Cynomolgus monkeys (Macaca fascicularis) experimentally infected with B19V and hepatitis A virus: no evidence of the co-infection as a cause of acute liver failure

This study was conducted to analyse the course and the outcome of the liver disease in the co-infected animals in order to evaluate a possible synergic effect of human parvovirus B19 (B19V) and hepatitis A virus (HAV) co-infection. Nine adult cynomolgus monkeys were inoculated with serum obtained from a fatal case of B19V infection and/or a faecal suspension of acute HAV. The presence of specific antibodies to HAV and B19V, liver enzyme levels, viraemia, haematological changes, and necroinflammatory liver lesions were used for monitoring the infections. Seroconversion was confirmed in all infected groups. A similar pattern of B19V infection to human disease was observed, which was characterised by high and persistent viraemia in association with reticulocytopenia and mild to moderate anaemia during the period of investigation (59 days). Additionally, the intranuclear inclusion bodies were observed in pro-erythroblast cell from an infected cynomolgus and B19V Ag in hepatocytes. The erythroid hypoplasia and decrease in lymphocyte counts were more evident in the co-infected group. The present results demonstrated, for the first time, the susceptibility of cynomolgus to B19V infection, but it did not show a worsening of liver histopathology in the co-infected group.

Detection of enteric viruses in activated sludge by feasible concentration methods.

Human enteric viruses are responsible to cause several diseases, including gastroenteritis and hepatitis, and can be present in high amounts in sewage sludge. This study compared virus recovery efficiency of two feasible concentration methods used for detecting human adenovirus (HAdV), rotavirus species A (RV-A), norovirus genogroup II (NoV GII) and hepatitis A virus (HAV) in sewage sludge from an activated sludge process. Twelve sewage sludge samples were collected bi-monthly from January to July, 2011. Ultracentrifugation was compared with a simplified protocol based on beef extract elution for recovering enteric viruses. Viruses were quantified by quantitative real-time PCR assays and virus recovery efficiency and limits of detection were determined. Methods showed mean recovery rates lower than 7.5%, presenting critical limits of detection (higher than 10(2) - 10(3) genome copies - GC L(-1) for all viruses analyzed). Nevertheless, HAdV were detected in 90% of the analyzed sewage sludge samples (range: 1.8 × 10(4) to 1.1 × 10(5) GC L(-1)), followed by RV-A and NoV (both in 50%) and HAV (8%). Results suggesting that activated sludge is contaminated with high viral loads and HAdV are widely disseminated in these samples. The low virus recovery rates achieved, especially for HAV, indicate that other feasible concentration methods could be developed to improve virus recovery efficiency in these environmental matrices.

Assessment of water quality in a border region between the Atlantic forest and an urbanised area in Rio de Janeiro, Brazil.

The preservation of water resources is one of the goals of the designation of parks that act as natural reservoirs. In order to assess the impact of the presence of humans in an environmental preservation area bordering urban areas, the presence of four pathogenic enteric viruses [group A rotavirus (RV-A), norovirus (NoV), human adenoviruses (HAdV), and hepatitis A virus (HAV)], as well as the physico-chemical parameters, and Escherichia coli levels were assessed in riverine water samples. From June 2008 to May 2009, monthly monitoring was performed along the Engenho Novo River. RV-A, NoV, and HAdV were observed in 29% (31/108) of the water samples, with concentrations of up to 10(3) genome copies/liter. The natural occurrence of infectious HAdV was demonstrated by Integrated Cell Culture-PCR (ICC-PCR). This study confirms the suitability of using the detection of fecal-oral transmitted viruses as a marker of human fecal contamination in water matrices and indicates the spread of pathogenic viruses occurring in an alleged area of environmental protection.

Detection of enteric viruses in activated sludge by feasible concentration methods

Human enteric viruses are responsible to cause several diseases, including gastroenteritis and hepatitis, and can be present in high amounts in sewage sludge. This study compared virus recovery efficiency of two feasible concentration methods used for detecting human adenovirus (HAdV), rotavirus species A (RV-A), norovirus genogroup II (NoV GII) and hepatitis A virus (HAV) in sewage sludge from an activated sludge process. Twelve sewage sludge samples were collected bi-monthly from January to July, 2011. Ultracentrifugation was compared with a simplified protocol based on beef extract elution for recovering enteric viruses. Viruses were quantified by quantitative real-time PCR assays and virus recovery efficiency and limits of detection were determined. Methods showed mean recovery rates lower than 7.5%, presenting critical limits of detection (higher than 102 103 genome copies -GC L-1 for all viruses analyzed). Nevertheless, HAdV were detected in 90% of the analyzed sewage sludge samples (range: 1.8 x 104 to 1.1 x 105 GC L-1), followed by RV-A and NoV (both in 50%) and HAV (8%). Results suggesting that activated sludge is contaminated with high viral loads and HAdV are widely disseminated in these samples. The low virus recovery rates achieved, especially for HAV, indicate that other feasible concentration methods could be developed to improve virus recovery efficiency in these environmental matrices.

The efficiency of concentration methods used to detect enteric viruses in anaerobically digested sludge.

The presence of enteric viruses in biosolids can be underestimated due to the inefficient methods (mainly molecular methods) used to recover the viruses from these matrices. Therefore, the goal of this study was to evaluate the different methods used to recover adenoviruses (AdV), rotavirus species A (RVA), norovirus genogroup II (NoV GII) and the hepatitis A virus (HAV) from biosolid samples at a large urban wastewater treatment plant in Brazil after they had been treated by mesophilic anaerobic digestion. Quantitative polymerase chain reaction (PCR) was used for spiking experiments to compare the detection limits of feasible methods, such as beef extract elution and ultracentrifugation. Tests were performed to detect the inhibition levels and the bacteriophage PP7 was used as an internal control. The results showed that the inhibitors affected the efficiency of the PCR reaction and that beef extract elution is a suitable method for detecting enteric viruses, mainly AdV from biosolid samples. All of the viral groups were detected in the biosolid samples: AdV (90%), RVA, NoV GII (45%) and HAV (18%), indicating the viruses' resistance to the anaerobic treatment process. This is the first study in Brazil to detect the presence of RVA, AdV, NoV GII and HAV in anaerobically digested sludge, highlighting the importance of adequate waste management.

The efficiency of concentration methods used to detect enteric viruses in anaerobically digested sludge

The presence of enteric viruses in biosolids can be underestimated due to the inefficient methods (mainly molecular methods) used to recover the viruses from these matrices. Therefore, the goal of this study was to evaluate the different methods used to recover adenoviruses (AdV), rotavirus species A (RVA), norovirus genogroup II (NoV GII) and the hepatitis A virus (HAV) from biosolid samples at a large urban wastewater treatment plant in Brazil after they had been treated by mesophilic anaerobic digestion. Quantitative polymerase chain reaction (PCR) was used for spiking experiments to compare the detection limits of feasible methods, such as beef extract elution and ultracentrifugation. Tests were performed to detect the inhibition levels and the bacteriophage PP7 was used as an internal control. The results showed that the inhibitors affected the efficiency of the PCR reaction and that beef extract elution is a suitable method for detecting enteric viruses, mainly AdV from biosolid samples. All of the viral groups were detected in the biosolid samples: AdV (90%), RVA, NoV GII (45%) and HAV (18%), indicating the viruses' resistance to the anaerobic treatment process. This is the first study in Brazil to detect the presence of RVA, AdV, NoV GII and HAV in anaerobically digested sludge, highlighting the importance of adequate waste management.

Detection of enteric viruses in recreational waters of an urban lagoon in the city of Rio de Janeiro, Brazil.

The aim of this study was to determine the occurrences of the group A rotavirus (RVA), norovirus (NoV) and human adenovirus (HAdV) in the surface waters of an urban lagoon (Rodrigo de Freitas Lagoon) in the city of Rio de Janeiro, Brazil. During one year of surveillance, water samples were obtained from the lagoon and other interconnected ecosystems (river and beach). The samples were concentrated using an adsorption-elution method with a negatively charged membrane and tested by qualitative and quantitative polymerase chain reaction assays. RVA was the most prevalent virus detected (24.3%) with a viral load ranging from 3.0 x 10¹-5.6 x 10(4) genome copies/L, followed by NoV (18.8%) and HAdV (16.7%). Considering water samples suitable for bathing, according to Escherichia coli criterion (< 2,000 most probable number/100 mL), viruses were detected in 50% (57/114) of them. Physicochemical parameters were also measured and showed possible correlations between turbidity and RVA presence and between pH and NoV presence. These data demonstrate the importance of considering viral parameters to ensure water quality and the utilisation of these parameters as additional tools for the characterisation of environmental contamination.

Detection of enteric viruses in recreational waters of an urban lagoon in the city of Rio de Janeiro, Brazil

The aim of this study was to determine the occurrences of the group A rotavirus (RVA), norovirus (NoV) and human adenovirus (HAdV) in the surface waters of an urban lagoon (Rodrigo de Freitas Lagoon) in the city of Rio de Janeiro, Brazil. During one year of surveillance, water samples were obtained from the lagoon and other interconnected ecosystems (river and beach). The samples were concentrated using an adsorption-elution method with a negatively charged membrane and tested by qualitative and quantitative polymerase chain reaction assays. RVA was the most prevalent virus detected (24.3%) with a viral load ranging from 3.0 x 10¹-5.6 x 10(4) genome copies/L, followed by NoV (18.8%) and HAdV (16.7%). Considering water samples suitable for bathing, according to Escherichia coli criterion (< 2,000 most probable number/100 mL), viruses were detected in 50% (57/114) of them. Physicochemical parameters were also measured and showed possible correlations between turbidity and RVA presence and between pH and NoV presence. These data demonstrate the importance of considering viral parameters to ensure water quality and the utilisation of these parameters as additional tools for the characterisation of environmental contamination.

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.

Importance of the cutoff ratio for detecting antibodies against hepatitis A virus in oral fluids by enzyme immunoassay.

Multiple studies have examined the use of oral fluids in modified serum-based assays aiming to replace serum in antibody detection for hepatitis A. However, the reliable detection of HAV immunity in oral fluid requires an extremely sensitive assay; most immunoassays designed for serum antibody determination lack sufficient sensitivity for this purpose. Consequently, an "in-house" competitive enzyme immunoassay (EIA) designed specifically for use with oral samples collected using a ChemBio(®) device was developed to detect total anti-HAV antibodies (IgG and IgM). This system was compared to an in-house competitive EIA and a commercial EIA considered to be the "gold standard" using corresponding serum samples (n=225) to determine the accuracy of the assay and to evaluate the importance of the cutoff ratio for the detection of anti-HAV antibodies in oral fluids. When the median serum cutoff and the optimal oral fluid cutoff (ROC analysis) obtained from the in-house competitive EIA were compared, the oral fluid cutoff was found to be 28.8% higher than the serum cutoff. When different oral fluid cutoff values were compared, a reduction of about 17% was shown to be essential to increase test accuracy. At an oral fluid cutoff value of 0.351, sensitivity and specificity were higher, reaching 91.7% and 86.2% (p<0.001, AUROC=0.915), respectively. The convenience, accuracy and non-invasive nature of the developed method make it a useful alternative to serum-based assays for discriminating between HAV-immune and non-immune individuals.

Hepatitis A virus subgenotypes dissemination during a community outbreak in a surrounding region of Rio de Janeiro

From December 1999 to December 2001, many cases of hepatitis A were notified in the county of Belford Roxo involving individuals aged 0 to 79 years. Serum samples were collected to evaluate the prevalence of anti-hepatitis A virus (HAV) antibodies, to detect HAV-RNA and to correlate with possible risk factors of HAV infection. Serum samples were screened by commercial IgM and total anti-HAV antibody ELISA and HAV-RNA was isolated and subsequently amplified by reverse transcription-polymerase chain reaction (RT-PCR) at VP1/2A region, sequenced and analyzed. Total anti-HAV prevalence was 87.9 percent (203/231) and IgM anti-HAV prevalence was 38.7 percent (89/231). Multivariate analysis showed that individuals under 20 years old are risks groups to acquire the infection suggesting that hygienic habits of young subjects are the principal factor of transmission and so they could be the target for vaccine programs. HAV-RNA was amplified from 29 (32.5 percent) IgM anti-HAV positive patients and 26 samples were sequenced and classified into subgenotypes IB (8 isolates) and IA (18 isolates). Isolates classified into subgenotype IB were identical representing one distinct strain. We could observe both subgenotypes circulating during the study which suggests different sources of infection. Prophylactic measures as vaccination strategies added to improvements in hygienic and sanitary conditions would be highly effective to reduction of infection.

Hepatitis B among female sex workers in Ribeirão Preto - São Paulo, Brazil

OBJECTIVE: To estimate the prevalence of hepatitis B markers and to study the risk factors for this disease among female sex workers in the city of Ribeirão Preto, Brazil. METHODS: A questionnaire was given to 449 female sex workers in order to obtain information about demographic, socioeconomic and behavioral variables. Blood samples were collected and analyzed by immunoenzymatic techniques for detection of HBsAg, anti-HBs, anti-HBc and anti-HCV markers. RESULTS: The mean age of participants was 23 years, varying from 13 to 64 years. A high spatial mobility was observed, with 47.9 percent of participants residing in places out of the region of Ribeirão Preto or in other Brazilian states. Complete absence of previous vaccine against hepatitis B was referred by 98.2 percent. Overall, the presence of any hepatitis B marker was observed in 106 participants (prevalence of 23.6 percent; 95 percent CI: 19.7 27.5), with 84 positive for anti-HBs (18.7 percent), 100 for anti-HBc (22.3 percent), and only 3 for HBsAg (0.7 percent). The logistic regression analysis showed association between hepatitis B markers and the following co-variables: residence in Ribeirão Preto, age, low socioeconomic level, consumption of crack, intercourse with HIV-infected individuals, history of previous hepatitis, intercourse with a case of hepatitis, and positivity for hepatitis C. CONCLUSIONS: Ribeirão Preto's female sex workers present several risk factors for hepatitis B and almost absence of previous specific vaccination, making it necessary to emphasize this low-cost preventive measure, preferably through the use of a mobile team, taking the vaccine to their places of work.

OBJETIVOS: Estimar a prevalência de marcadores de hepatite B e estudar os fatores de risco para esta doença entre mulheres profissionais do sexo na cidade de Ribeirão Preto, Brasil. MÉTODOS: Foi aplicado um questionário a 449 mulheres profissionais do sexo, com a finalidade de levantar informações demográficas, socioeconômicas e comportamentais. Amostras de sangue das participantes foram analisadas através de técnicas imunoenzimáticas, para detecção dos marcadores HBsAg, anti-HBs e anti-HBc. RESULTADOS: A idade média das participantes foi 23 anos, variando de 13 a 64 anos. Uma elevada mobilidade espacial foi verificada, com 47,9 por cento delas residindo em locais fora da região de Ribeirão Preto ou em outros estados brasileiros. Completa ausência de vacinação prévia contra hepatite B foi referida por 98,2 por cento. No total, observou-se presença de qualquer marcador de hepatite B em 106 participantes (prevalência de 23,6 por cento; IC95 por cento: 19,7 27,5), com 84 positivos para anti-HBs (18,7 por cento), 100 para anti-HBc (22,3 por cento) e apenas 3 para HBsAg (0,7 por cento). A análise por regressão logística evidenciou associação entre marcadores de hepatite B e as seguintes co-variáveis: idade, baixo nível socioeconômico, consumo de crack, relações sexuais com indivíduos infectados pelo HIV, história de hepatite prévia, relações sexuais com pessoas portadoras de hepatite e positividade para hepatite C. CONCLUSÕES: As profissionais do sexo em Ribeirão Preto apresentam diversos fatores de risco para hepatite B e quase total ausência de vacinação prévia específica, tornando necessários esforços concentrados na aplicação dessa medida de baixo custo, preferencialmente através do uso de equipes móveis que levem a vacina até os seus locais de trabalho.

Regressão clínica da forma hepatosplênica da esquistossomose mansônica e marcadores séricos do vírus da hepatite B / Clinical regression of the hepatosplenic form of schistosomiasis mansoni and serological markers for hepatitis B virus

A pesquisa da associação entre a ocorrência da regressão clínica da forma hepatosplênica (HE) da esquistosssomose mansônica (EM) e a ausência de marcadores sorológicos do vírus da hepatite B (VHB) foi pesquisada em uma área endêmica para EM no Brasil. O estudo longitudinal (coorte) incluiu membros das famílias residentes na área de transmissão de EM de Catolândia (Estado da Bahia, Brasil), o qual foi iniciado em 1975-76, com periódicos exames clínicos, parasitológicos (Kato-Katz), tratamentos (oxamniquine e/ou praziquantel) e medidas de desenvolvimento sanitário. Amostras séricas (n = 1.273) dos residentes foram testadas para marcadores do VHB (ELISA: AgHBs, anti-HBs e anti-HBc). De 1976 a 1996, após dois ou mais tratamentos específicos, em 72 casos foi observada a regressão clínica da forma HE e não observada em outros 15 casos, mas em ambos os grupos as frequências dos marcadores sorológicos do VHB foram similares (p >/ 0,17). Quando os residentes foram reclassificados em dois grupos (com ou sem prévio contato com o VHB, a distribuição do diagnóstico final de regressão dos pacientes hepatosplênicos (HE) foi também similar (p >0,64). Em conclusão, a presença de AgHBs ou infecção curada pelo não foi associada a não-regressão da forma HE

Prevalência de marcadores do vírus da hepatite B em crianças de 3 a 9 anos em um município da Amazônia brasileira / Prevalence of hepatitis B viral markers in children 3 to 9 years old in a town in the Brazilian Amazon

OBJETIVO: Estimar a prevalência da infecção pelo vírus da hepatite B e identificar as possíveis vias de infecção em crianças de 3 a 9 anos em um município da Amazônia brasileira. MÉTODOS: Foi realizado um estudo transversal com 487 crianças matriculadas em creches e escolas públicas. Foram obtidos dados acerca de migração, história médica, exposição a fatores de risco clássicos para infecção, aspectos socioeconômicos e hábito de compartilhar a cama ou objetos de uso pessoal. Também foi investigada a presença no domicílio de morador com doença sexualmente transmissível, usuário de drogas injetáveis, dependente de bebida alcoólica, homens que faziam sexo com outro homem e pessoa com múltiplos parceiros sexuais, além de história de icterícia ou hepatite e de transfusão de hemoderivados. Foram obtidas amostras de sangue das crianças para pesquisa de marcadores virais. RESULTADOS: A prevalência de marcadores do vírus da hepatite B foi de 11,1 por cento (54 crianças). Seis crianças (1,2 por cento) eram portadoras do HBsAg. Não houve associação entre marcadores virais e condições econômicas, escolaridade dos pais, condições higiênico-sanitárias, número de cômodos do domicílio, história prévia de icterícia ou hepatite, tratamento dentário, acidente com seringas e agulhas, reutilização de agulhas, transfusão de hemoderivados e hábito de compartilhar a cama ou objetos pessoais. Porém, as crianças que tiveram contato com usuário pesado de bebida alcoólica tenderam à maior prevalência de marcadores virais (P= 0,06). Dentre as crianças estudadas, 53 (10,9 por cento) haviam sido vacinadas contra a hepatite B, mas nenhuma havia completado o esquema vacinal de três doses. Na regressão logística, as variáveis "história de icterícia materna durante a gravidez," "mais de oito moradores no domicílio" e "profissão da mãe" mostraram associação com maior prevalência de marcadores virais. CONCLUSÃO: A população de Peixoto de Azevedo provavelmente apresenta endemicidade moderada de hepatite B, tendo a transmissão horizontal como a principal forma de infecção. Fatores ligados à promiscuidade domiciliar possivelmente são determinantes da infecção na primeira década da vida. O baixo índice de cobertura vacinal possibilita a livre circulação viral.

Objective. To compare two pediatric recombinant hepatitis B vaccines­the Engerix-B reference vaccine and the Euvax-B vaccine­in terms of immunogenicity and reactogenicity, and also to investigate their interchangeability, that is, whether a three-dose hepatitis B vaccination schedule begun with a first dose of Engerix-B could be completed with two doses of Euvax-B. Methods. This study was conducted in the city of Córdoba, Argentina, from March 1999 through February 2000. Three groups of Argentine newborns (100 per group) were vaccinated at 0, 1, and 6 months of age with hepatitis B vaccine: group A, three doses of Euvax-B; group B, three doses of Engerix-B; and group C, one dose of Engerix-B followed by two doses of Euvax-B. Reactogenicity was evaluated based on parental reporting of any solicited local or systemic event occurring during the 7-day period following vaccination. Whether Euvax-B and Engerix-B were clinically identical was assessed in terms of the seroprotection rates (antibodies to hepatitis B surface antigen (anti-HBsAg) ≥ 10 milli-international units per mL (mIU/mL) 2 months after the third vaccination). Results. Reactogenicity was low in all three groups. Five months after the second dose (that is, immediately prior to the third vaccination), seroprotection rates were 95.9%, 94.7%, and 90.2% for groups A, B, and C, respectively. Two months after the third dose all subjects were seroprotected, with geometric mean concentrations of anti-HBsAg of 2 468.1, 1 714.8, and 2 075.3 mIU/mL for groups A, B, and C, respectively. Conclusions. Both of the recombinant hepatitis B vaccines that we studied were well tolerated and highly immunogenic. Euvax-B was clinically identical (not inferior) to the EngerixB reference vaccine, and either vaccine could be used to achieve the World Health Organization goal of immunizing all infants against hepatitis B. Further, Euvax-B can be safely used in infants given an initial dose of either Euvax-B or Engerix-B

Prevalência da hepatite B em área rural de município hiperendêmico na Amazônia Mato-grossense: situação epidemiológico / Prevalence of Hepatitis B in a Hiperendemic Rural Area in the Amazon Region of Mato Grosso State: Epidemiological Situation

Em 1995, foi identificada epidemia comunitária de hepatite B entre colonos agrícolas recentemente assentados em Cotriguaçu,noroeste de Mato Grosso. Houve campanha de vacinação nos municípios da região. Nos anos seguintes, manteve-se a estratégia de vacinar a população e os migrantes que continuaram chegando àquele município. Em 2001, foi realizado novo inquérito sorológico de marcadores da hepatites B com o objetivo de avaliar a magnitude do problema e o impacto das medidas tomadaspreviamente. A comunidade de Nova União foi escolhida por ser a área de maior atração de migrantes a partir do final da década de 90. Foram estudados 838 indivíduos, encontrando-se 40,0 por cento já infectados pelo vírus da hepatite B (VHB), 2,1 por cento de portadores do vírus e 40,8 por cento protegidos por vacinação. A maioria dessa população era composta de migrantes vindos de Rondônia. Entre os portadores do VHB, 28,0 por cento tinham marcadores de hepatite D. As variáveis associadas à infecção pelo VHB foram: atividade sexual; etilismo; contato com caso de hepatite; e ter vivido em garimpo. Esse padrão de moderada prevalência é diferente do observado em 1995, ocasião em que se evidenciou alta prevalência. Observou-se associação do VHB com aumento da idade e com início da atividade sexual. Como a migração para aquela área continua intensa, recomenda-se que a atual estratégia de vacinação continue. Especial atenção deve ser dedicada à progressiva entrada do vírus da hepatite Delta (VHD) na região

Comparison of four extraction methods to detect hepatitis A virus RNA in serum and stool samples

The efficiency of extraction methods for hepatitis A virus (HAV) RNA in clinical samples is of great importance for molecular diagnosis, especially in regions endemic for HAV, such as Brazil. We compared the efficiency of four different extraction techniques in serum and stool samples for the detection of hepatitis A virus by reverse transcription PCR (RT-PCR). We used PCR to analyse serum and stool samples of 12 patients who were referred to the Brazilian Reference Center for Viral Hepatitis (BRCVH) in Rio de Janeiro. The methods tested were Proteinase K, Silica, TRIzol and Guanidine isothiocyanate. Proteinase K extraction was the best method for serum samples; it detected the HAV-RNA in 11 of the 12 samples. The guanidine isothiocyanate method was the most effective for stool samples, detecting HAV-RNAs in 9 of the samples. The TRIzol« method worked best with serum samples, and the silica method was unsatisfactory with both serum and stool samples. The RNA extraction method affected the outcome. The use of appropriate RNA extraction methods is a critical step for successful and valid PCR studies on clinical samples. We recommend that RNA extraction techniques be carefully selected for their efficiency with each type of specimen

Situação atual da hepatite B e D na região de Cotriguaçu, Amazônia Mato-grossense, 2001. (Nota Prévia) / Current situation of hepatitis B and D in the Amazon Region of Cotriguaçu, Mato Grosso State, 2001: (Previous Note)

DELINEAMENTO DO PROBLEMA: Em 1995, foi identificada epidemia comunitária de hepatite B entre colonos então recentemente assentados em Cotriguaçu, no noroeste mato-grossense. Houve campanha de vacinação nos municípios da região. Nos anos seguintes, manteve-se a estratégia de vacinar os migrantes que continuaram chegando. Para avaliar a atual situação da infecção pelo VHB na região rural de Cotriguaçu, foi planejado inquérito sobre a prevalência dos marcadores do vírus da hepatite B (VHB) e do vírus de hepatite D (VHD) em 2001. MATERIAL E MÉTODOS: Este estudo, tipo corte-transversal, foi desenhado para rastrear todos os habitantes das vilas. Aos moradores, foi solicitado consentimento por escrito. Foi realizada entrevista e coleta de sangue para testar presença de marcadores de hepatite B e D. Considerou-se como expostos à infecção pelo VHB os indivíduos com anti-HBc positivo, independente de serem positivos ou não para o HBsAg ou anti-HBs. Foram considerados como respondedores à vacina aqueles vacinados contra o VHB, com títulos de anti-HBs acima de 10U/L, e com HBsAg e anti-HBc negativos. Os portadores do VHB foram submetidos à pesquisa de anticorpos contra o VHD. Foram feitas análises univariada e multivariada, com regressões linear e logística. RESULTADOS: De 838 participantes, 335 (40%; IC95%: 36,7-43,4) foram infectados pelo VHB e 18 (2,1%; IC95%: 1,3-3,4) eram portadores do HBsAg. Foram considerados respondedores à vacina contra o VHB, 342 (40,8%; IC95%: 37,5-44,2). Os outros 161 (19,2%; IC95%: 16,6-22,1) eram ainda suscetíveis ao VHB. De 802 indivíduos que responderam à pergunta, 72,3% informaram vacinação prévia contra o VHB e 40,2% afirmaram ter tomado as três doses. Após análise multivariada, mantiveram associação independente com os marcadores do VHB: atividade sexual (p<0,01), uso regular de álcool (p<0,01), contato domiciliar com hepatite (p<0,02) e ter vivido em garimpo (p<0,02), assim como pertencer à faixa etária mais elevada (p<0,05). A avaliação de determinantes dos títulos de anti-HBs nos imunes, controlados por idade e fatores de risco, demonstrou que o fato de ter tomado ao menos uma dose da vacina (p<0,001) esteve associado independentemente a maiores títulos de anti-HBs, assim como ter tido a infecção natural pelo VHB (p<0,001). Cinco (28%) dos 18 portadores do HBsAg tiveram infecção também pelo VHD. CONCLUSÕES: Este inquérito demonstra mudança do padrão epidemiológico de hepatite B de alta para moderada prevalência (40% expostos e 2% de portadores). Análise dos índices de cobertura vacinal mostram que o pool de suscetíveis atualmente é pequeno na região e dificilmente sustentará novos surtos de hepatite B como vistos anteriormente. O crescimento dos casos de VHD pode tornar a região hiperendêmica para essa infecção.

BACKGROUND: A community outbreak of hepatitis B among recently settled farmers in Cotriguaçu County in the northwestern area of Mato Grosso State, Brazil, occurred in 1995. There was a vaccination campaign in the counties of the region. In the following years, efforts were made to maintain the vaccination of new immigrants. To assess the current status of HBV and HDV infection in the countryside of Cotriguaçu a cross-sectional survey was designed and carried out in 2001. MATERIAL AND METHODS:The cross-sectional survey was designed to include the entire population of the villages. The settlers who agreed to participate in the study were asked for written consent. Participants were interviewed and bled to test for HBV and HDV markers. Anti-HBc-positive subjects were considered as HBV exposed, independently if positive for HbsAg or anti-HBs. Participants were considered to be responders to vaccination if they were vaccinated, had anti-HBs titers higher than 10U/L and were negative for HbsAg and anti-HBc. HBsAg carriers were tested for HBD antibodies. Univariate and multivariate analyses, including linear and logistic regression models were performed. RESULTS: A total of 335 of 838 study participants (40%; 95% CI:36.7-43.4) had been exposed to HBV. Eighteen (2.1%; 95% CI:1.3-3.4) were HBV carriers; 342 (40.8%; 95% CI:37.5-44.2) were considered as vaccine responders, and 161 (19.2%; 95% CI:16.6-22.1) were susceptible to HBV infection. Previous vaccination was reported by 580/802 (72.3%) subjects, and 40.2% had received the complete schedule. After adjustment with multivariate analyses, the following variables were independently associated with HBV markers: sexual activity (p<0.01), regular use of alcohol (p<0.01), household contact with hepatitis cases (p<0.02), having lived in mining camps (p<0.02) and older age (p<0.05). Anti-HBs titers among immune subjects were assessed. Having received vaccination, at least one dose, and to have been naturally exposed to HBV were associated with higher anti-HBs titers (p<0.001). Five of 18 HBV carriers (28%) were coinfected with HDV. CONCLUSIONS: This survey showed a change in the HBV epidemiological pattern from high to moderate prevalence (40% exposed and 2% of HBV carriers). Vaccine coverage analysis shows that the pool of susceptibles is small. New HBV outbreaks will not likely take place again in this region. The increasing number of HDV infection can result in this area becoming hyperendemic.

Seasonal variation of hepatitis A virus infection in the city of Rio de Janeiro, Brazil

Hepatitis A virus (HAV) infection constitutes a major public health problem in Brazil. The transmission of HAV is primarily by fecal-oral route so the water is an important vehicle of HAV dissemination. There is a great incidence of acute cases of hepatitis A in some areas of Brazil however the seasonal variation of these cases was not documented. The aim of this study was to determine the seasonality of HAV infection in Rio de Janeiro. From January 1999 to December 2001, 1731 blood samples were collected at the National Reference Center for Hepatitis Viruses in Brazil (NRCHV). These samples were tested by a commercial enzyme-immunoassay to detect anti-HAV IgM antibodies. Yearly positive rates were 33.74 percent in 1999, 32.19 percent in 2000, and 30.63 percent in 2001. A seasonal variation was recognized with the highest incidence in spring and summer. Furthermore a seasonal increase in incidence of HAV infection was found during the rainy season (December to March) because the index of rains is very high. It is concluded that HAV infections occur all year round with a peak during hot seasons with great number of rains

Seroepidemiological markers of enterically transmitted viral hepatitis A and E in individuals living in a community located in the North Area of Rio de Janeiro, RJ, Brazil

We investigated the seroprevalence of hepatitis A virus (HAV) and hepatitis E virus (HEV) infection in subjects living in the community of Manguinhos, Rio de Janeiro, Brazil, and assisted at the Health Unit of Escola Nacional de Saúde Pública, Fundaçäo Oswaldo Cruz. After formal consent, individuals were submitted to an interview using a standardized questionnaire. Anti-HAV and anti-HEV antibodies were detected by ELISA. Statistical analysis was carried out using the Epi-Info 6.04b software, to investigate possible associations between serological markers and risk factors. Results were regarded as significant when p value < 0.05. Although a high prevalence of anti-HAV was observed (87 percent), almost 50 percent of subjects under the age of 10 were susceptible to HAV infection, an unexpected rate in endemic areas. This fact could be attributed to improvements in environmental sanitation, occurring in this area in the last years. The increasing proportion of susceptible people may result in outbreaks of HAV infection, since the virus still circulates in this area, as verified by the detection of anti-HAV IgM in some individuals. No statistical association was met between HAV infection and the risk factors here assessed. The anti-HEV IgG prevalence found in this population was 2.4 percent, consistent with the one found in non-endemic areas

Relation between hepatitis B carrier status and antibody against synthetic Plasmodium falciparum erythrocyte surface (pf155 - RESA) antigen.

A survey on Plasmodium infection was carried out in gold mine camps located in the Brazilian Amazon. Antibody against P. falciparum ring-infected erythrocyte surface antigen (RESA) was quantified by an enzyme-immunoassay in order to assess P. falciparum exposure. Hepatitis B, a common infection in this area, was also investigated by serologic markers. Among 520 sampled subjects, 517 (99.4%) admitted previous symptomatic malaria, 106 (20.4%) had positive thick smears for malaria, 82.9% had HBV markers, and 7.1% were HBsAg positive. Anti-RESA titers was significantly lower in HBV carriers than in people with resolved HBV infection suggesting that the anti-RESA immune response could be supressed by HBV carrier status. Moreover, immunedeficient responses to both infections may take place in some subjects causing concomitant lower anti-RESA response and incapacity to clear HBV.