RESUMEN
The Procleix HIV-1/HCV Assay is a high-throughput nucleic acid test for the simultaneous detection of human immunodeficiency virus type 1 (HIV-1) and hepatitis C virus (HCV) RNA during blood donor screening. This study evaluated the clinical sensitivity of the Procleix assay and assessed the assay's ability to identify HIV-1- and HCV-infected individuals undetected by standard serologic tests. Plasma samples were obtained prospectively from 539 individuals at high risk for HIV-1 and HCV infection at seven clinics affiliated with Johns Hopkins University. Samples were tested in the Procleix HIV-1/HCV Assay and, if reactive, were then tested in the Procleix HIV-1 and HCV discriminatory assays to differentiate the source of viral RNA positivity. Of these 539 subjects, 287 (53.2%) tested reactive in the Procleix HIV-1/HCV Assay. In discriminatory assay testing, 12 of 287 subjects (4.2%) were reactive for HIV-1 RNA only, 260 (90.6%) were reactive for HCV RNA only, and 11 (3.8%) were coinfected with HIV-1 and HCV. The clinical sensitivity for samples tested neat was 100% for HIV-1 and 99.3% for HCV. Three subjects with Procleix HCV reactive/seronegative results seroconverted upon follow-up and were confirmed as Procleix HCV yield cases. The Procleix HIV-1/HCV Assay is a highly sensitive test that detects ongoing and early HIV-1 and HCV infection in a significant number of subjects at high risk for these diseases. Confirmation of Procleix yield cases upon follow-up demonstrated the ability of the Procleix HIV-1/HCV Assay to detect the presence of HIV-1 and HCV in blood earlier than standard serologic tests.
Asunto(s)
VIH-1/aislamiento & purificación , Hepacivirus/aislamiento & purificación , ARN Viral/análisis , Virología/métodos , Serodiagnóstico del SIDA , Adolescente , Adulto , Anciano , Donantes de Sangre , Femenino , Infecciones por VIH/diagnóstico , VIH-1/genética , Hepacivirus/genética , Hepatitis C/diagnóstico , Humanos , Masculino , Tamizaje Masivo/métodos , Tamizaje Masivo/estadística & datos numéricos , Persona de Mediana Edad , ARN Viral/sangre , ARN Viral/genética , Factores de Riesgo , Sensibilidad y Especificidad , Virología/estadística & datos numéricosRESUMEN
Various nucleic acid assays have been developed and implemented for diagnostics and therapeutic monitoring of human immunodeficiency virus type 1 (HIV-1) and hepatitis C virus (HCV) infections. The high-throughput, semiautomated assays described here were developed to provide a method suitable for screening plasma specimens for the presence of HIV-1 and HCV RNAs. Three assays were developed: a multiplex HIV-1/HCV assay for simultaneous detection of HIV-1 and HCV, and discriminatory assays for specific detection of HIV-1 and HCV. The assay systems utilize three proprietary technologies: (i) target capture-based sample preparation, (ii) transcription-mediated amplification (TMA), and (iii) hybridization protection assay (HPA). An internal control is incorporated into each reaction to control for every step of the assay and identify random false-negative reactions. The assays demonstrated a sensitivity of at least 100 copies/ml for each target, and they detected with similar sensitivity all major variants of HCV and HIV-1, including HIV-1 group O strains. Assay sensitivity for one virus was not affected by the presence of the other. The specificity of these TMA-driven assays was >or=99.5% in both normal donor specimens and plasma containing potentially interfering substances or other blood-borne pathogens. Statistical receiver operating characteristic plots of 1 - specificity versus sensitivity data determined very wide analyte cutoff values for each assay at the point at which the assay specificity and sensitivity were both >or=99.5%. The sensitivity, specificity, and throughput capability predict that these assays will be valuable for large-volume plasma screening, either in a blood bank setting or in other diagnostic applications.
Asunto(s)
VIH-1/genética , VIH-1/aislamiento & purificación , Hepacivirus/genética , Hepacivirus/aislamiento & purificación , ARN Viral/análisis , ARN Viral/genética , Virología/métodos , Donantes de Sangre , Infecciones por VIH/complicaciones , Infecciones por VIH/diagnóstico , Infecciones por VIH/virología , Hepatitis C/complicaciones , Hepatitis C/diagnóstico , Hepatitis C/virología , Humanos , Tamizaje Masivo , Reproducibilidad de los Resultados , Sensibilidad y Especificidad , Virología/estadística & datos numéricosRESUMEN
Three assays, one based on monoclonal antibodies and the others on polyclonal antibodies, were employed to detect hepatitis B surface antigen (HBsAg)-reactive samples in both vaccinated and unvaccinated populations in areas of the world where hepatitis B virus (HBV) is endemic. Any discordant sera were tested by polymerase chain reaction (PCR) to confirm current infection, and sequence data were obtained from the DNA coding for the major hydrophilic region (MHR) of HBsAg of those samples positive for PCR. In all countries studied, samples that reacted in one HBsAg assay but not another were found. In the most extreme case, about 5% of viremic sera in Papua New Guinea were nonreactive in the monoclonal HBsAg assay; 9 of the 13 PCR-positive samples had novel or once-described variants, or a variant out of its usual genotype context. In South Africa, samples with sequences of subtype ayw2 reacted poorly, particularly in the polyclonal assay. Two had novel variants. In Sardinia, antibody to hepatitis B core antigen (anti-HBc) was analyzed as a marker of infection. A significant proportion of anti-HBc-positive, but monoclonal HBsAg-negative, vaccinees and unvaccinated persons were found to be PCR positive, as were some individuals without any markers of hepatitis B virus infection. Five more novel variants were found in these groups. There are implications for the design of HBsAg assays, which may have to be modified according to local sequence variability. Not all discordant samples were explained by variants, indicating that assay sensitivity is fundamental to diagnostic efficacy. Overall, this study defined 16 novel variants and 2 new potential epitope clusters.
Asunto(s)
Epítopos/análisis , Antígenos de Superficie de la Hepatitis B/inmunología , Virus de la Hepatitis B/inmunología , Hepatitis B/inmunología , Secuencia de Aminoácidos , Anticuerpos Monoclonales , ADN Viral/análisis , Reacciones Falso Positivas , Genotipo , Hepatitis B/epidemiología , Hepatitis B/prevención & control , Antígenos de Superficie de la Hepatitis B/genética , Vacunas contra Hepatitis B/administración & dosificación , Virus de la Hepatitis B/genética , Humanos , Italia/epidemiología , Tamizaje Masivo , Datos de Secuencia Molecular , Nueva Guinea/epidemiología , Reacción en Cadena de la Polimerasa , Prevalencia , Sensibilidad y Especificidad , Sudáfrica/epidemiologíaRESUMEN
Patients presenting with clinical and laboratory features consistent with a diagnosis of acute non-A, non-B hepatitis were evaluated for evidence of hepatitis C or hepatitis E infection and for evidence of severe or prolonged disease. Antibody to hepatitis C virus (anti-HCV) was detected in 75 of 108 (69%) patients, antibody to hepatitis E virus (anti-HEV) in three patients (3%), and neither antibody in 31 (29%) patients. One patient had both anti-HCV and anti-HEV. HCV RNA was not detected in sera from any of 20 patients with seronegative (non-ABCDE) hepatitis, but in all 10 patients with anti-HCV who were tested by polymerase chain reaction (PCR). Compared with patients with acute hepatitis C, those with non-ABCDE hepatitis had a lower incidence of parenteral risk factors (6% vs. 70%; P < .001), higher peak serum bilirubin levels (45% vs. 5% with peak levels > 15 mg/dL; P < .001), more prolonged jaundice (25% vs. 0% with peak bilirubin >5 weeks after onset; P < .01), more severe prothrombin time abnormalities (26% vs. 0% with >3 second prolongation; P < .001), more severe hypoalbuminemia (39% vs. 9% with albumin <3 g/dL; P < .01), and more frequent major clinical complications (13% vs. 0% with encephalopathy; P < .01; 10% vs. 0% with death or transplant; P = .024). Patients with acute non-ABCDE hepatitis were less likely to develop chronic hepatitis than those with acute hepatitis C (23% vs. 68%; P < .05). Thus, patients with acute non-ABCDE hepatitis are epidemiologically distinct from those with acute hepatitis C and have a significantly more severe acute illness.
Asunto(s)
Hepacivirus/inmunología , Hepatitis C/inmunología , Hepatitis C/virología , Virus de la Hepatitis E/inmunología , Hepatitis E/inmunología , Hepatitis E/virología , Adolescente , Adulto , Anciano , Bilirrubina/sangre , Demografía , Femenino , Hepatitis C/sangre , Hepatitis E/sangre , Humanos , Masculino , Persona de Mediana EdadRESUMEN
The structure of the small hepatitis B virus surface antigen (HBsAg) was investigated by epitope mapping of four anti-HBsAg monoclonal antibodies (mAbs). Amino acid sequences of epitopes were derived from affinity-enrichment experiments (biopanning) using a filamentous phage peptide library. The library consists of 10(9) different clones bearing a 30-residue peptide fused to gene III. Sequence homologies between peptides obtained from panning the library against the antibodies and the native HBsAg sequence allowed for precise description of the binding regions. Three of four mAbs were found to bind to distinct discontinuous epitopes between amino acid residues 101 and 207 of HBsAg. The fourth mAb was demonstrated to bind to residues 121-124. The sequence data are supported by ELISA assays demonstrating the binding of the HBsAg-specific peptides on filamentous phage to mAbs. The sequence data were used to map the surface of HBsAg and to derive a topological model for the alpha-carbon trace of the 101-207 region of HBsAg. The approach should be useful for other proteins for which the crystal structure is not available but a representative set of mAbs can be obtained.
Asunto(s)
Anticuerpos contra la Hepatitis B/inmunología , Antígenos de Superficie de la Hepatitis B/inmunología , Virus de la Hepatitis B/inmunología , Secuencia de Aminoácidos , Anticuerpos Monoclonales/inmunología , Unión Competitiva , Colifagos , Mapeo Epitopo , Biblioteca de Genes , Datos de Secuencia Molecular , Estructura Terciaria de ProteínaRESUMEN
OBJECTIVES: The goals of this study were to examine responses to corticosteroid-containing therapy in non-B chronic hepatitis patients with different anti-hepatitis C virus (HCV), autoantibody, and biochemical test results and to determine what factors correlate with response. METHODS: Patients with a prior or current history of steroid therapy for putative autoimmune or chronic non-A, non-B hepatitis were assessed. Responses during the first 6 months of therapy were categorized as "complete" (normal aminotransferases for > or = 1 month), "partial" ( > 50% reduction), or "no response." RESULTS: Sufficient data available to permit evaluation in 32 patients. Complete responses were noted in 17, partial responses in 12, and no response in three subjects. By multivariate analysis, only absence of anti-HCV and presence of cirrhosis were independent predictors of response. Nonresponders were found to have lower scores in a proposed autoimmune hepatitis scoring system, but scores of complete and partial responders were not significantly different. Despite a lower likelihood of a complete response, 80% (12/15) of patients with multiantigen positive anti-HCV tests had either partial or complete initial responses to corticosteroid-containing therapy, and, in nine patients, aminotransferases fell to < 2 times the upper limit of normal. All 15 anti-HCV-negative patients, but only three of 15 anti-HCV-positive patients, entered complete responses that were sustained (aminotransferases < twofold abnormal) on regimens containing < 20 mg/day or prednisolone or prednisone. CONCLUSIONS: Although anti-HCV-positive patients frequently exhibit partial initial responses to immunosuppressive therapy, the absence of specific anti-HCV antibodies was better as a predictor of completeness of response than assessment of autoantibodies or degree of biochemical abnormalities.
Asunto(s)
Corticoesteroides/uso terapéutico , Enfermedades Autoinmunes/tratamiento farmacológico , Hepacivirus/genética , Anticuerpos contra la Hepatitis C/análisis , Hepatitis C/tratamiento farmacológico , Hepatitis Crónica/tratamiento farmacológico , Hepatitis/tratamiento farmacológico , ARN Viral/análisis , Corticoesteroides/administración & dosificación , Adulto , Antiinflamatorios/administración & dosificación , Autoanticuerpos/análisis , Enfermedades Autoinmunes/diagnóstico , Enfermedades Autoinmunes/inmunología , Secuencia de Bases , Pruebas Enzimáticas Clínicas , Cartilla de ADN , Femenino , Glucocorticoides/administración & dosificación , Hepacivirus/inmunología , Hepatitis/diagnóstico , Hepatitis/inmunología , Hepatitis C/diagnóstico , Hepatitis Crónica/diagnóstico , Humanos , Masculino , Persona de Mediana Edad , Datos de Secuencia Molecular , Prednisolona/administración & dosificación , Prednisona/administración & dosificación , Técnica del ADN Polimorfo Amplificado Aleatorio , Estudios Retrospectivos , Factores de TiempoRESUMEN
The detection of a fatal case of reactivation of hepatitis B, in a previously vaccinated Indonesian patient after withdrawal of chemotherapy for lymphoma, was delayed because HBsAg was negative in a widely used monoclonal-antibody-based ELISA. The serum was later found to be strongly reactive for HBsAg by the polyclonal radioimmunoassay and for HBV DNA. PCR sequencing revealed a substitution of arginine for glycine at position 145 of HBsAg in the major neutralising epitope cluster, the a determinant, as well as a 2-aminoacid insertion of asparagine and threonine between positions 122 and 123, immediately upstream of this determinant.
Asunto(s)
Antígenos de Superficie de la Hepatitis B/genética , Virus de la Hepatitis B/genética , Hepatitis B/virología , Secuencia de Aminoácidos , Ensayo de Inmunoadsorción Enzimática , Resultado Fatal , Hepacivirus/inmunología , Anticuerpos Antihepatitis/aislamiento & purificación , Antígenos de Superficie de la Hepatitis B/aislamiento & purificación , Virus de la Hepatitis B/inmunología , Anticuerpos contra la Hepatitis C , Humanos , Masculino , Persona de Mediana Edad , Datos de Secuencia Molecular , Mutación , Reacción en Cadena de la Polimerasa , Proteínas del Envoltorio Viral/genéticaRESUMEN
BACKGROUND: Testing for antibody to hepatitis B core antigen (anti-HBc) as a surrogate for hepatitis C viremia is no longer needed for blood donor screening. Currently, the important question is how much its use supplements hepatitis B surface antigen (HBsAg) donor screening in preventing transfusion-transmitted hepatitis B virus (HBV) infection. STUDY DESIGN AND METHODS: In a study conducted in the 1970s, 64 blood donors were associated with 15 cases of HBV (1.0%) in 1533 transfusion recipients. Sera from 61 donors at donation and 29 follow-up visits were available for present-day assays for HBsAg, HBV DNA, anti-HBc, and antibody to HBsAg (anti-HBs). RESULTS: HBsAg was found in four previously negative blood donors; HBV DNA was limited to three of these four. Anti-HBc was detected in six HBsAg-negative donors. Two other donors were negative in all assays at donation, but positive for anti-HBc and anti-HBs 2 to 4 months later. The remaining donors were negative for all HBV markers, which left five recipient cases unexplained. No HBV transmission was observed when anti-HBs sample-to-negative control values were > or = 10. CONCLUSION: Some 33 to 50 percent of cases of hepatitis B that could be transmitted by transfusion of blood from HBsAg-negative donors are prevented by anti-HBc screening. Anti-HBc-positive donors unequivocally positive for anti-HBs should be considered noninfectious for HBV and should be allowed to donate. Anti-HBc screening of paid plasmapheresis donors, supplemented by anti-HBs testing, would reduce the amount of HBV to be processed by virus inactivation and increase the content of anti-HBs in plasma pools.
Asunto(s)
Donantes de Sangre , Anticuerpos Antihepatitis/análisis , Antígenos del Núcleo de la Hepatitis B/inmunología , Hepatitis B/transmisión , Reacción a la Transfusión , Humanos , Reacción en Cadena de la PolimerasaRESUMEN
The serological endpoint of response in the treatment of chronic hepatitis B is the loss of hepatitis B virus DNA and HBeAg. Because the quantitative measurement of hepatitis B virus DNA in serum has been shown to be useful for monitoring and predicting response to interferon-alpha therapy, we decided to evaluate whether changes in HBeAg concentration could also be used in this manner. Twenty-nine patients who were initially positive for HBeAg and HBV DNA were serially evaluated for HBeAg concentration with a microparticle-capture enzyme immunoassay. HBeAg levels in serum were calculated by means of comparison with a standard curve of fluorescence rate vs. HBeAg concentration. The results, expressed in milliunits per milliliter, were compared with hepatitis B virus DNA levels determined by means of solution hybridization. The baseline HBeAg concentration proved to be the best independent predictor of response on stepwise Cox regression analysis (p = 0.026). Similar disappearance curves were observed for the two markers, although hepatitis B virus DNA became undetectable at an earlier interval in 13 of 16 cases (81%). In the 16 responders, a decline in HBeAg concentration of more than 90% was observed by wk 12 of therapy (mean +/- S.D., 95% +/- 13%). Nonresponders did not demonstrate such steep declines in HBeAg values by wk 12 (mean +/- S.D., 45% +/- 27%), and levels tended to increase at subsequent time points. We conclude that serial monitoring of HBeAg concentration with a technique that should be readily adaptable to clinical laboratories may be useful in the initial evaluation and monitoring of patients undergoing antiviral therapy.
Asunto(s)
ADN Viral/sangre , Antígenos e de la Hepatitis B/sangre , Virus de la Hepatitis B/genética , Hepatitis B/terapia , Interferón-alfa/uso terapéutico , Anticuerpos Antihepatitis/sangre , Hepatitis B/inmunología , Hepatitis B/microbiología , Antígenos e de la Hepatitis B/inmunología , Virus de la Hepatitis B/inmunología , Humanos , Técnicas para Inmunoenzimas , Interferón alfa-2 , Valor Predictivo de las Pruebas , Pronóstico , Proteínas Recombinantes , Análisis de RegresiónRESUMEN
OBJECTIVE: To investigate the possible interference with acute hepatitis B virus infection by co-infection with hepatitis C virus. DESIGN: Analysis of stored sera collected for transfusion transmitted viruses study in 1970s. SETTING: Four major medical centres in the United States. PATIENTS: 12 recipients of blood infected with hepatitis B virus. MAIN OUTCOME MEASURES: In 1970s, presence of antibodies in hepatitis B virus and raised serum alanine aminotransferase concentration; detection of antibodies to hepatitis C virus with new enzyme linked immunoassays. RESULTS: Five of the 12 patients were coinfected with hepatitis C virus. Hepatitis B surface antigen was first detected at day 59 in patients infected with hepatitis B virus alone and at day 97 in those coinfected with hepatitis C virus (p = 0.01); median durations of antigenaemia were 83 and 21 days respectively (p = 0.05), and the antigen concentration was lower in the coinfected patients. Alanine aminotransferase patterns were uniphasic when hepatitis B virus infection occurred alone (range 479-2465 IU/l) and biphasic in patients with combined acute infection (no value > 380 IU/l; p = 0.0025). Four coinfected recipients developed chronic hepatitis C virus infection. The fifth patient was followed for only four months. CONCLUSIONS: Acute coinfection with hepatitis C virus and hepatitis B virus inhibits hepatitis B virus infection in humans, and onset of hepatitis B may reduce the severity of hepatitis C virus infection but not frequency of chronicity. Alanine aminotransferase concentration showed a biphasic pattern in dual infection.
Asunto(s)
Hepatitis B/complicaciones , Hepatitis C/complicaciones , Enfermedad Aguda , Alanina Transaminasa/metabolismo , Hepatitis B/enzimología , Hepatitis B/inmunología , Antígenos del Núcleo de la Hepatitis B/análisis , Antígenos de Superficie de la Hepatitis B/análisis , Hepatitis C/enzimología , Hepatitis C/inmunología , HumanosRESUMEN
A unique hepatitis B virus (HBV) variant has been identified in a gibbon (Hylobates lar) which could be passed to a chimpanzee by experimental inoculation. This HBV variant had been shown to have no reactivity to a monoclonal anti-preS2 antibody (preS2 mAb 116-34) differentiating it from all human HBV specimens tested. This gibbon sera also was not recognized by an anti-preS1 mAb which binds the preS1 hepatocyte receptor region, amino acids 27-35. In this paper, we report the DNA sequence of the gibbon HBV PreS gene. The lack of preS2 mAb (116-34) binding can be explained by a unique nucleotide substitution of A for C in the second codon of the preS2 region leading to the replacement of glutamine with lysine. Two other unique changes were observed at the seventh and 24th amino acid positions in the preS2 gene leading to a substitution of a valine for threonine and alanine, respectively. Unlike all human derived HBV sequences in the preS1 region, the gibbon HBV had a glutamic acid instead of an aspartic acid at amino acid residue 27. Another unique substitution was a leucine for alanine at preS1 position 33. These amino acid changes in the gibbon HBV may explain its unique preS mAb reactivity.
Asunto(s)
ADN Viral/aislamiento & purificación , Virus de la Hepatitis B/genética , Hylobates/microbiología , Secuencia de Aminoácidos , Animales , Anticuerpos Monoclonales , Secuencia de Bases , Clonación Molecular , ADN Viral/genética , Genes Virales , Genoma Viral , Antígenos de Superficie de la Hepatitis B/sangre , Virus de la Hepatitis B/aislamiento & purificación , Datos de Secuencia Molecular , Oligodesoxirribonucleótidos , Pan troglodytes , Reacción en Cadena de la Polimerasa , Homología de Secuencia de AminoácidoRESUMEN
HCV is the principal etiologic agent of post-transfusion (PTH) hepatitis. The incidence and course of HCV hepatitis in liver transplant recipients is not well established. To resolve this information deficit, all records of recipients of single liver transplant (OLTx) between March 1986 and March 1990 at the University of Pittsburgh in whom both the donor and recipients' pre-OLTx sera were available (n = 516) were reviewed. All sera were assayed for HCV antibody using a second generation ELISA method developed by Abbott Laboratories. On the basis of the anti-HCV status of the donor and recipient pre-OLTx sera, four groups could be classified: group I (donor-, recipient-) n = 375; group II (donor-, recipient+) n = 111; group III (donor+, recipient-) n = 25; and group IV (donor+, recipient+) n = 5. Post OLTx liver biopsies were obtained for a clinical indication in 473 of these 516 patients. The prevalence of anti-HCV among recipients pre-OLTx was 22.5% (116/516) which is three times greater than the 5.8% (30/516) prevalence in the donors. Histologic hepatitis not ascribable to any cause other than HCV occurred in 76/516 (15%) recipients: 42 in group I; 28 in group II; 6 in group III and none in group IV. The overall risk of HCV hepatitis at 6 months, 1 year and 2 years post-OLTx was 4.8% (25/516), 7.6% (39/516) and 10.1% (52/516), respectively. At each of these time intervals, no significant difference between groups for the prevalence of HCV hepatitis was evident.(ABSTRACT TRUNCATED AT 250 WORDS)
Asunto(s)
Anticuerpos Antihepatitis/análisis , Hepatitis C/epidemiología , Hepatitis C/etiología , Trasplante de Hígado/efectos adversos , Donantes de Tejidos , Adulto , Ensayo de Inmunoadsorción Enzimática , Femenino , Hepatitis C/inmunología , Humanos , Incidencia , Masculino , Persona de Mediana Edad , Índice de Severidad de la Enfermedad , Trasplante HomólogoRESUMEN
The IgM and IgG antibody response to various hepatitis C virus (HCV) antigens was studied in 8 patients who acquired posttransfusion HCV infection. IgM anti-HCV was detectable in only 4 of these patients, coincident with (1 patient) or later than (3 patients) the IgG anti-HCV response. Seven patients had initially decreasing IgG anti-HCV titres, indicating passive transfer of antibodies from donor to recipient. All 8 patients showed active IgG seroconversion, as demonstrated by increasing IgG anti-HCV titres, on average, 75 days after infection. Five years after infection, all patients were still reactive for IgG anti-HCV antibodies and 7 were positive for HCV RNA by the polymerase chain reaction (PCR). Two of these PCR positive patients were also reactive for IgM anti-HCV. It is concluded that the serology of HCV infection does not follow the classical pattern of IgM response preceding detection of IgG. The IgM response may be absent, late, or persistent after HCV infection. The serological diagnosis of recent HCV infection should be based on the polymerase chain reaction or rising IgG titres in at least 2 sequential patient blood samples.
Asunto(s)
Anticuerpos Antihepatitis/biosíntesis , Hepatitis C/inmunología , Inmunoglobulina M/biosíntesis , Antígenos Virales , Hepacivirus/inmunología , Hepatitis C/diagnóstico , Hepatitis C/transmisión , Anticuerpos contra la Hepatitis C , Antígenos de la Hepatitis C , Humanos , Inmunoglobulina G/biosíntesis , Factores de Tiempo , Reacción a la TransfusiónRESUMEN
A semiautomated dot blot assay and cDNA polymerase chain reaction (PCR) were used to study longitudinal anti-hepatitis C virus (HCV) recognition patterns in relation to presence of HCV-RNA in transfusion recipients and their infectious donors. In 9 recipients, 4 different patterns of HCV infection were observed: (A) persistent HCV carriage accompanied by chronic hepatitis in 6, (B) acute resolved hepatitis, but persistent HCV replication in one, and (C) continuous HCV replication without hepatitis in one and (D) acute resolved hepatitis with clearance of infection in one. This last self-limited infection was characterized by the disappearance of HCV-RNA as well as anti-HCV reactivity. In contrast, antibody reactivity persisted in 7 of 8 patients with chronic HCV infection who could be followed until 1990. Seven of the 9 recipients developed antibodies to all recombinant peptides in dot blot assay; one became positive for anti-C33 and anti-core and one developed anti-core only. The sequence of appearance of antibodies differed among individual patients. In 7 patients with full anti-HCV recognition patterns, the sequence of events was (mean and limits in days after transfusion): onset of hepatitis at day 50 (22-74), seroconversion of anti-C33 at day 91 (59-129), anti-core at day 133 (54-203), and anti-C100 at day 143 (59-365). The incorporation of C33 and core proteins, in addition to C100, in the second generation anti-HCV ELISA enhanced the detection rate in the HCV-infected transfusion recipients from 7/9 (78%) to 9/9 (100%).(ABSTRACT TRUNCATED AT 250 WORDS)
Asunto(s)
Hepacivirus/aislamiento & purificación , Anticuerpos Antihepatitis/sangre , Hepatitis C/inmunología , Hepatitis C/microbiología , Biomarcadores , Ensayo de Inmunoadsorción Enzimática/métodos , Ensayo de Inmunoadsorción Enzimática/estadística & datos numéricos , Estudios de Evaluación como Asunto , Hepacivirus/genética , Hepatitis C/transmisión , Humanos , Immunoblotting , Reacción en Cadena de la Polimerasa , ARN Viral/sangre , ARN Viral/genética , Sensibilidad y Especificidad , Reacción a la TransfusiónRESUMEN
We performed a quantitative study of serum hepatitis B virus (HBV) markers, including new parameters such as pre-S1 antigen (Ag), pre-S2 Ag, and anti-HBx, in 88 chronic hepatitis B surface antigen (HBsAg) carriers. New IMx assays for HBsAg and immunoglobulin M (IgM) anti-HBc detection were also used. The population studied was composed of 65 chronic hepatitis cases (40 positive for hepatitis B antigen [HBeAg] and 25 positive for anti-HBe) and 23 anti-HBe-positive, asymptomatic HBsAg carriers. Serum HBsAg levels detected by IMx were higher in HBeAg-positive than in anti-HBe-positive HBsAg carriers (all patient subgroups included) and correlated with the serum HBV DNA level (P = 0.0001). Both pre-S1 and pre-S2 Ags were detected by enzyme immunoassays in almost all HBsAg carriers. Both pre-S1 and pre-S2 Ag titers correlated positively with the serum HBsAg concentration (P = 0.0001), but only the pre-S1 Ag titer correlated with the level of serum HBV DNA (P = 0.02). The detection of low levels of IgM anti-hepatitis B core (anti-HBc) antibodies by IMx was associated with the presence of liver disease (P = 0.05) but not with the level of viral replication. The prevalence of anti-HBx antibodies detected by the enzyme immunoassay was slightly, although not significantly, higher in patients with high levels of HBV DNA (greater than 100 pg/ml) than in patients without detectable HBV DNA (P = 0.16). In anti-HBe-positive chronic HBsAg carriers, the quantitative detection of serum HBV DNA, pre-S Ag titers, and IgM anti HBc allowed us to predict which patients suffered from chronic liver disease and/or supported viral replication (P < 0.05). In a follow-up study of eight patients undergoing antiviral therapy, the clearance of both pre-S1 Ag and HBV DNA was associated with a subsequent clearance of HBV. Therefore, the quantitative determination of HBV DNA, pre-S Ags, IgM anti-HBc may prove useful for the decision to use and the monitoring of antiviral therapy, especially in anti-HBe-positive HBsAg carriers.
Asunto(s)
ADN Viral/análisis , Anticuerpos contra la Hepatitis B/análisis , Antígenos de Superficie de la Hepatitis B/análisis , Hepatitis B/inmunología , Inmunoglobulina M/análisis , Precursores de Proteínas/análisis , Portador Sano , Enfermedad Crónica , Hepatitis B/tratamiento farmacológico , Antígenos e de la Hepatitis B/análisis , Virus de la Hepatitis B/genética , Humanos , Replicación ViralRESUMEN
Serological markers for hepatitis C virus (HCV) infection were measured in serial samples from 14 posttransfusion chronic non-A, non-B hepatitis patients by a semiquantitative dot blot immunoassay. The assay detected antibodies to HCV by use of recombinant proteins that represent putative HCV capsid (core), nonstructural protein 3 (NS3) (33c), and NS4 (c100) epitopes. Seroconversion to anti-HCV antibodies (anti-HCV) was detected in all patients. The average time to active antibody production detected by any of the recombinant proteins was 13.8 (range, 3.6 to 22.0) weeks posttransfusion or 4.6 (range, -4.5 to 13.4) weeks after the first biochemical marker of illness. Anti-HCV were detected earliest by the core antigen in most cases; however, the patterns of anti-HCV responses varied significantly among individuals. Overall, the addition of the core and NS3 antigens to the assay enabled the detection of the antibody response 4 to 5 weeks earlier than did the addition of the c100 antigen, the sole antigen used in current screening tests in the United States. Passively transferred antibodies were detected by at least one antigen in early posttransfusion samples from 12 patients and decayed below detectable levels for all antigens in only 2 patients. Antibodies to all three gene products were evident in the last sample from all five patients monitored for greater than 3 years from transfusion indicating the persistence of antibodies in patients with chronic illness. Our data show that the period following the onset of hepatitis during which anti-HCV are not detected by current screening assays can be greatly shortened by the detection of anti-HCV responses by a combination of core, NS3, and c100 antigens.
Asunto(s)
Hepatitis C/inmunología , Hepatitis C/transmisión , Reacción a la Transfusión , Antígenos Virales , Biomarcadores , Hepacivirus/inmunología , Anticuerpos Antihepatitis/sangre , Anticuerpos contra la Hepatitis C , Humanos , Inmunización Pasiva , Factores de TiempoRESUMEN
Liver enzyme levels, viral RNA, and the immune response against both structural and nonstructural hepatitis C virus (HCV) proteins have been studied in experimentally infected chimpanzees in order to further understand the natural history of HCV infection. An ELISA for measuring both IgG and IgM responses to core (c22), 33c (NS3), and c100 (NS4) was employed. The IgG response rates were 5/8 for core, and 8/8 for both 33c and c100. Utilizing this antigen combination, at least one antibody response is measureable at, or within 3 weeks of, the major ALT peak. Although no individual antibody response is universally associated with initial detection of seroconversion, the combination of all three recombinant proteins measures seroconversion an average of 54 days earlier than with c100 alone, in 6/8 of the animals. IgM responses were measureable in 5/8 of the chimpanzees, were of shorter duration, and usually arose concomitantly with IgG responses. IgM appears to be a good indicator of primary infection since neither boosting nor recrudescence of disease during the chronic phase of disease elicited a secondary IgM response. Viral RNA can be measured 4-7 days (average = 9 days) postinfection with the period preceding the ALT peak being characterized by several PCR positive segments interrupted by periods in which no viral RNA can be measured. Following the ALT peak, chronically infected animals with recurring ALT elevations are generally PCR positive with intercedent PCR negative periods. Those animals that appear to have biochemically resolved disease generally have PCR negative profiles, although they still may periodically exhibit PCR positive sera. This indicates that with the recent advent of new screening techniques, a more stringent definition of HCV resolution will be required.
Asunto(s)
Hepacivirus/inmunología , Anticuerpos Antihepatitis/sangre , Hepatitis C/sangre , ARN Viral/sangre , Animales , Secuencia de Bases , ADN Viral/sangre , Hepacivirus/metabolismo , Anticuerpos Antihepatitis/inmunología , Hepatitis C/inmunología , Hepatitis C/patología , Anticuerpos contra la Hepatitis C , Inmunoglobulina G/inmunología , Inmunoglobulina M/inmunología , Leucocitos Mononucleares , Hígado/patología , Datos de Secuencia Molecular , Pan troglodytes , Reacción en Cadena de la Polimerasa , Factores de TiempoRESUMEN
IgM antibody against hepatitis C virus (IgM anti-HCV) was measured in serial samples from 15 transfusion recipients in whom posttransfusion chronic non-A, non-B hepatitis (NANBH) developed and three plasmapheresis donors during acute HCV infection using recombinant proteins derived from three immunodominant regions: core, NS-3, and NS-4 (c100). IgM anti-HCV core was detected in 13 of 15 posttransfusion patients. Nine of these patients had transient, acute-phase IgM anti-HCV core detected coincidentally or earlier than active IgG anti-HCV core response. The average duration of IgM anti-HCV core reactivity was 8.1 +/- 3.7 weeks. One patient lacking an IgM anti-HCV core response had detectable IgM anti-HCV NS-3 during the acute phase. Passive transfer of IgM anti-HCV was not observed in these posttransfusion cases, in contrast to the high frequency observed for IgG anti-HCV. Late IgM anti-HCV was detectable against core, c100, and NS-3 in three, two, and one posttransfusion patients, respectively. These data indicate that IgM anti-HCV core is a useful acute-phase marker in HCV infection.