RESUMO
Mutations in the internal ribosome entry site (IRES) of hepatitis A virus (HAV) have been associated with enhanced in vitro replication and viral attenuation in animal models. To address the possible role of IRES variability in clinical presentation, IRES sequences were obtained from HAV isolates associated with benign (n = 8) or severe (n = 4) hepatitis. IRES activity was assessed using a bicistronic dual-luciferase expression system in adenocarcinoma (HeLa) and hepatoma (HuH7) cell lines. Activity was higher in HuH7 than in HeLa cells, except for an infrequently isolated genotype IIA strain. Though globally low, significant variation in IRES-dependent translation efficiency was observed between field isolates, reflecting the low but significant genetic variability of this region (94.2% +/- 0.5% nucleotide identity). No mutation was exclusive of benign or severe hepatitis, and variations in IRES activity were not associated with a clinical phenotype, indirectly supporting the preponderance of host factors in determining the clinical presentation.
Assuntos
Regiões 5' não Traduzidas/genética , Vírus da Hepatite A/genética , Vírus da Hepatite A/patogenicidade , Hepatite A/virologia , RNA Viral/genética , Doença Aguda , Adolescente , Adulto , Sequência de Bases , Linhagem Celular , Criança , Primers do DNA/genética , DNA Viral/genética , França , Variação Genética , Genótipo , Células HeLa , Vírus da Hepatite A/isolamento & purificação , Vírus da Hepatite A/fisiologia , Humanos , Pessoa de Meia-Idade , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Mutação , Conformação de Ácido Nucleico , Filogenia , Biossíntese de Proteínas , RNA Viral/química , Virulência/genética , Adulto JovemRESUMO
Three hepatitis A virus (HAV) genotypes, I, II, and III, divided into subtypes A and B, infect humans. Genotype I is the most frequently reported, while genotype II is hardly ever isolated, and its genetic diversity is unknown. From 2002 to 2007, a French epidemiological survey of HAV identified 6 IIA isolates, mostly from patients who did not travel abroad. The possible African origin of IIA strains was investigated by screening the 2008 mandatory notification records of HAV infection: 171 HAV strains from travelers to West Africa and Morocco were identified. Genotyping was performed by sequencing of the VP1/2A junction in 68 available sera. Entire P1 and 5' untranslated regions of IIA strains were compared to reference sequences of other genotypes. The screening retrieved 5 imported IIA isolates. An additional autochthonous case and 2 more African cases were identified in 2008 and 2009, respectively. A total of 14 IIA isolates (8 African and 6 autochthonous) were analyzed. IIA sequences presented lower nucleotide and amino acid variability than other genotypes. The highest variability was observed in the N-terminal region of VP1, while for other genotypes the highest variability was observed at the VP1/2A junction. Phylogenetic analysis identified 2 clusters, one gathering all African and two autochthonous cases and a second including only autochthonous isolates. In conclusion, most IIA strains isolated in France are imported by travelers returning from West Africa. However, the unexplained contamination mode of autochthonous cases suggests another, still to be discovered geographical origin or a French reservoir to be explored.
Assuntos
Vírus da Hepatite A Humana/classificação , Vírus da Hepatite A Humana/genética , Hepatite A/epidemiologia , Hepatite A/virologia , Regiões 5' não Traduzidas , Adulto , Análise por Conglomerados , França/epidemiologia , Genótipo , Vírus da Hepatite A Humana/isolamento & purificação , Humanos , Pessoa de Meia-Idade , Epidemiologia Molecular , Dados de Sequência Molecular , Filogenia , RNA Viral/genética , Análise de Sequência de DNA , Homologia de Sequência , Viagem , Proteínas Estruturais Virais/genéticaRESUMO
BACKGROUND/AIMS: Liver transplantation (LT) in immune-suppressed human immunodeficiency virus (HIV) and hepatitis C virus (HCV) coinfected patients is feasible but raises questions regarding the severity of HCV recurrence on the liver graft and preservation of immune function. We investigated whether LT is deleterious to the immune system. METHODS: Fourteen HIV-HCV coinfected patients (HIV viral load [VL] <50 copies/ml; median CD4 count of 276/mm(3) pretransplantation) were grafted for HCV-cirrhosis and followed over 2 years. Nine patients received anti-HCV therapy post-transplantation. HCV and HIV VLs and degree of acute and chronic hepatitis were monitored. Peripheral blood T-cell phenotypes and interferon-gamma (IFN-gamma) immune responses against opportunistic pathogens, HCV, and HIV-1 p24 were evaluated. RESULTS: Median HCV VLs, CD4 counts, T-cell subsets, and IFN-gamma-producing T-cell frequencies against opportunistic pathogens and HIV-1 p24 did not change over time. HCV-specific T cells were observed ex vivo in two patients pretransplantation and in two others post-transplantation. HCV-specific in vitro amplification enabled the detection of HCV-specific IFN-gamma-producing responses in three further patients post-transplantation. Anti-HCV responses were observed independently of anti-HCV therapy and were undetectable in patients with severe hepatitis or liver fibrosis. CONCLUSIONS: These results demonstrate that LT in HIV-HCV coinfected patients is not deleterious to the immune system and does not alter immune responses directed against HCV, HIV, or opportunistic pathogens.
Assuntos
Infecções por HIV/complicações , Infecções por HIV/imunologia , Anticorpos Anti-Hepatite C/biossíntese , Hepatite C/complicações , Hepatite C/imunologia , Cirrose Hepática/imunologia , Cirrose Hepática/cirurgia , Transplante de Fígado/imunologia , Infecções Oportunistas Relacionadas com a AIDS/imunologia , Adulto , Contagem de Linfócito CD4 , Carcinoma Hepatocelular/etiologia , Carcinoma Hepatocelular/imunologia , Carcinoma Hepatocelular/cirurgia , Feminino , Proteína do Núcleo p24 do HIV/imunologia , Hepatite C/virologia , Humanos , Tolerância Imunológica , Técnicas In Vitro , Interferon gama/biossíntese , Cirrose Hepática/etiologia , Neoplasias Hepáticas/etiologia , Neoplasias Hepáticas/imunologia , Neoplasias Hepáticas/cirurgia , Masculino , Pessoa de Meia-Idade , Subpopulações de Linfócitos T/imunologia , Carga Viral/imunologiaRESUMO
BACKGROUND & AIMS: Hepatitis B virus (HBV) recurrence after orthotopic liver transplantation (OLT) is significantly reduced by prophylaxis with hyperimmune antibody to hepatitis B surface antigen (anti-HBs) globulins (HBIG) and antiviral drugs. The role of hepatocellular carcinoma (HCC) in HBV recurrence remains unclear. We investigated the association between HCC pre-OLT and HBV recurrence post-OLT. METHODS: We studied 99 hepatitis B surface antigen-positive patients who underwent OLT for cirrhosis. The median follow-up period was 43 months. All patients received HBIG, and 51 also received lamivudine and/or adefovir. Of these 99 patients, 31 had HCC before OLT. Total HBV DNA and covalently closed circular (ccc)-DNA were measured in tumor and nontumor tissues from the explanted livers of 16 of these 31 HCC patients and, also, in a context of tumor recurrence, in 3 patients who developed HBV/HCC recurrence. RESULTS: Fourteen patients (14.1%) developed HBV recurrence within a median period of 15 months post-OLT. HCC at OLT, a pre-OLT HBV DNA viral load > or = 100,000 copies/mL, and HBIG monoprophylaxis were independently associated with HBV recurrence post-OLT. Eleven out of the 31 patients with HCC at OLT presented with HBV recurrence and 3 out of the 68 patients without HCC had HBV recurrence (P < .0001). HBV recurrence was more frequent in patients who developed HCC recurrence (7/8 patients, 87.5%) than in those who did not (4/23 patients, 17.4%) (P < .0001). In the 16 explanted livers, cccDNA was detectable in HCC cells in 11 and in nontumor cells in 12. cccDNA was detected in a context of HCC recurrence in 2 of the 3 patients tested who developed HBV/HCC recurrence. CONCLUSIONS: The associations of HCC pre-OLT, and HCC recurrence with HBV recurrence post-OLT, and the detection of HBV DNA and cccDNA in HCC suggest that HBV replication in tumor cells may contribute to HBV recurrence post-OLT.
Assuntos
Carcinoma Hepatocelular/virologia , Vírus da Hepatite B , Hepatite B/etiologia , Cirrose Hepática/virologia , Neoplasias Hepáticas/virologia , Transplante de Fígado , Adenina/análogos & derivados , Adenina/uso terapêutico , Adolescente , Adulto , Idoso , Antivirais/uso terapêutico , Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/cirurgia , DNA Circular/análise , DNA Viral/análise , Feminino , Seguimentos , Hepatite B/complicações , Hepatite B/diagnóstico , Hepatite B/tratamento farmacológico , Hepatite B/cirurgia , Antígenos de Superfície da Hepatite B/análise , Vírus da Hepatite B/genética , Vírus da Hepatite B/crescimento & desenvolvimento , Vírus da Hepatite B/imunologia , Humanos , Imunoglobulinas/uso terapêutico , Imuno-Histoquímica , Lamivudina/uso terapêutico , Cirrose Hepática/tratamento farmacológico , Cirrose Hepática/cirurgia , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/cirurgia , Masculino , Pessoa de Meia-Idade , Organofosfonatos/uso terapêutico , Reação em Cadeia da Polimerase , Medição de Risco , Fatores de Risco , Prevenção Secundária , Análise de Sequência de DNA , Fatores de Tempo , Resultado do Tratamento , Carga Viral , Replicação ViralRESUMO
We assessed the feasibility of using dried serum spots (DSS) for the serological and molecular diagnosis of hepatitis A virus (HAV) infection. Sixty-eight sera spotted onto filter papers (Whatman International Ltd., United Kingdom) were used for detection of total anti-HAV antibodies, and 64 sera were used for detection of immunoglobulin M antibody to HAV. DSS were stored at 4 degrees C, room temperature, and 37 degrees C for 1, 2, and 4 weeks. Sensitivity and specificity of the serological assays were 100% regardless of temperature and storage duration. To assess the stability of HAV RNA, we performed qualitative and quantitative reverse transcription-PCRs (RT-PCRs) with human plasma spiked with serial dilutions of cultured HAV spotted on Flinders Technology Associates filter paper cards (Whatman International Ltd.). Filter papers were stored at room temperature and processed for RT-PCR assays. No reduction of viral load was observed after 5, 15, and 30 days of storage. The approximately 10-fold reduction of sensitivity from DSS was attributable to a smaller sample input in DSS samples. This method was further evaluated using 35 frozen sera. HAV RNA amplification showed 100% specificity and 92.3% sensitivity, and sequence analysis from DSS and sera provided identical results. HAV RNA can be accurately recovered from DSS for molecular epidemiology purposes, and we confirm the reliability of blotted samples in the serological diagnosis of HAV infection. The DSS method facilitates storage and shipment of samples from routine laboratories to reference centers for further investigations and large epidemiological studies.
Assuntos
Dessecação , Anticorpos Anti-Hepatite A/sangue , Vírus da Hepatite A/isolamento & purificação , Hepatite A/diagnóstico , Soro/virologia , Manejo de Espécimes/métodos , Humanos , Dados de Sequência Molecular , Filogenia , RNA Viral/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sensibilidade e Especificidade , Análise de Sequência de DNA , Homologia de Sequência , Reino UnidoRESUMO
A 63-year-old woman treated with prednisone for sinusitis developed fulminant liver failure due to a clinically unsuspected primary varicella zoster virus infection. The diagnosis of herpetic hepatitis was made from a liver biopsy, and varicella zoster virus viremia was detected by polymerase chain reaction. She was treated successfully with transplantation and perioperative administration of acyclovir.
Assuntos
Varicela/complicações , Herpesvirus Humano 3/metabolismo , Falência Hepática Aguda/complicações , Falência Hepática Aguda/terapia , Transplante de Fígado/métodos , Aciclovir/uso terapêutico , Antivirais/uso terapêutico , Biópsia , Varicela/terapia , Feminino , Humanos , Fígado/virologia , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Prednisona/efeitos adversos , Resultado do TratamentoRESUMO
BACKGROUND AND OBJECTIVES: Enzyme-linked immunoassays (ELISA) are the most widely used anti-hepatitis C virus (HCV) screening tests but simple, instrument and electricity-free screening tests have been developed with results available in a few minutes. METHODS: The sensitivity of a rapid immuno-chromatographic assay for the detection of anti-HCV antibodies was evaluated on 421 HCV RNA-positive samples from chronic carriers and compared with ELISA method. RESULTS: The sensitivity of the ELISA method was 99.3% and the sensitivity of the rapid test was 95.5%. False negative results were independent of HCV genotype, but were associated with human immunodeficiency virus (HIV)-positive status. Among HIV-negative people, sensitivities of the rapid test and the EIA assay were 99.2% and 100%, respectively. Whereas among HIV-positive people, sensitivities were 77.5% and 96.3%. CONCLUSIONS: The immuno-chromatographic test is rapid and simple, and could be used along with rapid anti-HIV determination, in settings with limited facilities or when rapid results are required.
Assuntos
Cromatografia/métodos , Hepacivirus/imunologia , Anticorpos Anti-Hepatite C/sangue , Hepatite C Crônica/diagnóstico , Adulto , Ensaio de Imunoadsorção Enzimática/métodos , Feminino , Hepatite C Crônica/imunologia , Hepatite C Crônica/virologia , Humanos , Masculino , Pessoa de Meia-Idade , Kit de Reagentes para Diagnóstico , Sensibilidade e Especificidade , Fatores de TempoRESUMO
BACKGROUND: Understanding the prevalence of potential antigenic variation of the hepatitis B virus (HBV) surface antigen (HBsAg) is fundamental for assay design and to future changes in vaccine formulation. In this study, the nature and frequency of HBsAg polymorphisms occurring in France in chronic carriers and in newly diagnosed patients were determined. We focused on variations in the major hydrophilic region (MHR), the central core of HBsAg known to be exposed on the surface and involved in antibody binding. METHODS: Two patient groups were identified: 51 chronic HBV carriers followed at our institution for > 1 year; and 129 newly diagnosed patients (63 of whom had a first HBsAg-positive result at our hospital laboratory and 66 a first positive result in a private laboratory). DNA sequences of HBsAg were obtained from these 180 patients and compared with consensus sequences built with 168 full-length HBV sequences imported from GenBank. Polymorphisms of the MHR of HBsAg were analysed with the Mutation Master Software. Literature review and BLOSUM scores were used to define potentially altered antigenicity. RESULTS: The global frequency of MHR variants was 27.8%. Occurrence of MHR variants was independent of viral load, HBeAg status and sex, but was associated with the chronic carriers' group, advancing age, the presence of antibodies to HBsAg, immunoprophylaxis administration, antiviral treatment and genotypic resistance to antivirals. In multivariate analysis, the independent variables associated with MHR variants were advancing age and the presence of genotypic resistance to nucleoside or nucleotide analogues. CONCLUSION: Most MHR variants emerge with longer disease duration and upon indirect selective pressure. Variation of the MHR may serve to restore virus replication of resistant strains. Combined envelope and polymerase variants could impair diagnostic assays and limit treatment alternatives.
Assuntos
Antígenos de Superfície da Hepatite B/genética , Vírus da Hepatite B/genética , Hepatite B/virologia , Fatores Etários , Sequência de Aminoácidos , Portador Sadio , Farmacorresistência Viral/genética , Feminino , França , Hepatite B/sangue , Hepatite B/tratamento farmacológico , Anticorpos Anti-Hepatite B/sangue , Antígenos de Superfície da Hepatite B/imunologia , Vírus da Hepatite B/efeitos dos fármacos , Hepatite B Crônica/virologia , Humanos , Masculino , Pessoa de Meia-Idade , Dados de Sequência Molecular , Nucleosídeos/farmacologia , Nucleotídeos/farmacologia , Polimorfismo Genético , Alinhamento de Sequência , Carga ViralAssuntos
Proteínas de Bactérias/biossíntese , Surtos de Doenças , Infecções por Klebsiella/epidemiologia , Infecções por Klebsiella/microbiologia , Klebsiella pneumoniae/enzimologia , beta-Lactamases/biossíntese , Infecção Hospitalar/epidemiologia , Infecção Hospitalar/microbiologia , França , Transferência Genética Horizontal , Hospitais , Humanos , Klebsiella pneumoniae/isolamento & purificação , Plasmídeos/análiseAssuntos
Técnicas de Tipagem Bacteriana , Infecções por Escherichia coli/transmissão , Saúde da Família , Filogenia , Escherichia coli Uropatogênica/enzimologia , beta-Lactamases/biossíntese , Adulto , Impressões Digitais de DNA , Infecções por Escherichia coli/microbiologia , Proteínas de Escherichia coli/biossíntese , Feminino , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Técnica de Amplificação ao Acaso de DNA Polimórfico , Infecções Urinárias/microbiologia , Infecções Urinárias/transmissão , Escherichia coli Uropatogênica/classificação , Escherichia coli Uropatogênica/isolamento & purificaçãoRESUMO
AIM: Diagnosis of acute hepatitis A virus (HAV) infection is classically based on the detection of HAV-IgM. Nevertheless, HAV-IgM can be positive for patients with polyclonal stimulation of their immune system (i.e. immune reactivation). To improve the diagnostic yield, an avidity test for HAV-IgG antibodies was developed and tested. METHODS: Avidity tests were performed in 128 sera: 11 selected samples from patients with past infection, 15 acute hepatitis A, 10 vaccinated subjects and 4 patients with immune reactivation as well as 84 HAV-IgM positive unselected sera, provided by routine laboratories. RESULTS: Patients with past infection had avidities over 70%, whereas avidities in patients with acute hepatitis A were below 50% during the first month following the onset of symptoms. As expected, patients with immune reactivation had avidities over 70% consistent with past infection. The results obtained for the 84 unselected sera allowed reconsidering the diagnosis of acute hepatitis A for nearly a third of patients. CONCLUSION: This test could improve the diagnosis of acute hepatitis A infection, particularly in elderly patients.
Assuntos
Afinidade de Anticorpos , Hepatite A/diagnóstico , Hepatite A/imunologia , Imunoglobulina G/imunologia , Adolescente , Adulto , Idoso , Estudos de Casos e Controles , Criança , Diagnóstico Diferencial , Feminino , Humanos , Imunoglobulina G/análise , Masculino , Pessoa de Meia-Idade , Sensibilidade e Especificidade , Testes SorológicosRESUMO
Sera from 12 patients infected with human immunodeficiency virus and hepatitis B virus (HBV), on lamivudine as part of an antiretroviral therapy, were retrospectively analysed for the presence of HBV polymerase mutations by the line probe assay, INNO-LiPA HBV DR, and by the direct sequencing assay, TRUGENE HBV genotyping kit. Results at codons 180, 204 and 207 were compared for 44 samples. Full concordance was observed for 81.4% of the 129 analysed codons. Discordance involved only mixed populations: LiPA detected additional species in 19 codons and TRUGENE in five. Viral breakthrough occurred in seven patients, 12-33 months after lamivudine initiation. In five cases with close sampling available, both assays detected mutations before the rise in viral load, although earlier by LiPA for three patients. The time interval between the first mutant detection and viral escape ranged from 2 to 22 months. Mutations were detected in four of the five remaining patients: 1) at therapy initiation in a primary non-responder; 2) after 37 months, but replication became undetectable after tenofovir introduction; 3) transiently at 6 months by LiPA but treatment was ceased thereafter; 4) after 23 months but replication levels remained low during a 5-year follow-up. Interestingly, TRUGENE sequencing identified on late samples from three patients a variant carrying rtV173L plus rtL180M plus M204V mutations, having the in vitro characteristics of 'vaccine escape' mutants. Both assays appear to be valuable tools for the early detection of mutated HBV strains. The detection of genotypic therapeutic decision-making, although clinical or other virological factors may determine the rapidity of the viral breakthrough.
Assuntos
Síndrome da Imunodeficiência Adquirida/virologia , Produtos do Gene pol/genética , HIV-1 , Vírus da Hepatite B/genética , Hepatite B/virologia , Lamivudina/uso terapêutico , Mutação , Adulto , Idoso , DNA Viral/análise , Farmacorresistência Viral , Genótipo , Hepatite B/tratamento farmacológico , Humanos , Masculino , Pessoa de Meia-Idade , Hibridização de Ácido Nucleico , Reação em Cadeia da Polimerase , Kit de Reagentes para Diagnóstico , Análise de Sequência de DNARESUMO
OBJECTIVES: To correlate human herpesvirus 6 (HHV-6) viral load with pathologic features in graft acute hepatitis of unknown origin. METHODS: Liver frozen samples from 26 patients with graft hepatitis of unknown origin were available for HHV-6 DNA quantification. RESULTS: In 10 (38.5%) of 26 liver samples, HHV-6 DNA was detectable, with a median viral load of 3.84 log10 copies/106 cells. Confluent periportal necrosis was observed in 4 of 10 patients and associated with high viral load. These 4 patients responded to antiviral therapy. Mild unspecific hepatitis was observed in 4 patients with low intragraft viral load and in 2 patients with high viral load in a context of deep immunosuppression. Patients with HHV-6-negative graft hepatitis disclosed lobular necrotico-inflammatory activity without periportal necrosis. CONCLUSIONS: Our study provides data supporting the pathogenic role of HHV-6 for liver allografts. The presence of confluent periportal necrosis could be a clue for prompt diagnosis of HHV-6-induced graft hepatitis.
Assuntos
Hepatite/patologia , Herpesvirus Humano 6/isolamento & purificação , Transplante de Fígado/patologia , Fígado/patologia , Infecções por Roseolovirus/patologia , Adolescente , Adulto , DNA Viral , Feminino , Hepatite/virologia , Humanos , Fígado/virologia , Masculino , Pessoa de Meia-Idade , Infecções por Roseolovirus/virologia , Carga ViralAssuntos
Anticorpos Anti-Hepatite A/sangue , Vírus da Hepatite A Humana/imunologia , Hepatite A/diagnóstico , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Afinidade de Anticorpos , Feminino , Anticorpos Anti-Hepatite A/imunologia , Humanos , Imunoglobulina G/imunologia , Masculino , Valor Preditivo dos Testes , Viremia/diagnósticoRESUMO
There is little information on the diverse infectious causes of jaundice and hepatitis in the Asiatic tropics. Serology (hepatitis A, B, C and E, leptospirosis, dengue, rickettsia), antigen tests (dengue), PCR assays (hepatitis A, C and E) and blood cultures (septicaemia) were performed on samples from 392 patients admitted with jaundice or raised transaminases (> or =x3) to Mahosot Hospital, Vientiane, Laos over 3 years. Conservative definitions suggested diagnoses of dengue (8.4%), rickettsioses (7.3%), leptospirosis (6.8%), hepatitis B (4.9%), hepatitis C (4.9%), community-acquired septicaemia (3.3%) and hepatitis E (1.6%). Although anti-hepatitis A virus (HAV) IgM antibody results suggested that 35.8% of patients had acute HAV infections, anti-HAV IgG antibody avidity and HAV PCR suggested that 82% had polyclonal activation and not acute HAV infections. Scrub typhus, murine typhus or leptospirosis were present in 12.8% of patients and were associated with meningism and relatively low AST and ALT elevation. These patients would be expected to respond to empirical doxycycline therapy which, in the absence of virological diagnosis and treatment, may be an appropriate cost-effective intervention in Lao patients with jaundice/hepatitis.
Assuntos
Hepatite Viral Humana/etiologia , Icterícia/microbiologia , Adolescente , Adulto , Criança , Pré-Escolar , Infecções Comunitárias Adquiridas/diagnóstico , Dengue/diagnóstico , Feminino , Febre/microbiologia , Hepatite Viral Humana/diagnóstico , Hospitalização , Humanos , Lactente , Recém-Nascido , Icterícia/virologia , Laos , Leptospirose/diagnóstico , Masculino , Pessoa de Meia-Idade , RNA Viral/isolamento & purificação , Infecções por Rickettsia/diagnóstico , Tifo por Ácaros/diagnóstico , Testes Sorológicos/métodos , Tifo Endêmico Transmitido por Pulgas/diagnóstico , Adulto JovemRESUMO
The Kaposi sarcoma-associated herpesvirus (KSHV)-DNA level was determined in samples from 71 patients with Kaposi sarcoma (KS), 28 patients with multicentric Castleman disease (MCD), and 8 patients with primary effusion lymphoma (PEL). KSHV-DNA levels were higher in patients with active KS or MCD than in those with KS or MCD in remission. Among patients with active disease, the highest KSHV-DNA levels were observed in effusion fluid samples from patients with PEL (7.2 log(10) copies/150,000 cells), followed by blood samples from patients with MCD and PEL (4.86 and 3.83 log(10) copies/150,000 cells, respectively), and the lowest levels were observed in blood samples from patients with KS (2.63 log(10) copies/150,000 cells). Determining the KSHV-DNA level may be useful in diagnosing KSHV-associated disease and for following up patients with KS when the development of MCD or PEL is suspected.
Assuntos
Hiperplasia do Linfonodo Gigante/virologia , DNA Viral/sangue , Herpesvirus Humano 8/genética , Linfoma/virologia , Sarcoma de Kaposi/virologia , Carga Viral , Adulto , Idoso , Idoso de 80 Anos ou mais , Líquido Ascítico/virologia , Contagem de Linfócito CD4 , Hiperplasia do Linfonodo Gigante/complicações , Feminino , Infecções por HIV/complicações , Herpesvirus Humano 8/patogenicidade , Humanos , Linfoma/complicações , Masculino , Pessoa de Meia-Idade , Derrame Pleural/virologia , Estudos Retrospectivos , Sarcoma de Kaposi/complicaçõesRESUMO
Mutations in hepatitis B virus surface antigen (HBsAg) involving amino acid substitution within the immunodominant "a" determinant may affect the performance of commercial HBsAg assays. The performances of four HBsAg assays that recently received Conformité Européene marking, Advia Centaur HBsAg (Bayer), Monolisa HBsAg Ultra (Bio-Rad), Liaison HBsAg (Dia Sorin), and Vidas HBsAg Ultra (bioMérieux), were compared with that of the routinely used HBsAg assay AxSYM HBsAg V2 (Abbott). Assays were evaluated for (i) analytical sensitivity performance with a national reference HBsAg panel (including 10 samples with calibrated HBsAg concentrations from 0.04 to 2.24 ng/ml) and (ii) the detection of HBsAg mutants by studying a panel of 35 HBsAg mutants (23 collected from patients and 12 recombinant mutants). The limits of detection of these assays were <0.15 ng/ml (from 0.089 to 0.121 ng/ml). The sensitivity performances for mutant virus detection varied, ranging from 37.1% to 91.4%. The lack of detection of these mutants by commercial assays was probably due to the epitope recognition of the anti-HBs assay reagents in the capture phase and in the conjugates. The prevalence and clinical impact of HBsAg mutants are under investigation. However, the manufacturers must be vigilant in the design of the assays in order to reduce the risk of missing a broad range of described S gene mutants.
Assuntos
Antígenos de Superfície da Hepatite B/análise , Imunoensaio , Proteínas Mutantes/análise , Antígenos de Superfície da Hepatite B/genética , Humanos , Proteínas Mutantes/genética , Kit de Reagentes para Diagnóstico , Sensibilidade e EspecificidadeRESUMO
OBJECTIVES: We studied eight imipenem-resistant isolates of Klebsiella pneumoniae involved in an outbreak in a French teaching hospital. METHODS: The eight isolates were recovered from clinical specimens or rectal swabs. Antibiotic susceptibilities were determined using standard agar diffusion and dilution methods including synergy tests. PFGE was used to study the relatedness of isolates. Genes encoding beta-lactamases were characterized by transfer assays, specific amplification and cloning. RESULTS: The eight isolates were closely related by PFGE analysis and highly related to a K. pneumoniae strain from Greece. They were highly resistant to beta-lactams, including aztreonam and imipenem (MIC > or =32 mg/L), and were positive by the imipenem-EDTA disc synergy test. Isolates were also resistant to aminoglycosides, newer quinolones and sulfamethoxazole, and showed an intermediate level of resistance to tetracycline. VIM-1 and SHV-5 beta-lactamases were revealed in all isolates by PCR. The analysis of plasmid contents of Escherichia coli DH10B electroporants expressing the VIM-1 beta-lactamase or the SHV-5 beta-lactamase confirmed that the two enzymes were coded by two different plasmids. The bla(VIM-1) gene was part of a class 1 integron that also included aac6, dhfrI and aadA genes and was similar to those reported from strains isolated in Greece. CONCLUSIONS: This study confirms the potential risk of spread of multiresistant bacteria with international transfer of patients.
Assuntos
Antibacterianos/farmacologia , Infecção Hospitalar/microbiologia , Surtos de Doenças , Farmacorresistência Bacteriana Múltipla/genética , Infecções por Klebsiella/microbiologia , Klebsiella pneumoniae/enzimologia , beta-Lactamases/genética , Conjugação Genética , Infecção Hospitalar/epidemiologia , DNA Bacteriano/genética , Eletroforese em Gel de Campo Pulsado , Escherichia coli/enzimologia , Escherichia coli/genética , França , Hospitais Universitários , Humanos , Infecções por Klebsiella/epidemiologia , Klebsiella pneumoniae/genética , Klebsiella pneumoniae/isolamento & purificação , Testes de Sensibilidade MicrobianaRESUMO
Investigation of hepatitis A virus (HAV) outbreaks often implies nucleotide sequence analysis. As an alternative method for the identification of related strains, single strand conformation polymorphism method (SSCP) was compared to sequence analysis. Twenty-three strains from sporadic and outbreak cases were studied retrospectively. SSCP, sequence identity and phylogenetic analyses were conducted on a 267 bp fragment of the VP1-2A variable region. The results of SSCP pattern comparison and sequence identity were highly correlated (r = 0.92, P < 0.001). If SSCP showed similar patterns, the VP1-2A fragments had a high and significant probability to have a sequence identity over 99.6%. Results were concordant for outbreak strains. The only discordant result concerned a cluster of three sporadic cases evidenced by phylogenetic analysis while SSCP showed similar patterns for only two of these three cases. A prospective SSCP analysis of a recent HAV outbreak confirmed the reliability of this technique. SSCP may thus provide a rapid and cost-effective tool for preliminary investigation of HAV outbreaks, before undertaking exhaustive nucleotide sequence analysis.
Assuntos
Surtos de Doenças , Vírus da Hepatite A/classificação , Vírus da Hepatite A/genética , Hepatite A/epidemiologia , Hepatite A/virologia , Polimorfismo Conformacional de Fita Simples , Adolescente , Adulto , Criança , Pré-Escolar , Cisteína Endopeptidases/genética , Feminino , Vírus da Hepatite A/isolamento & purificação , Humanos , Masculino , Pessoa de Meia-Idade , Epidemiologia Molecular , Filogenia , Estudos Retrospectivos , Análise de Sequência de DNA , Proteínas Virais/genética , Proteínas Estruturais Virais/genéticaRESUMO
Differences in hepatitis C virus (HCV) variants of the highly conserved 5' untranslated region (UTR) have been observed between plasma and peripheral blood mononuclear cells (PBMC). The prevalence and the mechanisms of this compartmentalization are unknown. Plasma and PBMC HCV variants were compared by single-strand conformation polymorphism (SSCP) and by cloning or by genotyping with a line probe assay (LiPA) in 116 chronically infected patients, including 44 liver transplant recipients. SSCP patterns differed between compartments in 43/109 analyzable patients (39%). Differences were significantly more frequent in patients with transplants (21/38 [55%] versus 22/71 [31%]; P < 0.01) and in those who acquired HCV through multiple transfusions before 1991 (15/20; 75%) or through drug injection (16/31; 52%) than in those infected through an unknown route (7/29; 24%) or through a single transfusion (5/29; 17%; P < 0.001). Cloning of the 5' UTR, LiPA analysis, and nonstructural region 5B sequencing revealed different genotypes in the two compartments from 10 patients (9%). In nine patients, the genotype detected in PBMC was not detected in plasma and was weak or undetectable in the liver in three cases. This genotypic compartmentalization persisted for years in three patients and after liver transplantation in two. The present study shows that a significant proportion of HCV-infected subjects harbor in their PBMC highly divergent variants which were likely acquired through superinfections.