Oswaldo Gonçalves Cruz

Multiplex cytokine profile from dengue patients: MIP-1beta and IFN-gamma as predictive factors for severity

BackgroundDengue virus pathogenesis is not yet fully understood and the identification of patients at high risk for developing severe disease forms is still a great challenge in dengue patient care. During the present study, we evaluated prospectively the potential of cytokines present in plasma from patients with dengue in stratifying disease severity.MethodsSeventeen-cytokine multiplex fluorescent microbead immunoassay was used for the simultaneous detection in 59 dengue patients. GLM models using bimodal or Gaussian family were determined in order to associate cytokines with clinical manifestations and laboratory diagnosis.ResultsIL-1β, IFN-γ, IL-4, IL-6, IL-13, IL-7 and GM-CSF were significantly increased in patients with severe clinical manifestations (severe dengue) when compared to mild disease forms (mild dengue). In contrast, increased MIP-1β levels were observed in patients with mild dengue. MIP-1β was also associated with CD56+NK cell circulating rates. IL-1β, IL-8, TNF-α and MCP-1 were associated with marked thrombocytopenia. Increased MCP-1 and GM-CSF levels correlated with hypotension. Moreover, MIP-1β and IFN-γ were independently associated with both dengue severity and disease outcome.ConclusionOur data demonstrated that the use of a multiple cytokine assay platform was suitable for identifying distinct cytokine profiles associated with the dengue clinical manifestations and severity. MIP-β is indicated for the first time as a good prognostic marker in contrast to IFN-γ that was associated with disease severity.

Spatial Evaluation and Modeling of Dengue Seroprevalence and Vector Density in Rio de Janeiro, Brazil

Background Rio de Janeiro, Brazil, experienced a severe dengue fever epidemic in 2008. This was the worst epidemic ever, characterized by a sharp increase in case-fatality rate, mainly among younger individuals. A combination of factors, such as climate, mosquito abundance, buildup of the susceptible population, or viral evolution, could explain the severity of this epidemic. The main objective of this study is to model the spatial patterns of dengue seroprevalence in three neighborhoods with different socioeconomic profiles in Rio de Janeiro. As blood sampling coincided with the peak of dengue transmission, we were also able to identify recent dengue infections and visually relate them to Aedes aegypti spatial distribution abundance. We analyzed individual and spatial factors associated with seroprevalence using Generalized Additive Model (GAM). Methodology/Principal Findings Three neighborhoods were investigated: a central urban neighborhood, and two isolated areas characterized as a slum and a suburban area. Weekly mosquito collections started in September 2006 and continued until March 2008. In each study area, 40 adult traps and 40 egg traps were installed in a random sample of premises, and two infestation indexes calculated: mean adult density and mean egg density. Sera from individuals living in the three neighborhoods were collected before the 2008 epidemic (July through November 2007) and during the epidemic (February through April 2008). Sera were tested for DENV-reactive IgM, IgG, Nested RT-PCR, and Real Time RT-PCR. From the before–after epidemics paired data, we described seroprevalence, recent dengue infections (asymptomatic or not), and seroconversion. Recent dengue infection varied from 1.3% to 14.1% among study areas. The highest IgM seropositivity occurred in the slum, where mosquito abundance was the lowest, but household conditions were the best for promoting contact between hosts and vectors. By fitting spatial GAM we found dengue seroprevalence hotspots located at the entrances of the two isolated communities, which are commercial activity areas with high human movement. No association between recent dengue infection and households high mosquito abundance was observed in this sample. Conclusions/Significance This study contributes to better understanding the dynamics of dengue in Rio de Janeiro by assessing the relationship between dengue seroprevalence, recent dengue infection, and vector density. In conclusion, the variation in spatial seroprevalence patterns inside the neighborhoods, with significantly higher risk patches close to the areas with large human movement, suggests that humans may be responsible for virus inflow to small neighborhoods in Rio de Janeiro. Surveillance guidelines should be further discussed, considering these findings, particularly the spatial patterns for both human and mosquito populations.

Utilização de ferramentas de análise espacial na vigilância epidemiológica de leishmaniose visceral americana - Araçatuba, São Paulo, Brasil, 1998-1999

The control of American visceral leishmaniasis (AVL) is based on combating the vector and eliminating the domestic reservoir of the focus area - defined as 200 meters around human or canine cases. This paper discusses the use of spatial analysis techniques in the epidemiological surveillance of AVL in Araçatuba, São Paulo State, in order to propose a model for territorial epidemiological surveillance, reformulating current control strategies. The results showed that AVL transmission was not homogeneous; human cases were more frequent in areas with higher canine prevalence rates. Vector dispersion appeared to be restricted to a few houses, although it was not possible to model the vector density. In order to study the vector distribution and correlated covariates, a field study based on house sampling is being conducted. The results will aid the development of new spatial analysis tools and possibly redefine protocols and routines for the control of this endemic disease in urban areas.

Aplicação de modelo bayesiano empírico na análise espacial da ocorrência de hanseníase

OBJETIVO: Analisar a distribuicao espacial da hanseniase, identificar areas de possivel sub-registro de casos ou de provavel alta transmissao (risco) e verificar a associacao dessa distribuicao a existencia de casos de formas multibacilares. METODOS: O estudo foi realizado em Recife, PE, de acordo com 94 bairros analisados. A fonte de coleta de dados foi o Sistema de Informacoes sobre Agravos de Notificacao do Ministerio da Saude. Foi adotada uma abordagem ecologica com utilizacao do metodo bayesiano empirico para suavizacao local de taxas, a partir de informacoes de bairros vizinhos por adjacencia. RESULTADOS: A ocorrencia media anual foi de 17,3% de casos novos em menores de 15 anos (28,3% de formas multibacilares), indicando um processo de intensa transmissao da doenca. A analise da distribuicao espacial de hanseniase apontou tres areas onde se concentram bairros com taxas de deteccao elevadas e que possuem baixa condicao de vida. CONCLUSOES: O emprego do modelo bayesiano, baseado em informacoes de unidades espaciais vizinhas, permitiu estimar novamente indicadores epidemiologicos. Foi possivel identificar areas prioritarias para o programa de controle de hanseniase no municipio, tanto pelo elevado numero de ocorrencias correlacionado a presenca de formas multibacilares de doenca em menores de 15 anos quanto pela existencia de subnotificacao.

Epigrass: a tool to study disease spread in complex networks.

BackgroundThe construction of complex spatial simulation models such as those used in network epidemiology, is a daunting task due to the large amount of data involved in their parameterization. Such data, which frequently resides on large geo-referenced databases, has to be processed and assigned to the various components of the model. All this just to construct the model, then it still has to be simulated and analyzed under different epidemiological scenarios. This workflow can only be achieved efficiently by computational tools that can automate most, if not all, these time-consuming tasks. In this paper, we present a simulation software, Epigrass, aimed to help designing and simulating network-epidemic models with any kind of node behavior.ResultsA Network epidemiological model representing the spread of a directly transmitted disease through a bus-transportation network connecting mid-size cities in Brazil. Results show that the topological context of the starting point of the epidemic is of great importance from both control and preventive perspectives.ConclusionEpigrass is shown to facilitate greatly the construction, simulation and analysis of complex network models. The output of model results in standard GIS file formats facilitate the post-processing and analysis of results by means of sophisticated GIS software.

Perfil de risco: método multivariado de classificação sócio-econômica de microáreas urbanas - os setores censitários da região metropolitana do Rio de Janeiro

A method based on small area data analysis was developed to build a health risk classification for the Greater Rio de Janeiro Metropolitan Area. The approach uses 1991 census data and studies data pertaining to sanitation, ownership and type of housing, size and occupancy of the household, demography, schooling, and income. Principal component analysis applied over each dimension allowed for the choice of 15 variables, which summarized most of the observed variances. Additional analysis with these variables suggested that just six variables are sufficient for the construction of a classification using k-means method of multivariate cluster analysis. Five classes were obtained: (A) high income; (B) lower income; (C) poor; (D) low schooling and income; (E) low-level access to sanitation. The existing inequality in each of the geopolitical established areas was clearly identified. The proposed method allowed for the construction of compound indices to evaluate quality of life, based on widespread and easily obtained data (the census). Moreover, the method contributed to the detection of socioeconomic inequality, identifying, not only the larger poor regions but also the small excluded areas.

Circulating cytokines and chemokines associated with plasma leakage and hepatic dysfunction in Brazilian children with dengue fever.

Dengue fever is usually a benign acute viral infection transmitted by arthropods but may evolve to severe clinical manifestations such as coagulation and/or hemodynamic disorders, caused mainly by an increase of vascular permeability. Deregulated circulating immunological factors have been associated with severity. In Brazil severe cases appeared in children only recently and we evaluated the profile of cytokine/chemokine kinetics in 134 hospitalized young patients during the epidemic in Rio de Janeiro in 2008. Inflammatory cytokines TNF and IFNγ were found elevated during the acute phase in children as well as the anti-inflammatory IL10 and chemokines MIF and CXCL10/IP10, all last three persisting longer during the recovery phase. Severe disease fitting the dengue hemorrhagic fever pattern (WHO, 1997) was associated with higher IL10 and CXCL10/IP10 circulating levels (peak levels at seven days with P<0.01 and P<0.001 respectively as compared to DF). These factors were higher in patients pulmonary effusion or ascites (P<0.05 for IL10 and P<0.01 for CXCL10/IP10). Both factors were also associated with liver changes such as AST increase correlated with CXCL10/IP10 (r=0.4300 with P<0.0001) and patients presenting painful hepatomegaly showed higher circulating levels of IL10 (P<0.01, at 7-9 days) and of CXCL10/IP10 (P<0.05, 4-6 days and P<0.001, 7-9 days) when compared to patients without apparent liver alterations. Most cases presented a history of prior infection (93%). This is the first study demonstrating cytokine and chemokine association with severity during dengue fever in Brazilian children. IL10 and CXCL10/IP10 play a role in the disease severity associated with induction of vascular leakage and a novel association with changes in liver dysfunction.

Single nucleotide polymorphisms in candidate genes and dengue severity in children: A case–control, functional and meta-analysis study

Dengue is an arthropod-borne emerging viral disease with high morbidity and mortality risk in tropical countries like Brazil. Clinical manifestations are vast, ranging from asymptomatic to most severe forms of dengue such as shock. Previous data have shown that host genetics play a role in disease susceptibility and severity. Herein, we have tested the association of single nucleotide polymorphisms (SNPs) at TNF, IL10, MIF, DCSIGN, CLEC5A, NOD2, CCR5 and MRC1 as candidate genes using a matched case-control study design including 88 severe children cases of dengue patients and 335 healthy unrelated subjects that was also separated in IgG(+) and IgG(-) controls. We demonstrated that the TT genotype of CLEC5A SNP (rs1285933 C>T) is associated with dengue severity (OR=2.25; p=0.03) and that GG genotype of -336G>A DCSIGN (CD209) SNP is associated with protection to severe dengue (OR=0.12; p=0.04). Both comparisons were borderline significant when cases were compared with IgG(+) controls subgroup. Nevertheless, genotype-phenotype correlation was also assessed using serum levels of TNF from infected patients at the onset of dengue fever, and CT/TT carriers in CLEC5A secreted higher levels of TNF than CC individuals in 5-7 days of infection. No significant difference was observed in TNF levels between genotypes GG versus AG/AA at DCSIGN promoter. Next, we performed a meta-analysis retrieving results from the literature for -336G>A DCSIGN and -308G>A TNF SNPs demonstrating that the consensus estimates of these SNPs indicated no association with dengue severity (when compared to Dengue fever) in the overall analysis. But, a subgroup analysis in the -336G>A DCSIGN, the G allele was associated with severe dengue susceptibility in Asians (ORallele=2.77; p=0.0001; ORcarriers=2.99; p=0.0001) and protection in Brazilians (ORallele=0.66; p=0.013). In summary, our results suggest that genetic variations at CLEC5A increase the risk and regulate TNF secretion in dengue severity among Brazilians. Also, combined data of the literature suggest population-specific effect of the -336 DCSIGN SNP more prominent in Asians and in a different direction than Brazilians.

Clinical and hepatic evaluation in adult dengue patients: a prospective two-month cohort study

INTRODUCTION To analyze the liver dysfunction and evolution of signs and symptoms in adult dengue patients during a two-month follow-up period. METHODS A prospective cohort study was conducted in Campos dos Goytacazes, Rio de Janeiro, Brazil, from January to July, 2008. The evolution of laboratory and clinical manifestations of 90 adult dengue patients was evaluated in five scheduled visits within a two-month follow-up period. Twenty controls were enrolled for the analysis of liver function. Patients with hepatitis B, hepatitis C, those known to be human immunodeficiency virus (HIV) seropositive and pregnant women were excluded from the study. RESULTS At the end of the second month following diagnosis, we observed that symptoms persisted in 33.3% (30/90) of dengue patients. We also observed that, 57.7% (15/26) of the symptoms persisted at the end of the second month. The most persistent symptoms were arthralgia, fatigue, weakness, adynamia, anorexia, taste alteration, and hair loss. Prior dengue virus (DENV) infection did not predispose patients to a longer duration of symptoms. Among hepatic functions, transaminases had the most remarkable elevation and in some cases remained elevated up to the second month after the disease onset. Alanine aminotransferase (ALT) levels overcame aspartate aminotransferase (AST) during the convalescent period. Male patients were more severely affected than females. CONCLUSIONS Dengue fever may present a wide number of symptoms and elevated liver transaminases at the end of the second month.

Declining prevalence of hepatitis A virus antibodies among children from low socioeconomic groups reinforces the need for the implementation of hepatitis A vaccination in Brazil

Age-related seroprevalence studies that have been conducted in Brazil have indicated a transition from a high to a medium endemicity of hepatitis A virus (HAV) infection in the population. However, most of these studies have focused on urban populations that experience lower incidence rates of HAV infection. In the current study, the prevalence of anti-HAV antibodies was investigated in children with a low socioeconomic status (SES) that live on the periphery of three capital cities in Brazil. A total of 1,162 dried blood spot samples were collected from individuals whose ages ranged from one-18 years and tested for anti-HAV antibodies. A large number of children under five years old (74.1-90%) were identified to be susceptible to HAV infection. The anti-HAV antibody prevalence reached ≥ 50% among those that were 10-14 years of age or older. The anti-HAV prevalence rates observed were characteristics of regions with intermediate level of hepatitis A endemicity. These data indicated that a large proportion of children with a low SES that live at the periphery of urban cities might be at risk of contracting an HAV infection. The hepatitis A vaccine that is currently offered in Brazil is only available for high-risk groups or at private clinics and is unaffordable for individuals with a lower SES. The results from this study suggest that the hepatitis A vaccine should be included in the Brazilian National Program for Immunisation.