Vigorous proliferative CD4(+) T cell responses are the hallmark of spontaneous clearance of acute hepatitis C virus (HCV) infection, whereas comparable responses are absent in chronically evolving ...infection. Here, we comprehensively characterized the breadth, specificity, and quality of the HCV-specific CD4(+) T cell response in 31 patients with acute HCV infection and varying clinical outcomes. We analyzed in vitro T cell expansion in the presence of interleukin-2, and ex vivo staining with HCV peptide-loaded MHC class II tetramers. Surprisingly, broadly directed HCV-specific CD4(+) T cell responses were universally detectable at early stages of infection, regardless of the clinical outcome. However, persistent viremia was associated with early proliferative defects of the HCV-specific CD4(+) T cells, followed by rapid deletion of the HCV-specific response. Only early initiation of antiviral therapy was able to preserve CD4(+) T cell responses in acute, chronically evolving infection. Our results challenge the paradigm that HCV persistence is the result of a failure to prime HCV-specific CD4(+) T cells. Instead, broadly directed HCV-specific CD4(+) T cell responses are usually generated, but rapid exhaustion and deletion of these cells occurs in the majority of patients. The data further suggest a short window of opportunity to prevent the loss of CD4(+) T cell responses through antiviral therapy.
Lamivudine (LAM) is associated with the highest known rate of resistance mutations among nucleotide analogs used to treat chronic hepatitis B virus (HBV) infection. Despite this, LAM continues in ...widespread use, especially in combination therapies. The primary LAM resistance mutation (rtM204V/I) occurs in the YMDD motif of HBV polymerase. The aim of this study was to characterize Brazilian HBV isolates from acute and chronic cases by direct sequencing, and to identify HBV quasispecies in the YMDD motif using a pyrosequencing method capable of detecting single-nucleotide polymorphisms. HBV DNA from serum samples of 20 individuals with acute HBV infection and 44 with chronic infection undergoing antiviral therapies containing LAM were analyzed by direct sequencing and pyrosequencing methods.
Phylogenic analyses of direct-sequenced isolates showed the expected genotypes (A, D and F) for the Brazilian population in both acute and chronic infections. However, within genotype A isolates, subgenotype A2 was more frequently detected in acute cases than in chronic cases (P = 0.012). As expected, none of the individuals with acute hepatitis B had LAM-resistant isolates as a dominant virus population, whether detected by direct sequencing or pyrosequencing. However, pyrosequencing analyses showed that 45% of isolates (9/20) had minor subpopulations (4-17%) of LAM-resistant isolates. Among chronic patients undergoing LAM treatment, YMDD mutants were frequently found as a dominant virus population. In cases where wild-type virus was the dominant population, subpopulations of YMDD variants were usually found, demonstrating the complexity of HBV quasispecies.
YMDD variants were frequently detected as a minor population in acute HBV infection. The occurrence of pre-existing variants may lead to a high frequency of resistant mutants during antiviral therapy in the chronic phase. In chronic infection, detection of YMDD variants before virological or biochemical breakthrough might contribute to making better therapy choices and thus improving treatment outcome.
We evaluated the stability of hepatitis B virus (HBV) DNA in plasma samples stored at 42°C for external quality assessment (EQA) panels of viral load. To assess the stability of plasma samples ...containing different concentrations of HBV DNA, serial dilutions of HBV-infected samples with a viral load of 6.40 log(10) IU/mL were made to yield viral loads of 5, 4, and 3 log(10) IU/mL. These were incubated at 42°C for up to 7 days and then frozen at -70°C. Viral load testing for HBV DNA was performed for all samples using COBAS® AmpliPrep/COBAS® TaqMan® HBV Test (v.2.0, Roche, Switzerland). Results were compared with fresh frozen plasma samples as a benchmark to establish acceptable measurements on the days following sample collection. Although the results of this study demonstrated a decrease in HBV DNA viral load ranging from 0.005 to 0.30 log(10) IU/mL after storage at 42°C for up to 7 days, these values did not exceed 0.5 log(10), which is the estimated intra-assay variation for molecular tests. Thus, the insignificant decrease in viral load suggests that shipment of HBV in plasma samples at temperatures of up to 42°C is permissible if they are frozen within 7 days.
We evaluated the accuracy of a commercial rapid immunochromatographic test (rapid test RT) for hepatitis A (HA) diagnosis and epidemiological studies.
The accuracy of a RT was evaluated in laboratory ...and in field conditions. Predictive modeling estimated the test performance in a hypothetical population.
The RT showed sensitivities of 66-86%, and specificities of 21-100%, depending on the antibody isotype (IgM or IgG) analyzed and prevalence of infection.
The RT is a good alternative for diagnostic in HA outbreaks. The predictive model indicates that it should not be used alone for HA diagnosis in low prevalence populations. These data can be used in the future to strengthen decision-making during the implementation of rapid diagnostic methods in health services.
AbstractBackground. The relationship between 25-hydroxyvitamin D 25(OH)D serum levels and response to antiviral therapy and laboratory data in HCV infection remains unclear. The aim of this study was ...to determine pre-treatment 25(OH)D serum level among HCV infected individuals and to evaluate the association between vitamin D status, virological response, and laboratory data. Material and methods. Baseline serum 25(OH)D levels were measured in 237 chronic HCV infected patients (139 female, age 53.7 ± 11.2 years) using chemiluminescence immunoassay. Correlations between serum 25(OH)D levels, virological and laboratory data regarding HCV infection as well as sustained virological response (SVR) to antiviral therapy were evaluated. Results. Mean serum values of 25(OH)D was 26.2 ± 12 ng/mL and prevalence of vitamin D deficiency (< 30 ng/mL) was 66.2%. Advanced age (> 55 years), high mean values of LDL, total cholesterol, HDL and low mean values of alkaline phosphatase and hemoglobin were statistically associated to vitamin D deficiency. Antiviral treatment was underwent by 133 HCV patients and 44.3% of them achieved SVR. Most of individuals that presented SVR also presented 25(OH)D level higher than 30ng/mL (55.9%). SVR was associated to low mean values of LDL, total cholesterol and platelets; high mean values of ALT, AST and low fibrosis grade. Conclusions: In conclusion, low vitamin D levels were observed among HCV infected patients and was associated to laboratory findings, however baseline 25(OH)D level is not independently associated with SVR.
Background: Hepatitis A virus (HAV) infection is the leading cause of clinically apparent viral hepatitis in many parts of the world, including developed and developing countries. Only limited ...information is available regarding the seronegative viremic window that follows HAV infection, and no systematic search has been reported for HAV RNA positive, IgM anti-HAV negative serum samples during hepatitis A outbreaks.
Objectives: To determine the proportion of HAV infected individuals among (i) children who were tested negative for anti-HAV antibodies during hepatitis A outbreaks which occurred in a public school (
n=157) and a child care center (
n=38); (ii) subjects (
n=46) initially classified as acute non-A–C hepatitis patients after clinical examination and serological tests (sporadic cases).
Study design: Reverse transcription (RT)-PCR was performed to detect the presence of HAV genome in serum samples collected from anti-HAV negative, susceptible subjects.
Results: HAV RNA was detected in 19/157 (12%) and 5/38 (13%) anti-HAV negative children from the public school and child care center, respectively. Twelve (26%) out of the 46 acute hepatitis patients (sporadic cases) were also HAV RNA positive. From nine of these 12 patients, a second blood sample was obtained 18–34 days after the first one: all nine had seroconverted to IgM anti-HAV, and their serum transaminases had reached elevated levels (mean ALT, 418; mean AST, 241).
Conclusions: Detection of HAV RNA before IgM anti-HAV seroconversion may be used as an early diagnosis method during hepatitis A outbreaks. HAV RNA testing should also help to elucidate acute hepatitis cases of unknown etiology.
Zika virus (ZIKV) is a flavivirus that is closely related to other human pathogens, such as dengue virus (DENV)
. Primary transmission usually involves Aedes aegypti, which has expanded its ...distribution range considerably
, although rarer infection routes, including mother-to-fetus transmission, sexual contact and blood transfusion, have also been observed
. Primary ZIKV infection is usually asymptomatic or mild in adults, with quickly resolved blood viraemia, but ZIKV might persist for months in saliva, urine, semen, breast milk and the central nervous system
. During a recent ZIKV outbreak in South America, substantial numbers of neurological complications, such as Guillain-Barré syndrome, were reported
together with cases of microcephaly and associated developmental problems in infants born to women infected with ZIKV during pregnancy
, highlighting the clinical importance of this infection. Analyses of the human immune response to ZIKV are lacking
, but the recent outbreak has provided an opportunity to assess ZIKV immunity using current immunological methods. Here, we comprehensively assess the acute innate and adaptive immune response to ZIKV infection in ten women who were recruited during early infection and followed through reconvalescence. We define a cascade of events that lead to immunological control of ZIKV, with previous exposure to DENV impacting some, but not all, mediators of antiviral immunity.
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