Hepatitis B virus DNA (HBV DNA) in serum was measured by a Spot hybridization technique in a consecutive series of 79 cases with chronic HBV infection from Taiwan. HBV DNA was found in 96.3% (52/54) of HBeAg-positive, 66% (2/3) with neither HBeAg or anti-HBe and in 63.6% (14/22) of anti-HBe positive patients. The levels of HBV DNA in the HBe-Ag-positive patients were significantly higher than in the anti-HBe positive patients (median, 944 vs. 58 pg per ml, p less than 0.001). The mean ages increased from 28.7 years for the cases with high levels of HBV DNA, to 34.7 years for those with low levels (p less than 0.01) and to 41.0 years in those without HBV DNA in serum (p less than 0.05 when compared with those with low level of HBV DNA). Ninety per cent of patients (27/30) with high levels of HBV DNA showed only minor hepatic inflammatory activity, as did 91% (10/11) of those without HBV DNA. In contrast, histologic signs of chronic active hepatitis or chronic lobular hepatitis were demonstrated in 76% of cases (29/38) with low levels of HBV DNA. These data are consistent with the hypothesis that liver damage occurs during the period of clearance of hepatocytes supporting HBV replication, and are inconsistent with the view that HBV may be directly cytopathic. Thus, the natural history of chronic HBV infection may be divided into three phases.(ABSTRACT TRUNCATED AT 250 WORDS)

The hypervariable region (HVR) of the E2/NS1 region of hepatitis C virus (HCV) varies greatly between viral isolates with high rates of genomic change reported during the course of chronic infection. The HVR is thought to encode a structurally unconstrained envelope protein containing several linear B cell epitopes recognized by neutralizing antibody. It has been postulated that amino acid changes in the HVR could result from humoral immune pressure leading to the selection of escape mutants. The aim of this study was to compare the rates of nucleotide and amino acid variation in the HVR of control patients to patients with common variable immunodeficiency (CVID) where the effect of the humoral immune system is reduced. Five controls and four patients with CVID were studied. Serum samples were taken over periods of between 1 and 6 years. HCV was detected by polymerase chain reaction (PCR) with primers derived from conserved flanking regions of the HVR. PCR products were cloned into a plasmid vector and recombinant clones identified by restriction enzyme digestion. Purified DNA from at least three individual clones from each time point was sequenced by the dideoxynucleotide chain-termination method. Consensus sequences were extracted from the three clones, and the DNA and deduced protein sequences were compared. Control patients had a mean rate of nucleotide change of 6.954 nucleotide substitutions per year, compared with patients with CVID with a rate of 0.415 nucleotide substitutions per year (P F .02). The corresponding rates for amino acid variation were 3.868 amino acid substitutions per year for the control patients compared with 0.185 amino acid substitutions per year for the patients with CVID. These findings suggest that in the absence of humoral immune selective pressure, the frequency of occurrence of genetic variation in the major viral species is reduced. The mutations occur, but in the absence of immune selection remain as minor species. The evolution of viral mutants capable of evading the host's immune system may contribute to the ability of HCV to establish chronic infection. (HEPATOLOGY 1998;26:223-227.)Genomic analysis has shown hepatitis C virus (HCV) to be distantly related to both flaviviruses and pestiviruses with structural genes at the 5Ј end preceding those encoding the nonstructural proteins. Analysis of HCV isolates from around the world has revealed extensive heterogeneity between isolates, although the sequence variation is not distributed evenly throughout the genome. In particular, a 27-amino acid region at the amino terminus of E2/NS1 has been shown to vary greatly between isolates and has become known as hypervariable region 1 (HVR). 1-3 The HVR appears to be structurally unconstrained and tolerates extensive amino acid variation; it probably encodes part of an envelope peptide that is exposed to the host' s immune response. 4 Studies have indicated the presence within this region of linear B-cell epitopes recognized by neutralizing antibody, and amino acid variation occurring du...

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Hepatitis delta virus (HDV) infection of individuals infected with hepatitis B virus (HBV)is associated with more severe liver damage and an increased risk of fulminant disease. HDV is a single-stranded RNA virus that encodes a single protein, the delta antigen, which is expressed in two forms, small (S-HDAg) and large (L-HDAg). Here we show that although HDV ribonucleoproteins are mainly detected in the nucleus, they are also present in the cytoplasm of cells infected with HDV or transfected with HDV cDNA. Making use of an heterokaryon assay, we demonstrate that HDV ribonucleoproteins shuttle continuously between the nucleus and the cytoplasm. In the absence of HDV RNA, both forms of the delta antigen are retained in the nucleus, whereas in the absence of the delta antigen, HDV RNA is predominantly detected in the cytoplasm. Coexpression of HDV RNA and S-HDAg (which binds to the viral RNA and contains a nuclear localization signal) results in nuclear accumulation of the viral RNA. This suggests that HDV RNA mediates export of viral particles to the cytoplasm whereas the delta antigen triggers their reimport into the nucleus.

Hepatitis delta virus (HDV) is a human pathogen that can greatly increase the severity of liver damage caused by an hepatitis B infection. HDV contains a circular, single-stranded RNA genome that encodes a unique protein, the delta antigen. Two forms of the delta antigen, dAg-S and dAg-L, are derived from a single open reading frame by RNA editing. Here we analyze the subcellular distribution of HDV RNA and its spatial relationship to known intranuclear structures. The human hepatoma cell line Huh7 was stably transfected with wild-type HDV cDNA and the viral RNAs were localized by in situ hybridization and fluorescence confocal microscopy. HDV RNA is detected throughout the nucleoplasm, with additional concentration in focal structures closely associated with nuclear speckles or clusters of interchromatin granules. Both the smaller form of the delta antigen (dAg-S), which is required for HDV genomic replication, and the larger form of the delta antigen (dAg-L), which represses replication, co-localize with delta RNA throughout the nucleoplasm and in the foci. However, the foci do not incorporate bromo-UTP and do not concentrate either RNA polymerase II or cleavage and polyadenylation factors required for viral RNA synthesis and 39 end processing, respectively. Thus, it is unlikely that the delta foci represent major sites of viral transcription or replication. In conclusion, the data show that viral RNA-protein complexes accumulate in structures closely associated with interchromatin granules, a subnuclear domain highly enriched in small nuclear ribonucleoproteins, poly(A + ) RNA, and RNA splicing protein factors. This implies a specific compartmentalization of ribonucleoproteins in the nucleus.