RESUMO
Hepatitis A is a viral disease with a significant public health impact, especially in developing countries. Improvements in sewage services could prevent hepatitis A virus (HAV) dissemination into the environment and minimize the risk of infection. The aim of this study was to monitor HAV for one year in urban sewage samples from the largest wastewater treatment plant in Rio de Janeiro, Brazil, to assess environmental contamination with HAV and its dissemination after treatment by an activated sludge process. For this purpose, 48 samples (24 raw sewage samples and 24 treated effluent samples) were collected from August 2009 to July 2010 for HAV detection. Using quantitative real-time PCR 14 (58%) raw sewage samples were positive for HAV, and the highest viral genome loads were detected in the spring and summer. HAV was not detected in treated effluent samples, which suggests that the viral loads observed could be easily removed by the activated sludge process, thus preventing the dissemination of HAV into the environment. All of the HAV strains sequenced belonged to subgenotype IA, which clustered closely with Brazilian and Argentine HAV strains. These data demonstrate that environmental monitoring can be a useful tool in epidemiological studies.
Assuntos
Monitoramento Ambiental , Vírus da Hepatite A/isolamento & purificação , Hepatite A/transmissão , Esgotos/virologia , Brasil/epidemiologia , DNA Viral/isolamento & purificação , Monitoramento Epidemiológico , Feminino , Hepatite A/epidemiologia , Vírus da Hepatite A/genética , Humanos , Masculino , Filogenia , Saúde Pública , RNA Viral/isolamento & purificação , Reação em Cadeia da Polimerase em Tempo Real , Análise de Sequência de DNA , Saúde da População UrbanaRESUMO
Hospital wastewater has been described as an important source of spreading pathogenic microorganisms in the environment. However, there are few studies reporting the presence and concentrations of gastroenteric viruses and hepatitis A viruses in these environmental matrices. The aim of this study was to assess the contamination by viruses responsible for acute gastroenteritis and hepatitis derived from hospital wastewater treatment plants (WWTPs). Rotavirus A (RV-A), human adenoviruses (HAdV), norovirus genogroup I and II (NoV GI/GII) and hepatitis A viruses (HAV) were detected and quantified in sewage samples from two WWTPs located in Rio de Janeiro (Brazil) that operates different sewage treatments. WWTP-1 uses an Upflow Anaerobic Sludge Blanket (UASB reactor) and three serial anaerobic filters while WWTP-2 uses aerobic processes, activated sludge with extended aeration and final chlorination of the effluents. Viruses' detection was investigated by using conventional PCR/RT-PCR, quantitative real-time PCR (qPCR) and partial sequencing of the genome of the viruses detected. Rate of viruses detection ranged from 7% (NoV GI in WWTP-1) to 95% (RV-A in WWTP-2) and genome from all viruses were detected. The most prevalent genotypes were RV-A SG I, HAdV species D and F, NoV GII/4 and HAV subgenotype IA. Mean values of viral loads (genome copies (GC)/ml) obtained in filtered effluents from anaerobic process was 1.9 × 10(3) (RV-A), 2.8 × 10(3) (HAdV) and 2.4 × 10(3) (NoV GII). For chlorinated effluents from activated sludge process, the mean values of viral loads (GC/ml) was 1.2 × 10(5) (RV-A), 1.4 × 10(3) (HAdV), 8.1 × 10(2) (NoV GII) and 2.8 × 10(4) (HAV). Data on viral detection in treated effluents of hospital WWTPs confirmed the potential for environmental contamination by viruses and could be useful to establish standards for policies on wastewater management.
Assuntos
Enterovirus/genética , Hospitais , Esgotos/virologia , Eliminação de Resíduos Líquidos , Anaerobiose , Enterovirus/isolamento & purificação , Reação em Cadeia da Polimerase , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
This work studied the replication sites of hepatitis A virus (HAV) in cynomolgus monkeys (Macaca fascicularis) after intravenous inoculation. The cynomolgus monkeys were inoculated with the Brazilian hepatitis A virus strain (HAF-203). Monkeys were euthanized on days 15, 30, 45 and 60 postinoculation (pi). Liver samples, submandibular salivary gland, mesenteric lymph node and tonsils were removed for virological and pathological evaluation. Immunofluorescence analyses on liver and salivary gland sections using confocal laser scanning microscopy revealed the presence of HAV antigen (HAV Ag). The presence of HAV genome was monitored by real-time PCR. The HAV RNA was detected at 7 days postinoculation (dpi), concomitantly in serum, saliva and faeces. The highest HAV viral load was observed in faeces at 15 dpi (10(5) copies/ml), followed by serum viral load of 10(4) copies/ml at 20 dpi and saliva viral load of 10(3 )copies/ml at 7 dpi. The animals showed first histological and biochemical signs of hepatitis at 15 dpi. The HAV antigen (Ag) was present from day 7 until day 60 pi in the liver and salivary glands. The HAV replicative intermediate was also detected in the liver (4.5 x 10(4) copies/mg), salivary glands (1.9 x 10(3) copies/mg), tonsils (4.2 x 10(1) copies/mg) and lymph nodes (3.4 x 10(1) copies/mg). Our data demonstrated that the salivary gland as an extrahepatic site of early HAV replication could create a potential risk of saliva transmitted infection. In addition, the cynomolgus monkey was confirmed as a suitable model to study the pathogenesis of HAV human infection.
Assuntos
Vírus da Hepatite A/patogenicidade , Hepatite A/diagnóstico , Replicação Viral , Alanina Transaminase/sangue , Animais , Modelos Animais de Doenças , Fezes/virologia , Imunofluorescência , Hepatite A/patologia , Hepatite A/transmissão , Anticorpos Anti-Hepatite A/sangue , Antígenos da Hepatite A/isolamento & purificação , Vírus da Hepatite A/genética , Vírus da Hepatite A/imunologia , Injeções Intravenosas , Fígado/enzimologia , Fígado/virologia , Linfonodos/virologia , Macaca fascicularis , Masculino , Microscopia Confocal , Tonsila Palatina/virologia , RNA Viral/sangue , RNA Viral/isolamento & purificação , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Saliva/virologia , Glândulas Salivares/virologia , Fatores de Tempo , Carga ViralRESUMO
The hepatitis A virus (HAV) HAF-203 strain was isolated from an acute case of HAV infection. The primary isolation of HAF-203 in Brazil and its adaptation to the FRhK-4 cell lineage allowed the production of large amounts of viral particles enabling molecular characterization of the first HAV isolate in Brazil. The aim of our study was to determine the nucleotide sequence of the HAF-203 strain genome, compare it to other HAV genomes and highlight its genetic variability. The complete nucleotide sequence of the HAF-203 strain (7472 nucleotides) was compared to those obtained earlier by others for other HAV isolates. These analyses revealed 19 HAF-specific nucleotide sequence differences with 10 amino acid substitutions. Most of the non-conservative changes were located at VP1, 2C, and 3D genes, but the 3B region was the most variable. The availability of HAF-203 complementary DNA was useful for the production of the recombinant VP1 protein, which is a major determinant of viral infectivity. This recombinant protein was shown by enzyme-linked immunoassay and blotting, to be immunogenic and resemble the native protein, therefore suggesting its value as a reagent for incorporation into diagnostic tests.
Assuntos
Escherichia coli/genética , Variação Genética , Vírus da Hepatite A/genética , Proteínas Estruturais Virais/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Brasil , Expressão Gênica , Genoma Viral , Vírus da Hepatite A/imunologia , Humanos , Immunoblotting , Dados de Sequência Molecular , Filogenia , Reação em Cadeia da Polimerase , RNA Viral , CoelhosRESUMO
The hepatitis A virus (HAV) HAF-203 strain was isolated from an acute case of HAV infection. The primary isolation of HAF-203 in Brazil and its adaptation to the FRhK-4 cell lineage allowed the production of large amounts of viral particles enabling molecular characterization of the first HAV isolate in Brazil. The aim of our study was to determine the nucleotide sequence of the HAF-203 strain genome, compare it to other HAV genomes and highlight its genetic variability. The complete nucleotide sequence of the HAF-203 strain (7472 nucleotides) was compared to those obtained earlier by others for other HAV isolates. These analyses revealed 19 HAF-specific nucleotide sequence differences with 10 amino acid substitutions. Most of the non-conservative changes were located at VP1, 2C, and 3D genes, but the 3B region was the most variable. The availability of HAF-203 complementary DNA was useful for the production of the recombinant VP1 protein, which is a major determinant of viral infectivity. This recombinant protein was shown by enzyme-linked immunoassay and blotting, to be immunogenic and resemble the native protein, therefore suggesting its value as a reagent for incorporation into diagnostic tests.
Assuntos
Humanos , Animais , Coelhos , Variação Genética , Vírus da Hepatite A/genética , Proteínas Estruturais Virais , Sequência de Aminoácidos , Sequência de Bases , Brasil , Escherichia coli/genética , Expressão Gênica , Genoma Viral , Vírus da Hepatite A/imunologia , Immunoblotting , Dados de Sequência Molecular , Filogenia , Reação em Cadeia da Polimerase , RNA ViralRESUMO
A cross-sectional study was conducted in order to identify hepatitis A virus (HAV) serological markers in 418 individuals (mean age, 16.4 years; range, 1 month-80 years) at a public child care center in Rio de Janeiro, Brazil, as well as to analyze risk factors and determine circulating genotypes. Serum samples were tested using an enzyme immunoassay. Reverse transcription polymerase chain reaction (RT-PCR) was used to detect and characterize HAV RNA, and sequencing was performed. Anti-HAV antibodies and IgM anti-HAV antibodies were detected, respectively, in 89.5% (374/418) and 10.5% (44/418) of the individuals tested. Acute HAV infection in children was independently correlated with crawling (p < 0.05). In 56.8% (25/44) of the IgM anti-HAV-positive individuals and in 33.3% (5/15) of the IgM anti-HAV-negative individuals presenting clinical symptoms, HAV RNA was detected. Phylogenetic analysis revealed co-circulation of subgenotypes IA and IB in 93.3% (28/30) of the amplified samples. In present study, we verify that 79% (30/38) of children IgM anti-HAV-positive were asymptomatic. In child care centers, this asymptomatic spread is a more serious problem, promoting the infection of young children, who rarely show signs of infection. Therefore, vaccinating children below the age of two might prevent the asymptomatic spread of hepatitis A.
Assuntos
Creches/estatística & dados numéricos , Vírus da Hepatite A/imunologia , Hepatite A/epidemiologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Brasil/epidemiologia , Criança , Pré-Escolar , Estudos Transversais , Feminino , Hepatite A/diagnóstico , Anticorpos Anti-Hepatite A/sangue , Vacinas contra Hepatite A , Vírus da Hepatite A/genética , Humanos , Imunoglobulina M/sangue , Lactente , Recém-Nascido , Masculino , Pessoa de Meia-Idade , Prevalência , RNA Viral/análise , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
The prevalence of hepatitis A virus (HAV) infection is high in developing countries, in which low standards of sanitation promote the transmission of the virus. In Latin America, which is considered an area of high HAV endemicity, most HAV-positive individuals are infected in early childhood However recent studies have shown that prevalence rates are decreasing. Herein, we review the data on HAV prevalence and outbreaks available in scientific databases. We also use official government data in order to evaluate mortality rates in Brazil over the last two decades. Studies conducted in the northernmost regions of Brazil have indicated that, although improved hygiene has led to a reduction in childhood exposure to HAV, the greatest exposure still occurs early in life. In the Southeastern region, the persistence of circulating HAV has generated outbreaks among individuals of low socioeconomic status, despite adequate sanitation. Nationwide, hepatitis A mortality rates declined progressively from 1980 to 2002. During that period, mortality rates in the Northern region consistently exceeded the mean national rate and those for other regions. Excluding the North, the rates in all regions were comparable. Nevertheless, the trend toward decline observed in the South was paralleled by a similar trend in the North.
Assuntos
Surtos de Doenças , Doenças Endêmicas , Hepatite A/mortalidade , Adolescente , Adulto , Idoso , Brasil/epidemiologia , Criança , Pré-Escolar , Hepatite A/epidemiologia , Hepatite A/prevenção & controle , Humanos , Incidência , Lactente , Recém-Nascido , Pessoa de Meia-Idade , Prevalência , Fatores SocioeconômicosRESUMO
In this report, we examine the adaptability of commercially available serological kits to detect antibodies markers for viral hepatitis in oral fluid samples. We also assessed the prevalence of hepatitis A, B, and C virus-specific antibodies, and related risk factors for these infectious diseases through sensitivity of the tests in saliva samples to evaluate if oralfluid can be an alternative tool to substitute serum in diagnosis of acute viral hepatitis and in epidemiological studies. One hundred and ten paired serum and saliva specimens from suspect patients of having acute hepatitis were collected to detect antibodies to hepatitis A (total and IgM), hepatitis B (anti-HBs, total anti-HBc and IgM anti-HBc), and hepatitis C (anti-HCV) using commercially available enzyme-linked immunossorbent assay (EIA). In relation to serum samples, oral fluid assay sensitivity and specificity were as follows: 87 and 100% for total anti-HAV, 79 and 100% for anti-HAVIgM, 6 and 95% for anti-HBs, 13 and 100%for total anti-HBc, 100 and 100% for anti-HBc IgM, and 75 and 100% for anti-HCV The consistency observed between antibodies tests in saliva and expected risk factors for hepatitis A and C suggests that the saliva method could replace serum in epidemiological studies for hepatitis A and C.
Assuntos
Hepacivirus/imunologia , Vírus da Hepatite A/imunologia , Anticorpos Anti-Hepatite/análise , Vírus da Hepatite B/imunologia , Hepatite Viral Humana/diagnóstico , Saliva/virologia , Doença Aguda , Adolescente , Adulto , Idoso , Biomarcadores/análise , Brasil/epidemiologia , Criança , Ensaio de Imunoadsorção Enzimática , Métodos Epidemiológicos , Feminino , Hepatite A/diagnóstico , Hepatite B/diagnóstico , Hepatite C/diagnóstico , Humanos , Masculino , Pessoa de Meia-Idade , Kit de Reagentes para DiagnósticoRESUMO
A cross-sectional study was conducted in order to identify hepatitis A virus (HAV) serological markers in 418 individuals (mean age, 16.4 years; range, 1 month-80 years) at a public child care center in Rio de Janeiro, Brazil, as well as to analyze risk factors and determine circulating genotypes. Serum samples were tested using an enzyme immunoassay. Reverse transcription polymerase chain reaction (RT-PCR) was used to detect and characterize HAV RNA, and sequencing was performed. Anti-HAV antibodies and IgM anti-HAV antibodies were detected, respectively, in 89.5 percent (374/418) and 10.5 percent (44/418) of the individuals tested. Acute HAV infection in children was independently correlated with crawling (p < 0.05). In 56.8 percent (25/44) of the IgM anti-HAV-positive individuals and in 33.3 percent (5/15) of the IgM anti-HAV-negative individuals presenting clinical symptoms, HAV RNA was detected. Phylogenetic analysis revealed co-circulation of subgenotypes IA and IB in 93.3 percent (28/30) of the amplified samples. In present study, we verify that 79 percent (30/38) of children IgM anti-HAV-positive were asymptomatic. In child care centers, this asymptomatic spread is a more serious problem, promoting the infection of young children, who rarely show signs of infection. Therefore, vaccinating children below the age of two might prevent the asymptomatic spread of hepatitis A.
Assuntos
Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Criança , Pré-Escolar , Feminino , Humanos , Lactente , Recém-Nascido , Masculino , Pessoa de Meia-Idade , Creches/estatística & dados numéricos , Vírus da Hepatite A/imunologia , Hepatite A/epidemiologia , Brasil/epidemiologia , Estudos Transversais , Vacinas contra Hepatite A , Anticorpos Anti-Hepatite A/sangue , Vírus da Hepatite A/genética , Hepatite A/diagnóstico , Imunoglobulina M/sangue , Prevalência , Reação em Cadeia da Polimerase Via Transcriptase Reversa , RNA Viral/análiseRESUMO
The prevalence of hepatitis A virus (HAV) infection is high in developing countries, in which low standards of sanitation promote the transmission of the virus. In Latin America, which is considered an area of high HAV endemicity, most HAV-positive individuals are infected in early childhood. However, recent studies have shown that prevalence rates are decreasing. Herein, we review the data on HAV prevalence and outbreaks available in scientific databases. We also use official government data in order to evaluate mortality rates in Brazil over the last two decades. Studies conducted in the northernmost regions of Brazil have indicated that, although improved hygiene has led to a reduction in childhood exposure to HAV, the greatest exposure still occurs early in life. In the Southeastern region, the persistence of circulating HAV has generated outbreaks among individuals of low socioeconomic status, despite adequate sanitation. Nationwide, hepatitis A mortality rates declined progressively from 1980 to 2002. During that period, mortality rates in the Northern region consistently exceeded the mean national rate and those for other regions. Excluding the North, the rates in all regions were comparable. Nevertheless, the trend toward decline observed in the South was paralleled by a similar trend in the North.