, Chicago, IL). Student’s t test was performed to compare continuous variables. A two-tailed P value of <0.05 was considered statistically significant. RESULTS Replacement of dominant species of HDV in CHD. To determine the selection of quasispecies in patients, large-scale screening of at least 100 colonies was carried out by using variant-specific selleck bio RFLP analysis. Percentages of the original HDV dominant species of genotypes 1, 2, or 4 in the early stage of the disease course were very high (about 90 to 95%) (Fig. 1). Following marked elevations of ALT, the percentages of the original HDV dominant species decreased and they finally became minor ones. In contrast, the percentages of the novel HDV dominant species increased from preexisting minor variants in the original quasispecies after hepatitis flares.

Fig 1 Replacement of the original dominant species by novel dominant species during the disease courses of three patients chronically infected with different genotypes of HDV. At least 100 HDV colonies were randomly selected by large-scale screening carried … Comparison of viral replication and HDAg expression levels of the original and novel dominant HDV species. To clarify if the novel dominant species emerged because of a growth advantage, seven pairs of original and novel dominant quasispecies separated by hepatitis flares were cloned from seven patients (labeled I to VII) and cotransfected with an HBV-producing plasmid (a genotype B or C HBV-producing plasmid according to the patient’s clinical data) into the Huh-7 human hepatoma cell line to evaluate the replication activity of the dominant HDV strains at early and late time points of the clinical courses of different patients.

HBV serves as the helper for HDV virion production. Representative patients II and III, who went into biochemical remission in the absence of cirrhosis or HCC, are first illustrated in Fig. 2A and andB.B. These two patients had been treated with short-acting alpha interferon 2b at 5 million IU thrice weekly for 1 year. Their serum ALT levels remained elevated, and they fluctuated after interferon treatment in patient II. Remission did not occur until more than 110 months after treatment was stopped and was unlikely to have been the result of direct effects of treatment (Fig. 2A). In patient III, remission occurred immediately after interferon treatment (Fig. 2B).

In patient I, remission occurred 26 months after interferon treatment (Fig. 3A). The intracellular HDV replication, HDAg expression, and secreted HDV virions of the novel dominant HDV strains late in the clinical course were lower Dacomitinib than those of the original dominant strains (Fig. 2C and andD).D). The novel dominant HDV species with a lower relocation capacity was detected about 90 months after the cessation of interferon treatment and 24 months before remission in patient II and about 8 months before the start of interferon treatment, about 15 months before remission, in patient III (Fig.