RESUMO
Hepatitis B virus (HBV) DNA and RNA were quantified by digital PCR assays in 20-30 tissue pieces from each of 4 liver explants with cirrhosis caused by HBV. The within-patient variability of HBV RNA levels between pieces was up to a 1000-fold. Core RNA and S RNA levels were similar and correlated strongly when replication was high, supporting that transcription was from covalently closed circular DNA (cccDNA). By contrast, enhanced expression of S RNA relative to cccDNA and core RNA in patients with medium-high or low replication supports that HBV surface antigen (HBsAg) can be expressed mainly from integrated HBV DNA in such patients.
Assuntos
Hepatite B Crônica , Hepatite B , Antígenos de Superfície , DNA Circular/genética , DNA Viral/análise , Antígenos de Superfície da Hepatite B/metabolismo , Vírus da Hepatite B/genética , Humanos , Fígado , RNA Viral/análiseRESUMO
BACKGROUND: Hepatitis B virus (HBV) integration has implications for cancer development and surface antigen (HBsAg) production, but methods to quantify integrations are lacking. The aim of this study was to develop a droplet digital PCR (ddPCR) assay discriminating between circular and integrated HBV DNA, and to relate the distribution between the two forms to other HBV markers. METHODS: ddPCR with primers spanning the typical linearization breakpoint in the HBV genome allowed for quantification of the absolute copy numbers of total and circular HBV DNA, and calculation of linear HBV DNA. RESULTS: Analysis of 70 liver biopsies from patients with chronic HBV infection revealed that the fraction of linear HBV DNA, which includes integrations, was higher in HBeAg-negative patients than HBeAg-positive. The ratio between HBsAg and HBV DNA levels in serum correlated with the intrahepatic proportion of linear HBV DNA. Furthermore, ddPCR experiments on serum samples and experiments with nuclease indicated the contribution of encapsidated double-stranded linear DNA and replication intermediates to be limited. CONCLUSIONS: The degree of integration of intrahepatic HBV DNA in the HBeAg-negative stage may be higher than previously anticipated, and integrated DNA may explain the persistence of high HBsAg serum levels in patients with low HBV DNA levels.
Assuntos
Hepatite B Crônica , Hepatite B , DNA Circular/genética , DNA Viral , Antígenos de Superfície da Hepatite B , Antígenos E da Hepatite B , Vírus da Hepatite B/genética , Humanos , FígadoRESUMO
The family Hepeviridae includes enterically transmitted small quasi-enveloped or non-enveloped positive-sense single-stranded RNA viruses infecting mammals and birds (subfamily Orthohepevirinae) or fish (Parahepevirinae). Hepatitis E virus (genus Paslahepevirus) is responsible for self-limiting acute hepatitis in humans; the infection may become chronic in immunocompromised individuals and extrahepatic manifestations have been described. Avian hepatitis E virus (genus Avihepevirus) causes hepatitis-splenomegaly syndrome in chickens. This is a summary of the International Committee on Taxonomy of Viruses (ICTV) Report on the family Hepeviridae, which is available at www.ictv.global/report/hepeviridae.
Assuntos
Hepevirus , Vírus de RNA , Animais , Galinhas , Peixes , Genoma Viral , Hepevirus/genética , Humanos , Mamíferos , Vírus de RNA/genética , Vírion , Replicação ViralRESUMO
Respiratory infections are often caused by enteroviruses (EVs). The aim of this study was to identify whether certain types of EV were more likely to cause severe illness in 2016, when an increasing spread of upper respiratory infections was observed in Gothenburg, Sweden. The EV strain in 137 of 1341 nasopharyngeal samples reactive for EV by polymerase chain reaction could be typed by sequencing the viral 5'-untranslated region and VP1 regions. Phylogenetic trees were constructed. Patient records were reviewed. Hospital care was needed for 46 of 74 patients with available medical records. The majority of the patients (83) were infected with the rhinovirus (RV). The remaining 54 were infected with EV A, B, C, and D strains of 13 different types, with EV-D68 and CV-A10 being the most common (17 vs. 14). Significantly more patients with EV-D68 presented with dyspnea, both when compared with other EV types (p = 0.003) and compared to all other EV and RV infections (p = 0.04). Phylogenetic analysis of the sequences revealed the spread of both Asian and European CV-A10 strains and 12 different RV C types. This study showed an abundance of different EV types spreading during a year with increased upper respiratory increased infections. EV-D68 infections were associated with more severe disease manifestation. Other EV and RV types were more evenly distributed between hospitalized and nonhospitalized patients. The EV type CV-A10 was also found in infected patients, which warrants further studies and surveillance, as this pathogen could cause more severe disease and outbreaks of hand, foot, and mouth disease.
Assuntos
Enterovirus Humano D , Infecções por Enterovirus , Enterovirus , Infecções Respiratórias , Surtos de Doenças , Enterovirus/genética , Humanos , Lactente , Filogenia , Rhinovirus/genéticaRESUMO
The family Hepadnaviridae comprises small enveloped viruses with a partially double-stranded DNA genome of 3.0-3.4 kb. All family members express three sets of proteins (preC/C, polymerase and preS/S) and replication involves reverse transcription within nucleocapsids in the cytoplasm of hepatocytes. Hepadnaviruses are hepatotropic and infections may be transient or persistent. There are five genera: Parahepadnavirus, Metahepadnavirus, Herpetohepadnavirus, Avihepadnavirus and Orthohepadnavirus. This is a summary of the International Committee on Taxonomy of Viruses (ICTV) Report on the family Hepadnaviridae, which is available at ictv.global/report/hepadnaviridae.
Assuntos
Hepadnaviridae/classificação , Hepadnaviridae/genética , Citoplasma/virologia , Genoma Viral/genética , Hepatócitos/virologia , Humanos , Replicação Viral/genéticaRESUMO
In this recommendation, we update our 2016 table of reference sequences of subtypes of hepatitis E virus (HEV; species Orthohepevirus A, family Hepeviridae) for which complete genome sequences are available (Smith et al., 2016). This takes into account subsequent publications describing novel viruses and additional proposals for subtype names; there are now eight genotypes and 36 subtypes. Although it remains difficult to define strict criteria for distinguishing between virus subtypes, and is not within the remit of the International Committee on Taxonomy of Viruses (ICTV), the use of agreed reference sequences will bring clarity and stability to researchers, epidemiologists and clinicians working with HEV.
Assuntos
Vírus da Hepatite E/classificação , Vírus da Hepatite E/genética , Animais , Sequência de Bases , Bases de Dados de Ácidos Nucleicos , Genótipo , Hepatite E/virologia , Vírus da Hepatite E/isolamento & purificação , Humanos , Filogenia , RNA Viral/genética , Especificidade da EspécieRESUMO
Influent wastewater and effluent wastewater at the Rya treatment plant in Gothenburg, Sweden, were continuously monitored for enteric viruses by quantitative PCR (qPCR) during 1 year. Viruses in effluent wastewater were also identified by next-generation sequencing (NGS) in samples collected during spring, early summer, and winter. Samples of incoming wastewater were collected every second week. Seasonal variations in viral concentrations in incoming wastewater were found for noroviruses GII, sapovirus, rotavirus, parechovirus, and astrovirus. Norovirus GI and GIV and Aichi virus were present in various amounts during most weeks throughout the year, while hepatitis A virus, enterovirus, and adenovirus were identified less frequently. Fluctuations in viral concentrations in incoming wastewater were related to the number of diagnosed patients. The viruses were also detected in treated wastewater, however, with a 3- to 6-log10 reduction in concentration. Seven different hepatitis E virus (HEV) strains were identified in the effluents. Five of these strains belonged to genotype 3 and have been isolated in Sweden from swine, wild boars, and humans and in drinking water. The other two strains were divergent and had not been identified previously. They were similar to strains infecting rats and humans. Surveillance of enteric viruses in wastewater is a tool for early detection and follow-up of gastroenteritis outbreaks in society and for the identification of new viruses that can cause infection in humans.IMPORTANCE Both influent wastewater and treated wastewater at a wastewater treatment plant (WWTP) contain a high variety of human viral pathogens with seasonal variability when followed for 1 year. The peak of the amount of 11 different viruses in the inlet wastewater preceded the peak of the number of diagnosed patients by 2 to 4 weeks. The treatment of wastewater reduced viral concentrations by 3 to 6 log10 Despite the treatment of wastewater, up to 5 log10 virus particles per liter were released from into the surrounding river. Hepatitis E virus (HEV) strains previously identified in drinking water and two new strains, similar to those infecting rats and humans, were identified in the treated wastewater released from the WWTP.
Assuntos
Metagenoma , Vírus/isolamento & purificação , Águas Residuárias/virologia , Metagenômica , Reação em Cadeia da Polimerase em Tempo Real , Estações do Ano , Fenômenos Fisiológicos Virais , Vírus/classificação , Vírus/genéticaRESUMO
Hepatitis B virus (HBV) is a major cause of hepatocellular carcinoma (HCC). Integration of HBV DNA into the human genome may contribute to oncogenesis and to the production of the hepatitis B surface antigen (HBsAg). Whether integrations contribute to HBsAg levels in the blood is poorly known. Here, we characterize the HBV RNA profile of HBV integrations in liver tissue in patients with chronic HBV infection, with or without concurrent hepatitis D infection, by transcriptome deep sequencing. Transcriptomes were determined in liver tissue by deep sequencing providing 200 million reads per sample. Integration points were identified using a bioinformatic pipeline. Explanted liver tissue from five patients with end-stage liver disease caused by HBV or HBV/HDV was studied along with publicly available transcriptomes from 21 patients. Almost all HBV RNA profiles were devoid of reads in the core and the 3' redundancy (nt 1830-1927) regions, and contained a large number of chimeric viral/human reads. Hence, HBV transcripts from integrated HBV DNA rather than from covalently closed circular HBV DNA (cccDNA) predominated in late-stage HBV infection, in particular in cases with hepatitis D virus co-infection. The findings support the suggestion that integrated HBV DNA can be a significant source of HBsAg in humans.
Assuntos
Carcinoma Hepatocelular , DNA Viral , Vírus da Hepatite B , Hepatite B Crônica , Hepatite B , Sequenciamento de Nucleotídeos em Larga Escala , Neoplasias Hepáticas , Antígenos de Superfície da Hepatite B , Vírus da Hepatite B/genética , Humanos , Fígado , TranscriptomaRESUMO
A third of humans carry genetic variants of the ITP pyrophosphatase (ITPase) gene (ITPA) that lead to reduced enzyme activity. Reduced ITPase activity was earlier reported to protect against ribavirin-induced hemolytic anemia and to diminish relapse following ribavirin and interferon therapy for hepatitis C virus (HCV) genotype 2 or 3 infections. While several hypotheses have been put forward to explain the antiviral actions of ribavirin, details regarding the mechanisms of interaction between reduced ITPase activity and ribavirin remain unclear. The in vitro effect of reduced ITPase activity was assessed by means of transfection of hepatocytes (Huh7.5 cells) with a small interfering RNA (siRNA) directed against ITPA or a negative-control siRNA in the presence or absence of ribavirin in an HCV culture system. Low ribavirin concentrations strikingly depleted intracellular GTP levels in HCV-infected hepatocytes whereas higher ribavirin concentrations induced G-to-A and C-to-U single nucleotide substitutions in the HCV genome, with an ensuing reduction of HCV RNA expression and HCV core antigen production. Ribavirin triphosphate (RTP) was dephosphorylated in vitro by recombinant ITPase to a similar extent as ITP, a naturally occurring substrate of ITPase, and reducing ITPA expression in Huh 7.5 cells by siRNA increased intracellular levels of RTP in addition to increasing HCV mutagenesis and reducing progeny virus production. Our results extend the understanding of the biological impact of reduced ITPase activity, demonstrate that RTP is a substrate of ITPase, and may point to personalized ribavirin dosage according to ITPA genotype in addition to novel antiviral strategies.IMPORTANCE This study highlights the multiple modes of action of ribavirin, including depletion of intracellular GTP and increased hepatitis C virus mutagenesis. In cell culture, reduced ITP pyrophosphatase (ITPase) enzyme activity affected the intracellular concentrations of ribavirin triphosphate (RTP) and augmented the impact of ribavirin on the mutation rate and virus production. Additionally, our results imply that RTP, similar to ITP, a naturally occurring substrate of ITPase, is dephosphorylated in vitro by ITPase.
Assuntos
Antivirais/farmacologia , Hepacivirus/efeitos dos fármacos , Mutagênese , Pirofosfatases/genética , Ribavirina/farmacologia , Antivirais/metabolismo , Linhagem Celular Tumoral , Relação Dose-Resposta a Droga , Regulação da Expressão Gênica , Guanosina Trifosfato/metabolismo , Hepacivirus/genética , Hepacivirus/crescimento & desenvolvimento , Hepacivirus/metabolismo , Antígenos do Núcleo do Vírus da Hepatite B/genética , Antígenos do Núcleo do Vírus da Hepatite B/metabolismo , Hepatócitos/efeitos dos fármacos , Hepatócitos/enzimologia , Hepatócitos/virologia , Sequenciamento de Nucleotídeos em Larga Escala , Interações Hospedeiro-Patógeno , Humanos , Nucleotídeos/metabolismo , Pirofosfatases/antagonistas & inibidores , Pirofosfatases/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , RNA Viral/genética , RNA Viral/metabolismo , Ribavirina/metabolismo , Transdução de SinaisRESUMO
Objective: Hepatitis E virus (HEV) genotype 3 is endemic in Northern Europe and despite a high seroprevalence of anti-HEV IgG antibodies among blood donors (≈17%), few clinical cases are notified in Sweden. Low awareness of hepatitis E and its possible symptoms may contribute to this discrepancy. The aim of this study was to investigate the prevalence of acute HEV infection among hospital admitted patients with abdominal pain and elevated liver enzymes.Materials and methods: During 2016-2017, 148 adult patients with serum alanine aminotransferase (ALT) or aspartate aminotransferase (AST) > twice normal levels were prospectively enrolled at surgical wards at three Swedish hospitals. Serum samples were analyzed for HEV RNA as well as anti-HEV IgM and IgG, and medical records were reviewed.Results: Six (6/148, 4.1%) patients were HEV infected confirmed by detectable HEV RNA, but only one of these patients had detectable anti-HEV antibodies. Four of the HEV infected patients were diagnosed with gallstone-related disease: three with biliary pancreatitis and one with biliary colic. The remaining two were diagnosed with bowel obstruction and pancreatic malignancy. Four HEV strains were typed by sequencing to genotype 3.Conclusions: This study identified acute HEV3 infection in 4% of the patients with elevated liver enzymes admitted to a surgical ward. HEV infection was not the solitary disease leading to hospitalization, instead it was found to be associated with other surgical conditions such as gallstone-related disease including biliary pancreatitis. Additionally, HEV RNA might be the preferential diagnostic tool for detecting ongoing HEV infection.
Assuntos
Cólica/virologia , Cálculos Biliares/virologia , Genótipo , Vírus da Hepatite E/genética , Hepatite E/diagnóstico , Pancreatite/virologia , Doença Aguda , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Cólica/diagnóstico , Feminino , Seguimentos , Cálculos Biliares/diagnóstico , Hepatite E/complicações , Hepatite E/epidemiologia , Hepatite E/virologia , Vírus da Hepatite E/isolamento & purificação , Humanos , Masculino , Pessoa de Meia-Idade , Pancreatite/diagnóstico , Prevalência , Estudos Prospectivos , RNA Viral/análise , Suécia , Adulto JovemRESUMO
Hepatitis delta virus, the only member of the only species in the genus Deltavirus, is a unique human pathogen. Its ~1.7 kb circular negative-sense RNA genome encodes a protein, hepatitis delta antigen, which occurs in two forms, small and large, both with unique functions. Hepatitis delta virus uses host RNA polymerase II to replicate via double rolling circle RNA synthesis. Newly synthesized linear RNAs are circularized after autocatalytic cleavage and ligation. Hepatitis delta virus requires the envelope of the helper virus, hepatitis B virus (family Hepadnaviridae), to produce infectious particles. This is a summary of the International Committee on Taxonomy of Viruses (ICTV) Report on the taxonomy of Deltavirus which is available at www.ictv.global/report/deltavirus.
Assuntos
Hepatite D/virologia , Vírus Delta da Hepatite/classificação , Vírus Delta da Hepatite/genética , RNA Viral/genética , Genoma Viral , Vírus Auxiliares/fisiologia , Vírus da Hepatite B/fisiologia , Vírus Delta da Hepatite/crescimento & desenvolvimento , Interações Hospedeiro-Patógeno , Humanos , RNA/genética , RNA/metabolismo , RNA Polimerase II/metabolismo , RNA Circular , RNA Viral/metabolismo , Proteínas do Envelope Viral/metabolismo , Replicação ViralRESUMO
Seroprevalence studies provide information on the susceptibility to infection of certain populations, including women of childbearing age. Such data from Central Africa are scarce regarding two viruses that cause congenital infections: Zika virus (ZIKV), an emerging mosquito-borne infection, and Rubella virus (RuV), a vaccine-preventable infection. We report on the seroprevalence of both ZIKV and RuV from Rwanda, a country without any known cases of ZIKV, but bordering Uganda where this virus was isolated in 1947. Anti-ZIKV-specific and anti-RuV-specific immunoglobulin G (IgG) antibodies were analyzed by enzyme-linked immunosorbent assay (ELISA) in serum samples from 874 Rwandan and 215 Swedish blood donors. Samples positive for IgG antibodies against ZIKV were examined for viral RNA using real-time reverse transcription polymerase chain reaction (RT-qPCR). The seroprevalence of ZIKV IgG in Rwanda was 1.4% (12/874), of which the predominance of positive findings came from the Southeastern region. All anti-ZIKV IgG-positive samples were PCR-negative. Among 297 female blood donors of childbearing age, 295 (99.3%) were seronegative and thus susceptible to ZIKV. All Swedish blood donors were IgG-negative to ZIKV. In contrast, blood donors from both countries showed high seroprevalence of IgG to RuV: 91.2% for Rwandan and 92.1% for Swedish donors. Only 10.5% (31/294) of female donors of childbearing age from Rwanda were seronegative for RuV. In Rwanda, seroprevalence for ZIKV IgG antibodies was low, but high for RuV. Hence, women of childbearing age were susceptible to ZIKV. These data may be of value for decision-making regarding prophylactic measures.
Assuntos
Anticorpos Antivirais/sangue , Vírus da Rubéola/imunologia , Rubéola (Sarampo Alemão)/epidemiologia , Infecção por Zika virus/epidemiologia , Zika virus/imunologia , Adolescente , Adulto , Idoso , Doadores de Sangue , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Imunoglobulina G/sangue , Masculino , Pessoa de Meia-Idade , RNA Viral/sangue , Reação em Cadeia da Polimerase em Tempo Real , Ruanda/epidemiologia , Estudos Soroepidemiológicos , Suécia/epidemiologia , Adulto JovemRESUMO
BACKGROUND: Hepatocytes infected by hepatitis B virus (HBV) produce different HBV RNA species, including pregenomic RNA (pgRNA), which is reverse transcribed during replication. Particles containing HBV RNA are present in serum of infected individuals, and quantification of this HBV RNA could be clinically useful. METHODS: In a retrospective study of 95 patients with chronic HBV infection, we characterised HBV RNA in serum in terms of concentration, particle association and sequence. HBV RNA was detected by real-time PCR at levels almost as high as HBV DNA. RESULTS: The HBV RNA was protected from RNase and it was found in particles of similar density as particles containing HBV DNA after fractionation on a Nycodenz gradient. Sequencing the epsilon region of the RNA did not reveal mutations that would preclude its binding to the viral polymerase before encapsidation. Specific quantification of precore RNA and pgRNA by digital PCR showed almost seven times lower ratio of precore RNA/pgRNA in serum than in liver tissue, which corresponds to poorer encapsidation of this RNA as compared with pgRNA. The serum ratio between HBV DNA and HBV RNA was higher in genotype D as compared with other genotypes. CONCLUSIONS: The results suggest that HBV RNA in serum is present in viral particles with failing reverse transcription activity, which are produced at almost as high rates as viral particles containing DNA. The results encourage further studies of the mechanisms by which these particles are produced, the impact of genotype, and the potential clinical utility of quantifying HBV RNA in serum.
Assuntos
DNA Viral/genética , Vírus da Hepatite B/genética , Hepatite B Crônica/virologia , RNA Viral/genética , Transcrição Reversa , Vírion/genética , Adulto , Sequência de Bases , DNA Viral/sangue , Feminino , Expressão Gênica , Genótipo , Antígenos do Núcleo do Vírus da Hepatite B/genética , Antígenos do Núcleo do Vírus da Hepatite B/metabolismo , Antígenos de Superfície da Hepatite B/genética , Antígenos de Superfície da Hepatite B/metabolismo , Vírus da Hepatite B/classificação , Vírus da Hepatite B/isolamento & purificação , Vírus da Hepatite B/metabolismo , Hepatite B Crônica/sangue , Hepatócitos/virologia , Humanos , Fígado/virologia , Masculino , RNA Viral/sangue , RNA Viral/classificação , Estudos Retrospectivos , Vírion/metabolismo , Replicação ViralRESUMO
Hepatitis E virus (HEV) is a pathogen that causes hepatitis worldwide. Molecular studies have identified HEV RNA in blood products although its significance is not understood. This study was undertaken to characterize HEV genomes in asymptomatic plasma donors from Sweden and Germany lacking anti-HEV. Complete open reading frames (ORFs) were obtained from HEV strains in 5 out of 18 plasma donors who tested positive for HEV RNA. All strains had CUG as the start codon of ORF3, while 147 GenBank strains all had AUG as the start codon (p < 0.0001). This substitution was found in both interrelated and unrelated strains belonging to different phylogenetic clades. The HEV strains from the seronegative plasma donors had no other substitution in common, which may be why the CUG substitution seems to explain the seronegativity.
Assuntos
Doadores de Sangue , Genoma Viral , Vírus da Hepatite E/classificação , Vírus da Hepatite E/genética , Hepatite E/virologia , RNA Viral , Proteínas Virais/genética , Sequência de Aminoácidos , Códon de Iniciação , Genótipo , Vírus da Hepatite E/imunologia , Humanos , Imunoglobulina G/imunologia , Imunoglobulina M/imunologia , Fases de Leitura Aberta , Filogenia , Proteínas Virais/químicaRESUMO
BACKGROUND: Hepatitis C virus (HCV) is the leading cause of severe liver disease worldwide and is highly endemic in Africa, where it often has nosocomial spread. Little is known on the HCV prevalence, risk for transfusion-transmitted HCV, and circulating genotypes in Rwanda. This study was performed to investigate the prevalence of anti-HCV among blood donors from all regions of the country and genetically characterize identified HCV strains. STUDY DESIGN AND METHODS: Data on anti-HCV reactivity for all 45,061 Rwandan blood donations during 2014 were compiled. Samples from 720 blood donors were reanalyzed for anti-HCV in Sweden. Line immunoassay INNO-LIA HCV and detection of HCV RNA by polymerase chain reaction were used to confirm anti-HCV reactivity. The NS5B and core regions were sequenced and phylogenetic analysis was performed. RESULTS: The anti-HCV prevalence among all first-time blood donors was 1.6%, with the highest occurrence in donors from the eastern region. On further analysis, only 25 of 120 primarily anti-HCV-reactive samples could be confirmed reactive and 15 samples had indeterminate results by INNO-LIA. Confirmed reactivity was more common among females than males (p = 0.03) with no regional difference. Phylogenetic analysis of the sequences showed a predominance of subtypes 4k, 4q, and 4r, with no geographical difference in their distribution. CONCLUSION: The prevalence of anti-HCV among Rwandan blood donors has probably been overestimated previously due to the high rate of nonconfirmable anti-HCV reactivity. Further study of the involved mechanism is needed to avoid loss of blood products and distress for blood donors and other test recipients.
Assuntos
Doadores de Sangue , Hepacivirus/genética , Anticorpos Anti-Hepatite/sangue , RNA Viral/sangue , Sequência de Bases , Doadores de Sangue/provisão & distribuição , Feminino , Genótipo , Humanos , Masculino , Filogenia , Prevalência , Ruanda , Fatores SexuaisRESUMO
BACKGROUND: Rwanda is a central African country with about 12 million inhabitants. The 1994 genocide against the Tutsi destroyed much of the infrastructure, including the health system. Although this has improved significantly, many challenges remain to be addressed. In this study, the prevalence of serological markers of past and ongoing hepatitis B virus (HBV) infection and HBV vaccine related immunity was investigated in samples from blood donors from all regions of Rwanda. METHODS: The results from hepatitis B surface antigen (HBsAg) analyses of all (45,061) blood donations collected countrywide in 2014 from 13,637 first time and 31,424 repeat blood donors were compiled. Samples from 581 HBsAg negative blood donors were selected for further analysis for antibodies against HBV, anti-HBs and anti-HBc. Additional 139 samples from HBsAg positive donors were analyzed for HBeAg/anti-HBe (132 samples) and for HBV DNA. The S-gene was amplified by PCR, products sequenced, and phylogenetic analysis was performed. RESULTS: HBsAg was found in 4.1% of first time donors with somewhat higher prevalence among those from the Central and Eastern regions than from other parts of the country. Indications of past infection was found in 21% of the HBsAg negative donors, 4.3% had only anti-HBs suggesting HBV vaccination. HBeAg was detected in 28 (21%), anti-HBe in 97 (73%), and both HBeAg and anti-HBe in 4 of 132 HBsAg positive donors. HBV DNA was found in 85 samples, and the complete S-gene was sequenced in 58 of those. Phylogenetic analysis of the sequences revealed that all HBV strains belonged to subgenotype A1, and formed one clade in the phylogenetic tree. In addition, 12 strains from first time donors had a unique 18 amino acid deletion in the N-terminal part of the pre-S2 region. CONCLUSION: This study indicated that the prevalence of hepatitis B is intermediate in Rwanda and that the vaccination coverage is relatively low in young adults. All surveyed Rwandan blood donors were infected with similar subgenotype A1 strains, and a high frequency of those with anti-HBe had detectable HBV DNA. Several strains had in addition a unique pre-S2 deletion, the virulence of which needs to be further studied.
Assuntos
Vírus da Hepatite B/genética , Hepatite B/virologia , Adulto , Doadores de Sangue , DNA Viral/genética , Feminino , Hepatite B/epidemiologia , Anticorpos Anti-Hepatite B/sangue , Antígenos do Núcleo do Vírus da Hepatite B/imunologia , Antígenos de Superfície da Hepatite B/sangue , Antígenos E da Hepatite B/sangue , Vírus da Hepatite B/imunologia , Humanos , Masculino , Filogenia , Reação em Cadeia da Polimerase , Ruanda/epidemiologia , Proteínas Virais/genéticaRESUMO
Determination of anti-hepatitis E virus (anti-HEV) antibodies is still enigmatic. There is no gold standard, and results obtained with different assays often diverge. Herein, five assays were compared for detection of anti-HEV IgM and IgG. Serum samples from 500 Swedish blood donors and 316 patients, of whom 136 had suspected HEV infection, were analyzed. Concordant results for IgM and IgG with all assays were obtained only for 71% and 70% of patients with suspected hepatitis E, respectively. The range of sensitivity for anti-HEV detection was broad (42% to 96%); this was reflected in the detection limit, which varied up to 19-fold for IgM and 17-fold for IgG between assays. HEV RNA was analyzed in all patients and in those blood donors reactive for anti-HEV in any assay, and it was found in 26 individuals. Among all of the assays, both anti-HEV IgG and IgM were detected in 10 of those individuals. Twelve had only IgG and, in 7 of those 12, IgG was only detected with the two most sensitive assays. Three of the HEV-RNA-positive samples were negative for anti-HEV IgM and IgG in all assays. With the two most sensitive assays, anti-HEV IgG was identified in 16% of the blood donor samples and in 66% of patients with suspected HEV infection. Because several HEV-RNA-positive samples had only anti-HEV IgG without anti-HEV IgM or lacked anti-HEV antibodies, analysis for HEV RNA may be warranted as a complement in the laboratory diagnosis of ongoing HEV infection.
Assuntos
Testes Diagnósticos de Rotina/métodos , Anticorpos Anti-Hepatite/sangue , Vírus da Hepatite E/imunologia , Hepatite E/diagnóstico , Imunoensaio/métodos , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Adolescente , Adulto , Idoso , Doadores de Sangue , Estudos de Coortes , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , RNA Viral/sangue , Sensibilidade e Especificidade , Adulto JovemRESUMO
AIM: The first hepatitis C virus (HCV) recombinant, RF2k/1b, was initially described from Russia and has since then been identified from patients in Ireland, Estonia, Uzbekistan and Cyprus. Many of these patients originated from Georgia; however, there is no information on its prevalence in Georgia or its susceptibility to antiviral treatment. METHODS: We retrospectively sequenced the non-structural region 5B (NS5B) of the HCV genome in samples from 72 Georgian patients, 36 of whom had been treated with pegylated interferon and ribavirin. RESULTS: The HCV genotype was determined using the Versant HCV Genotype v2 kit. Based on this typing, 32 patients (44.4%) were infected with genotype 1, 21 (29.1%) genotype 2 and 19 (26.3%) genotype 3. Partial NS5B of these strains was sequenced and analyzed for type, with concordant genotype results for all type 1 and 3 strains. Discrepant results were observed for genotyped 2 strains, with 16 (76%) having NS5B of subtype 1b. On phylogenetic analysis, 15 NS5B sequences of these strains were found in a clade formed by recombinant RF2k/1b strains. The remaining discordant sequence was found within a clade formed by 1b strains. CONCLUSION: Our findings show that the RF2k/1b recombinant strain is common among Georgian patients previously assumed to be infected with genotype 2. Because genotyping is mainly performed to decide treatment strategies, there is a need to determine the genotype by analysis of at least two genomic regions in strains from Georgian patients considered infected with genotype 2 based on standard HCV genotyping methods.
RESUMO
Amassments of heterochromatin in somatic cells occur in close contact with the nuclear envelope (NE) but are gapped by channel- and cone-like zones that appear largely free of heterochromatin and associated with the nuclear pore complexes (NPCs). To identify proteins involved in forming such heterochromatin exclusion zones (HEZs), we used a cell culture model in which chromatin condensation induced by poliovirus (PV) infection revealed HEZs resembling those in normal tissue cells. HEZ occurrence depended on the NPC-associated protein Tpr and its large coiled coil-forming domain. RNAi-mediated loss of Tpr allowed condensing chromatin to occur all along the NE's nuclear surface, resulting in HEZs no longer being established and NPCs covered by heterochromatin. These results assign a central function to Tpr as a determinant of perinuclear organization, with a direct role in forming a morphologically distinct nuclear sub-compartment and delimiting heterochromatin distribution.
Assuntos
Núcleo Celular/metabolismo , Heterocromatina/química , Complexo de Proteínas Formadoras de Poros Nucleares/química , Proteínas Proto-Oncogênicas/química , Animais , Cromatina/química , Cromatina/metabolismo , Inativação Gênica , Células HeLa , Humanos , Camundongos , Microscopia Confocal/métodos , Microscopia Eletrônica de Transmissão/métodos , Microscopia de Fluorescência/métodos , Modelos Biológicos , Poliovirus/metabolismo , Interferência de RNARESUMO
A novel virus was detected in a sample collected from a Swedish moose (Alces alces). The virus was suggested as a member of the Hepeviridae family, although it was found to be highly divergent from the known four genotypes (gt1-4) of hepatitis E virus (HEV). Moose are regularly hunted for consumption in the whole of Scandinavia. Thus, the finding of this virus may be important from several aspects: (a) as a new diverged HEV in a new animal species, and (b) potential unexplored HEV transmission pathways for human infections. Considering these aspects, we have started the molecular characterization of this virus. A 5.1 kb amplicon was sequenced, and corresponded to the partial ORF1, followed by complete ORF2, ORF3 and poly(A) sequence. In comparison with existing HEVs, the moose HEV genome showed a general nucleotide sequence similarity of 37-63% and an extensively divergent putative ORF3 sequence. The junction region between the ORFs was also highly divergent; however, two putative secondary stem-loop structures were retained when compared to gt1-4, but with altered structural appearance. In the phylogenetic analysis, the moose HEV deviated and formed its own branch between the gt1-4 and other divergent animal HEVs. The characterization of this highly divergent genome provides important information regarding the diversity of HEV infecting various mammalian species. However, further studies are needed to investigate its prevalence in the moose populations and possibly in other host species, including the risk for human infection.