Figure 2.

HCV RNA association with infectious B cells. (A) Selected virions are infectious after passage in new uPA-SCID-mice. Chimeric mice were injected with 10⁴ IU of mouse-passaged virions originating after B cell mediated HCV transmission (marked with prefix ‘m’ and ID of the originating mouse). HCV RNA levels (IU/ml) were measured in mouse plasma and plotted against time (days, X-axis). The animal injected with mP09_K1461R died before the end of the observation period. All animals demonstrate high viral replication detectable as soon as one week after viral inoculation. (B) Incubating B cells from a naïve donor with infectious plasma does not transmit infection. Chimeric mice were challenged (IS) with 10⁶ B cells derived from a healthy individual (B_healthy) that were pre-incubated (37°C, 2h) with 5x10⁴ IU infectious mK983 virions alone (B_healthy + mK983) or immune-complexed with 100 µg autologous IgG (B_healthy + mK983 + IgG_P12) and subsequently washed to remove non-adherent virions. As controls, the infectivity of B cells from patient P12 (B_P12) and of 10⁴ IU mouse-derived virions (mK983) is shown. Limit of HCV-RNA quantification (LOQ) is 750 IU/ml (dotted horizontal line). (C) Detection of (+) and (-) strand HCV RNA in infectious and non-infectious B cells. Graphic presentation of (+)ssRNA and (-)ssRNA detection in 3 non-infectious (upper panels) and 5 infectious B cells using the Tth-based RT-PCR-method as described. Bars represent the absolute amounts of B cells tested, normalized for total RNA extraction (y-axis). Progressive dilution of B cells is represented on the x-axis. Black bars refer to the simultaneous detection of (+) and (-)ssRNA; grey bars to single (+)ssRNA detection and empty bars to RNA not detectable ( Supplementary Figure 5B ).