Several types of naturally occurring pre-S mutants in sera or liver tissues in patients with chronic hepatitis B virus (HBV) infection have been identified. To clarify the prevalence and significance of emergence of pre-S mutants, 140 sera and 18 resected livers from patients with HBV were studied. Replicative status was designated as high, intermediate, and low based on the HBV-DNA levels in serum or the expression of HBV antigens in liver. In vitro transfection and Western blot analysis were performed to characterize expression and secretion of HBsAg by the mutant constructs. Five major types (I to V) of pre-S deletion mutants in serum and liver and 2 types (VI and VII) in liver were identified. Pre-S mutant was 6.4% at high replicative phase, 13% at intermediate, and 37.5% at low or nonreplicative phases in serum. In livers, the same tendency existed: pre-S2 deletion mutants emerged and prevailed at a low replicative phase in hepato- Hepatitis B virus (HBV) is a small, enveloped DNA virus that causes acute and chronic liver diseases. The majority of acute HBV infection are usually self-limited, whereas patients with chronic HBV infection usually pursue a life-long course that can be categorized into 3 phases based on the replicative status of HBV in serum and liver. [1][2][3][4][5][6][7][8][9] In the early or high replicative phase, there are high levels of hepatitis B e antigen (HBeAg), hepatitis B surface antigen (HBsAg), and HBV DNA in serum and an active replication of HBV genome with nuclear expression of core antigen (HBcAg) in liver. 3,[8][9][10] The HBsAg-expressing hepatocytes or ground glass hepatocytes (GGH) are usually singly scattered in distribution. In the intermediate or immune clearance phase, there is an HLA-restricted cytotoxic T lymphocyte (CTL) response to viral antigen-expressing hepatocytes, which may result in a decrease of HBeAg and HBV DNA in serum. In the late or nonreplicative phase, there is a low titer or absence of viremia and an integration of HBV genomes in host cells. The serum HBsAg, albeit decreased in level, usually persists for life. Contrary to that at the active replicative stage, the HBsAg-containing hepatocytes are inversely increased and clustered in groups at the late or nonreplicative stage, usually associated with an absence of intrahepatic HBcAg expression. [9][10][11] The underlying mechanism of the life-long persistence of serum HBsAg and the novel discrepancy between the levels of HBsAg in serum and liver have not yet been clarified. In the past years, many studies have documented the accumulation of pre-S1 or large surface antigen in the hepatocytes during chronic HBV infection. [12][13][14][15] The large (pre-S1), middle (pre-S2), and small (major) surface proteins have been found to be differentially expressed in hepatocytes at different stages of HBV replication. 13,14 Most liver samples from viremic carriers show a prevalence of 2 smaller surface proteins whereas the large surface antigen is the predominant intrahepatic protein in nonviremic carriers. 14 ...