Genetic variability of hepatitis C virus in chronically infected patients with viral breakthrough during interferon-ribavirin therapy.

Little is known about hepatitis C virus (HCV) breakthrough during antiviral therapy, although it would help in understanding HCV resistance to current antiviral treatments. To analyse the implication of virological factors and the vigour of humoral immune responses in this phenomenon, we studied nine chronic hepatitis C patients with a viral breakthrough during IFN/ribavirin combination therapy, as well as five responders and five non-responders. The IRES and regions coding for the capsid protein, the PePHD domain of envelope glycoprotein E2 and the NS5A and 5B proteins were amplified by RT-PCR before treatment, before and during breakthrough, and after treatment. The major variant sequence was obtained by direct sequencing. The heterogeneity of quasispecies was studied by SSCP in all patients and sequencing after cloning in seven genotype 1b-infected patients. Humoral responses against HCV epitopes were also analysed. The major sequences of IRES, PePHD, and NS5B remained stable during treatment, regardless of the treatment response. However, the capsid protein and the regions flanking PePHD showed sequence variations in breakthrough patients, although no specific mutation was identified. The variable V3 region of NS5A, but not the PKR-binding domain and the ISDR, seemed to be associated with differences in response to treatment. The analysis of HCV quasispecies revealed no characteristic pattern during treatment in breakthrough patients, whose HCV genome profiles looked most similar to that of non-responders. The humoral response was similar between groups. In conclusion, viral breakthrough does not seem to be due to selection of resistant strains with signature mutations

Validation of the INNO-LIA Syphilis Kit as a Confirmatory Assay for <i>Treponema pallidum</i> Antibodies

ABSTRACT The commercially available diagnostic tests for syphilis are mostly based on the use of extracted antigens of Treponema pallidum . Pronounced cross-reactivities with other spirochete antigens are often reported. The aim of this study was to validate a novel multiparametric assay (the assay performed with the kit) INNO-LIA Syphilis for the confirmation of syphilis antibodies in a set of 840 documented human serum samples. All serum samples were previously tested at the French World Health Organization reference center for venereal diseases (Institute Alfred Fournier, Paris, France), with a consensus result provided for each sample. The study was conducted in two phases, with each phase involving a validation set (500 well-documented serum samples) and an exploratory set (340 serum samples) of serum samples, respectively. By measuring the sensitivity and specificity, we compared the result of the new assay with the consensus result on the basis of the results of a variable number of classical serological methods and clinical information when available. A sensitivity of 99.6% (95% confidence internal [CI], 98.5 to 99.9%) and a specificity of 99.5% (95% CI, 98.1 to 99.9%) were found for the new line immunoassay. Six of seven samples with indeterminate results by classical serology tested positive with the INNO-LIA Syphilis kit. This single multiparametric assay provides reliable confirmatory diagnostic information that must currently be obtained by the performance and interpretation of results of a combination of serological assays. </jats:p