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An association between the presence of HCV-Ab and immune reconstitution after HAART has been shown elsewhere. Results of our large, prospective study support a direct role of HCV viremia in the CD4+ cell count response to HAART. Moreover, our results underline the fact that, in individuals coinfected with HIV and HCV, the goal of treating HCV infection is to eradicate HCV, to both slow the rate of HCV progression and limit potential interference with the response to HAART.

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We describe a simple, sensitive, and reproducible method for using whole blood collected onto filter paper (dried blood spots) for detection and genotyping of hepatitis C virus RNA that can be useful in large field studies, particularly in settings where collection, preparation, storage, and shipment of samples at controlled temperature can be difficult.Dried blood spots (DBS) have been used worldwide for the neonatal screening of congenital disorders (2, 8). Seroepidemiological studies have been conducted on DBS residual to neonatal screening to assess human immunodeficiency virus (HIV) prevalence among childbearing women (9, 10). Recently, several studies were focused on DBS for detecting drug resistance mutations (6) and for tracking global spreading of HIV type 1 subtypes (4) in proviral HIV DNA. However, RNA is notoriously less stable, and standardization of DBS for viral RNA detection would be of great benefit for application to large field studies, since DBS collection is easy, does not require skilled phlebotomists and expert technicians, and is suitable for storage and shipment to laboratory in settings where these issues are problematic. Recently, dried plasma spots and DBS have been used for HIV RNA detection and quantification, showing good correlation with titers obtained with conventional plasma samples (5, 11). However, these observations were limited to short storage at room temperature (11) or at 37°C (5), and a loss of viral titers occurred during storage.For hepatitis C virus (HCV) RNA detection on dried spot samples, the available data are much less exhaustive. A complete match between frozen serum and dried plasma spots, though with a loss of titers after room temperature storage, has been observed (1).This study was aimed at developing a simple, sensitive, and reproducible method for using DBS in HCV RNA detection and genotyping.The study complied with all relevant national guidelines and institutional policies. Residual laboratory samples of EDTAwhole blood of 39 HCV antibody (Ab)-positive and 16 HCV Ab-negative patients, undergoing routine hematological controls, were used. HCV Abs were determined by third-generation assay (Abbott Diagnostics). Among the Ab-positive patients, 34 had HCV RNA levels ranging between 9,640 and 5,100,000 IU/ml (Amplicor HCV Monitor; Roche Molecular Systems Inc.), and 5 were HCV RNA negative (Versant HCV TMA; Bayer Diagnostic Inc.). The HCV genotype was known for eight patients (four had 1b; two had 2a/2c; one had 3a; and one had 4c/4d).DBS were realized within 5 h from venipuncture by carefully spotting, in multiple replicates for each patient, 50 l of EDTA-whole blood on SS grade 903 filter paper (Schleicher & Schuell Inc.). Two DBS from each patient were pooled and processed for each assay.HCV RNA detection was performed with both in-house reverse transcriptase PCR (RT-PCR) and transcription-mediated amplification (TMA). For RT-PCR, RNA extraction was performed with Boom technology, utilizing silica-based RNA isolation (3), which was purchased from Organon ...