and 31.5cm for males and females, respectively. The cutoff point identified by the Fenton method (32cm) coincided with the Brazilian Ministry of Health recommendation. Adopting Intergrowth as the standard, the choice would be 32cm for males and 31.5cm for females. The study identified the need to conduct critical and on-going analyses to evaluate cutoff points, including other characteristics for microcephaly case definition

Reflexões e inquietações sobre o alcance da promoção da saúde: Reflections and concerns about the scope of health promotion

INTRODUÇÃO: A Política Nacional de Promoção da Saúde atualizou-se aos auspícios de múltiplos olhares, com diretrizes “ambiciosas” que submergem a um contexto esfacelado de desenvolvimento e crescimento do país. Ante esta complexidade, o cenário brasileiro sobrevive às turbulências políticas, econômicas, sociais, aos confrontos dos valores éticos, morais, legais, dentre outras ameaças, que acenam na contramão das diretrizes de um sistema de saúde universal, equânime e integral. Percebe-se o fosso de um distanciamento crescente entre um discurso apregoado e uma prática ambígua, anuviada e infértil. OBJETIVO: Propor uma reflexão sobre a complexidade que confronta o impacto da política nacional de promoção da saúde no atual cenário brasileiro. MÉTODOS: Ensaio reflexivo, utilizando-se de etapas da estratégia pedagógica Ciclo de Discussão de Problemas (CDP), um desdobramento do método de Aprendizagem Baseada em Problemas e similares. RESULTADOS: A participação social é uma prática de cidadania que não se limita no brandir das bandeiras, mas se sustenta no ato de tomar parte em decisões, as quais envolvem competências que precisam ser adquiridas. Para tal, se defende dois elementos inseparáveis: a igualdade e a liberdade, onde a busca da primeira possibilita a ampliação da segunda, em uma relação que promova o contínuo desenvolvimento dos direitos humanos e uma efetiva democratização da sociedade. CONCLUSÃO: O alcance da promoção enquanto ação se limita ao campo da saúde, que desenvolve experiências intersetoriais isoladas, divulgadas como grandes vitórias espaçadas no meio de algumas derrotas. Uma reflexão crítica faz-se necessária para incorporação dos princípios epistêmicos que subjazem o campo da saúde coletiva na reorganização da atenção à saúde das populações.INTRODUÇÃO: A Política Nacional de Promoção da Saúde atualizou-se aos auspícios de múltiplos olhares, com diretrizes “ambiciosas” que submergem a um contexto esfacelado de desenvolvimento e crescimento do país. Ante esta complexidade, o cenário brasileiro sobrevive às turbulências políticas, econômicas, sociais, aos confrontos dos valores éticos, morais, legais, dentre outras ameaças, que acenam na contramão das diretrizes de um sistema de saúde universal, equânime e integral. Percebe-se o fosso de um distanciamento crescente entre um discurso apregoado e uma prática ambígua, anuviada e infértil. OBJETIVO: Propor uma reflexão sobre a complexidade que confronta o impacto da política nacional de promoção da saúde no atual cenário brasileiro. MÉTODOS: Ensaio reflexivo, utilizando-se de etapas da estratégia pedagógica Ciclo de Discussão de Problemas (CDP), um desdobramento do método de Aprendizagem Baseada em Problemas e similares. RESULTADOS: A participação social é uma prática de cidadania que não se limita no brandir das bandeiras, mas se sustenta no ato de tomar parte em decisões, as quais envolvem competências que precisam ser adquiridas. Para tal, se defende dois elementos inseparáveis: a igualdade e a liberdade, onde a busca da primeira possibilita a ampliação da segunda, em uma relação que promova o contínuo desenvolvimento dos direitos humanos e uma efetiva democratização da sociedade. CONCLUSÃO: O alcance da promoção enquanto ação se limita ao campo da saúde, que desenvolve experiências intersetoriais isoladas, divulgadas como grandes vitórias espaçadas no meio de algumas derrotas. Uma reflexão crítica faz-se necessária para incorporação dos princípios epistêmicos que subjazem o campo da saúde coletiva na reorganização da atenção à saúde das populações

Neighbourhood-level income and Zika virus infection during pregnancy in Recife, Pernambuco, Brazil: an ecological perspective, 2015-2017.

Zika virus (ZIKV) infections during pregnancy can lead to adverse neurodevelopmental and clinical outcomes in congenitally infected offspring. As the city of Recife in Pernambuco State, Brazil-the epicentre of the Brazilian microcephaly epidemic-has considerable disparities in living conditions, this study used an ecological approach to investigate the association between income at the neighbourhood level and the risk of ZIKV infections in pregnant individuals between December 2015 and April 2017. The spatial distribution of pregnant individuals with ZIKV infection was plotted on a map of Recife stratified into four categories based on mean monthly income of household heads. Additionally, a Poisson regression model with robust variance was fitted to compare proportions of ZIKV infections among pregnant individuals in relation to the mean monthly income of household heads, based on the 2010 census data, across 94 neighbourhoods in Recife. The results provide evidence that the risk of ZIKV infection to pregnant individuals was higher among those residing in lower-income neighbourhoods: relative to neighbourhoods that had a mean monthly income of ≥5 times minimum wage, neighbourhoods with <1 and 1 to <2 times minimum wage had more than four times the risk (incidence rate ratio, 95% CI 4.08, 1.88 to 8.85 and 4.30, 2.00 to 9.20, respectively). This study provides evidence of a strong association between neighbourhood-level income and ZIKV infection risks in the pregnant population of Recife. In settings prone to arboviral outbreaks, locally targeted interventions to improve living conditions, sanitation, and mosquito control should be a key focus of governmental interventions to reduce risks associated with ZIKV infections during pregnancy

Association between Zika virus infection and microcephaly in Brazil, January to May, 2016: preliminary report of a case-control study.

BACKGROUND: The microcephaly epidemic, which started in Brazil in 2015, was declared a Public Health Emergency of International Concern by WHO in 2016. We report the preliminary results of a case-control study investigating the association between microcephaly and Zika virus infection during pregnancy. METHODS: We did this case-control study in eight public hospitals in Recife, Brazil. Cases were neonates with microcephaly. Two controls (neonates without microcephaly), matched by expected date of delivery and area of residence, were selected for each case. Serum samples of cases and controls and cerebrospinal fluid samples of cases were tested for Zika virus-specific IgM and by quantitative RT-PCR. Laboratory-confirmed Zika virus infection during pregnancy was defined as detection of Zika virus-specific IgM or a positive RT-PCR result in neonates. Maternal serum samples were tested by plaque reduction neutralisation assay for Zika virus and dengue virus. We estimated crude odds ratios (ORs) and 95% CIs using a median unbiased estimator for binary data in an unconditional logistic regression model. We estimated ORs separately for cases with and without radiological evidence of brain abnormalities. FINDINGS: Between Jan 15, 2016, and May 2, 2016, we prospectively recruited 32 cases and 62 controls. 24 (80%) of 30 mothers of cases had Zika virus infection compared with 39 (64%) of 61 mothers of controls (p=0·12). 13 (41%) of 32 cases and none of 62 controls had laboratory-confirmed Zika virus infection; crude overall OR 55·5 (95% CI 8·6-∞); OR 113·3 (95% CI 14·5-∞) for seven cases with brain abnormalities; and OR 24·7 (95% CI 2·9-∞) for four cases without brain abnormalities. INTERPRETATION: Our data suggest that the microcephaly epidemic is a result of congenital Zika virus infection. We await further data from this ongoing study to assess other potential risk factors and to confirm the strength of association in a larger sample size. FUNDING: Brazilian Ministry of Health, Pan American Health Organization, and Enhancing Research Activity in Epidemic Situations

Co-circulation of Chikungunya Virus during the 2015-2017 Zika Virus Outbreak in Pernambuco, Brazil: An Analysis of the Microcephaly Epidemic Research Group Pregnancy Cohort.

Co-circulation of arthropod-borne viruses, particularly those with shared mosquito vectors like Zika (ZIKV) and Chikungunya (CHIKV), is increasingly reported. An accurate differential diagnosis between ZIKV and CHIKV is of high clinical importance, especially in the context of pregnancy, but remains challenging due to limitations in the availability of specialized laboratory testing facilities. Using data collected from the prospective pregnancy cohort study of the Microcephaly Epidemic Research Group, which followed up pregnant persons with rash during the peak and decline of the 2015-2017 ZIKV epidemic in Recife, Pernambuco, Brazil, this study aims to describe the geographic and temporal distribution of ZIKV and CHIKV infections and to investigate the extent to which ZIKV and CHIKV infections may be clinically differentiable. Between December 2015 and June 2017, we observed evidence of co-circulation with laboratory confirmation of 213 ZIKV mono-infections, 55 CHIKV mono-infections, and 58 sequential ZIKV/CHIKV infections (i.e., cases with evidence of acute ZIKV infection with concomitant serological evidence of recent CHIKV infection). In logistic regressions with adjustment for maternal age, ZIKV mono-infected cases had lower odds than CHIKV mono-infected cases of presenting with arthralgia (aOR, 99% CI: 0.33, 0.15-0.74), arthritis (0.35, 0.14-0.85), fatigue (0.40, 0.17-0.96), and headache (0.44, 0.19-1.90). However, sequential ZIKV/CHIKV infections complicated discrimination, as they did not significantly differ in clinical presentation from CHIKV mono-infections. These findings suggest clinical symptoms alone may be insufficient for differentiating between ZIKV and CHIKV infections during pregnancy and therefore laboratory diagnostics continue to be a valuable tool for tailoring care in the event of arboviral co-circulation