Antiviral treatment could block mother-to-child transmission (MTCT) of hepatitis B virus (HBV) effectively. We examined whether maternal use of telbivudine (LdT) could decrease the proportion of CD4⁺CD25⁺ regulatory T cells and explored the immunological mechanism. A total of 89 pregnant women with HBsAg positive were enrolled, where 30 pregnant women with HBeAg negative (viral load<10⁶ IU/ml) and the other 59 pregnant women with HBeAg positive (viral load≥10⁶ IU/ml) were followed in the study. The women with high viral load were divided to the LdT-treated group where they were prescribed with 600mg LdT daily (29 cases) during the third trimester of pregnancy or to the non-treated group (30 cases) on a voluntary basis. Samples of neonates were taken for analyzing CD4⁺CD25⁺ Tregs with flow cytometric techniques. A more significant decrease in the proportion of CD4⁺CD25⁺Tregs in neonatal peripheral blood had been observed with maternal use of telbivudine (2.8%±1.1%) than those without any treatment (7.0%±1.6%, P< 0.01). None of the infants in the LdT-treated group were HBsAg positive at 7 months of age. In addition, neonates whose mothers received telbivudine had a significant improvement in cellular immune function, as indicated by the proportion of CD8⁺ T cells. For HBV carriers with high viral load, maternal use of LdT may be useful in regulating neonatal immune function involved in mother-to-child transmission of hepatitis B virus.