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.

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 10(4) to 1.1 x 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.

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 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.

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.

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% (203/231) and IgM anti-HAV prevalence was 38.7% (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%) 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.

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% 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%. No total, observou-se presença de qualquer marcador de hepatite B em 106 participantes (prevalência de 23,6%; IC95%: 19,7 27,5), com 84 positivos para anti-HBs (18,7%), 100 para anti-HBc (22,3%) e apenas 3 para HBsAg (0,7%). 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.

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% 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%. Overall, the presence of any hepatitis B marker was observed in 106 participants (prevalence of 23.6%; 95% CI: 19.7 27.5), with 84 positive for anti-HBs (18.7%), 100 for anti-HBc (22.3%), and only 3 for HBsAg (0.7%). 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% 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%. No total, observou-se presença de qualquer marcador de hepatite B em 106 participantes (prevalência de 23,6%; IC95%: 19,7 27,5), com 84 positivos para anti-HBs (18,7%), 100 para anti-HBc (22,3%) e apenas 3 para HBsAg (0,7%). 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.

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% 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%. Overall, the presence of any hepatitis B marker was observed in 106 participants (prevalence of 23.6%; 95% CI: 19.7 27.5), with 84 positive for anti-HBs (18.7%), 100 for anti-HBc (22.3%), and only 3 for HBsAg (0.7%). 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.

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% (54 crianças). Seis crianças (1,2%) 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%) 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 estimate the prevalence of hepatitis B infection and to identify possible means of transmission in children from 3 to 9 years of age in Peixoto de Azevedo, a town in the Brazilian Amazon. METHODS: A cross-sectional study was carried out with 487 children enrolled in public schools and day-care centers. We obtained data concerning migration, medical history, exposure to classical risk factors for hepatitis B infection, socioeconomic factors, and the habit of sharing a bed or personal objects. We also investigated the presence in the home of persons with sexually transmitted diseases, injecting drug users, heavy alcohol users, men who had sex with other men, persons with multiple sexual partners, and a history of jaundice or hepatitis or of receiving a transfusion of blood products. Blood samples were obtained from the children to test for viral markers. RESULTS: The prevalence of hepatitis B viral markers was 11.1% (54 of 487 children). Six of the children (1.2%) tested positive for hepatitis B surface antigen. There was no association between viral markers and economic conditions, parental schooling, sanitary conditions (e.g., source of drinking water, method of wastewater disposal, method of trash disposal), number of rooms in the home, the child's history of jaundice or hepatitis, dental treatment, any accidents with needles or syringes, reusing needles or syringes, transfusion of blood products, and the habit of sharing a bed or personal objects. However, children who had contact with heavy alcohol users tended to have a higher prevalence of viral markers (P = 0.06). Among the children studied, 53 of them (10.9%) had received one or two doses of hepatitis B vaccine, but none had completed the three-dose series. Logistic regression showed that an increased prevalence of viral markers was associated with a maternal history of jaundice during the pregnancy, more than eight persons living in the home, and the mother not working in a professional occupation. CONCLUSION: Hepatitis B is probably moderately endemic in the population of Peixoto de Azevedo, with transmission mainly occurring horizontally. Such factors as being exposed in the home to persons who are heavy users of alcohol are probably important determinants of infection in the first decade of life. The low vaccination coverage promotes free circulation of the virus.

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.

A infecção pelo vírus da hepatite A constitui um dos maiores problemas de saúde pública no Brasil. A transmissão do HAV ocorre principalmente pela via orofecal deste modo a água é um importante veículo da disseminação do HAV. Existe uma alta incidência de casos agudos de hepatite A em algumas áreas do Brasil entretanto a variação sazonal destes casos não foi relatada. O objetivo deste estudo foi avaliar a sazonalidade da infecção pelo HAV no Rio de Janeiro. De janeiro de 1999 a dezembro de 2001, 1731 amostras de sangue foram coletadas no Centro Nacional de Referência para Hepatites Virais no Brasil (CRNHV). Estas amostras foram testadas por um ensaio imunoenzimático comercial para detectar anticorpos IgM anti-HAV. Taxas de positividade anuais foram de 33,74% em 1999, 32,19% em 2000, e 30,63% em 2001. Uma variação sazonal foi observada com um alto índice na primavera e verão. Também foi observado um aumento sazonal na incidência da infecção pelo HAV durante a estação chuvosa (Dezembro a Março) pois o índice de chuvas é muito alto. Foi concluído que a infecção pelo HAV ocorre o ano todo com um pico durante estações quentes com grande número de chuvas.

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% in 1999, 32.19% in 2000, and 30.63% 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.

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, Fundação 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%), almost 50% 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%, consistent with the one found in non-endemic areas.

Avaliou-se a prevalência de anticorpos para as hepatites A e E em 487 crianças de 2 a 9 anos, de um município da Amazônia matogrossense. As prevalências de anti-VHA e anti-VHE foram 86,4% (IC95% 83 -- 89,3) e 4,5% (IC95% 2,9 -- 6,9) respectivamente. A prevalência de anti-VHA já se mostrou elevada desde as faixas etárias mais baixas, distribuição típica das regiões de alta endemicidade da doença. Não se constatou associação de prevalência de marcadores das hepatites A e E com gênero, classe sócio-econômica, escolaridade dos pais, condições higiênicas, número de moradores por residência, densidade de moradores por cômodo e entre as escolas estudadas. A prevalência de hepatite E foi baixa, semelhante às encontradas entre adultos em outras regiões do Brasil.

The prevalence of antibodies to hepatitis A anti-HAV total and E viruses anti-HEV IgG was assessed in 487 children ranging from 2 to 9 years old who were students of nurseries and public schools in a county of the Amazonian region, in Mato Grosso State, Brazil. The anti-HAV and anti-HEV prevalence were 86.4% (CI 95% 83 -- 89.3) and 4.5% (CI95% 2.9 -- 6.9), respectively. The anti-HAV prevalence was high in all ages, suggesting that this is an area of high endemicity for HAV infection. There was no association between the anti-HAV markers and gender, socioeconomic level, parental educational level, hygienic conditions, number or density of residents per room or schools in which they studied. Anti-HEV prevalence was low and similar to that found among adults in another Brazilian regions.

From June 1 to July 1 1999, an outbreak involving 25 cases of hepatitis A occurred in a public school in Rio de Janeiro, Brazil. Since these cases were notified to the State Health Department, the National Reference Center for Hepatitis Viruses (CNRHV) was required to investigate the extent of hepatitis A virus (HAV) dissemination. Blood samples from all students were tested for IgM and total anti-HAV antibodies using a commercial enzyme-linked immunoassay (ELISA). At the same time, a questionnaire was completed in order to identify possible risk factors for HAV infection. The environmental investigation showed that there was no fecal contamination of the water supply. The epidemiological investigation demonstrated that almost 50% of this population was susceptible to HAV infection and probably person-to-person transmission was the principal mode of virus dissemination. In this situation, a massive vaccination campaign could control the HAV infection.

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.