Pregnant women and women of childbearing age were enrolled in our study and their knowledge about the Hepatitis B virus (HBV) and chronic hepatitis B (CHB) was evaluated. A questionnaire was distributed to every woman in the cross-sectional study. The questionnaire was answered by all participants before they received health education and advice about HBV and CHB from the doctors visited. Data collected from all answers were analyzed using the χ2 test and logistic regression models. A total of 206 pregnant women and women of childbearing age with CHB infection were enrolled in the study during their first visit to the Infectious Diseases Clinic of the Third Affiliated Hospital of Guangzhou Medical University. Some women of childbearing age (40.8%) and pregnant women with CHB infection (30.6%) still believed HBV could be transmitted through diet and/or mosquito bites. Some women of childbearing age and pregnant women with CHB infection had limited knowledge of the prevention of HBV transmission (111 of 206, 53.9%). Women with higher levels of education had more knowledge about HBV (senior middle school, P = 0.02; university, P <0.01). The majority of participants were willing to take antiviral medicine to decrease the mother-to-child transmission (MTCT) rate of HBV. Some women of childbearing age and/or pregnant women with CHB infection have relatively limited knowledge about HBV or CHB. This situation contributes to the timeliness, or lack thereof, of these women with CHB to see a doctor and receive antiviral therapy. As a result, the morbidity and mortality of HBV-related complications could increase along with the rate of MTCT of HBV.