Background: Hepatitis B virus is a public health burden in Uganda, yet little is known about its epidemiology in pregnancy.Objective: This study aimed at determining the prevalence and associated risk factors of hepatitis B virus infection among pregnant women attending antenatal care at the Kyazanga Health Centre IV in Lwengo District, Uganda.Methods: This cross-sectional study was conducted from April 2021 to June 2021 and analysed qualitative data that were collected using a structured in-person questionnaire. Aseptically collected blood specimens were screened for hepatitis B virus infection using an immunochromatographic rapid diagnostic test kit. Participants who were positive for the hepatitis B surface antigen (HBsAg) were further screened for hepatitis B envelope antigen (HBeAg) using commercial rapid diagnostic test kits.Results: Out of 384 pregnant women studied, eight tested positive for HBsAg. This gave a prevalence of 2.1% (95% confidence interval: 1.0% – 4.1%); 5/8 (62.5%) were positive for HBeAg. None of the variables studied were significantly associated with HBsAg positivity among pregnant women.Conclusion: Hepatitis B viral infection is still a public health challenge in pregnant women with possible risk for vertical transmission to their babies in the study area. We recommend routine screening for hepatitis B virus in pregnancy in addition to strengthening current strategies aimed at controlling and preventing hepatitis B infection spread and transmission.