<p>The immunopathogenesis of chronic hepatitis B (CHB) with unclear immune phases hinders functional cure. For highly infectious HBeAg-positive (HBeAg⁺) patients where functional cure is unattainable, achieving sustained HBeAg seroconversion constitutes the key therapeutic objective. This study used single-cell sequencing (scRNA-seq) to analyze B-cell subsets dynamics during HBeAg seroconversion and their roles in restricting HBsAg seroconversion. ScRNA-seq datasets (GSE182159, GSE234241) were retrieved from the GEO database, comprising 67 peripheral blood/liver samples. After batch correction and integration, compared with the two HBeAg⁺ phases, patients with HBeAg-negative (HBeAg⁻) chronic HBV infection show opposite distribution trends of naive and memory B cells in peripheral blood and liver. Compared with healthy controls and chronic resolved (CR) cases, the HBeAg⁻ chronic HBV infection and HBeAg⁻ groups showed decreased plasma cell proportions in both peripheral blood and liver along with increased hepatic non-switched and switched memory B cells. Functional analysis indicates that B cell subsets in the HBeAg⁺ group may present more severe metabolic dysfunctions, epigenetic abnormalities, as well as signaling and differentiation defects compared to the HBeAg⁻ group, suggesting a potential pivotal role of B cell subsets in HBeAg seroconversion. Moreover, relative to the CR group, B cell subsets in the HBeAg⁻ group are likely to exhibit more profound energy metabolic impairment and aberrant immune activation, which may consequently facilitate the persistence of HBV infection. B-cell subsets may play a pivotal role in HBeAg seroconversion, and their altered proportions and functional defects may further contribute to the persistence of HBV infection.</p>