Background: Hepatitis B virus infection is a potentially life-threatening infection of the liver from hepatitis B virus (HBV). It is a major global health burden accounting for 2.7% of all deaths globally. Being part of the health care system, the risk of exposure to hepatitis B viral infection among medical and health science students is even higher than other health care workers. In this study area, very little is known about the practice of medical and health Science students towards Hepatitis B virus infection prevention and its associated factors. Objective: The main aim of this study was to assess the practice towards hepatitis B virus infection prevention and its associated factors among undergraduate students at Hawassa University College of Medicine and Health Sciences, Hawassa, Ethiopia, 2021. Methodology: Institution-based cross-sectional study was conducted from May 15 to June 15, 2021, among students who had clinical exposure. The 404 sampled participants were recruited using a systematic random sampling technique. Data was collected using a structured selfadministered questionnaire. Data was entered into Epi.Data version 4.6.0 and was exported to SPSS version 25 for analysis. Association between the dependent and independent variables was computed using the bivariate and multivariate logistic regression model. OR was calculated. Results were interpreted as significant if p-value is < 0.05 at 95% CI. Result: This study revealed that 277(69.9%) of the students were in the age group of 20-24 years and 266(67.2%) were males. Out of 396 participants, about half 199(50.3%) had a good practice towards hepatitis B virus infection prevention. Only 43.4% of the study participants had completely vaccinated against hepatitis B virus. Age (20-24) (AOR = 2.736), 95% CI (1.130-6.625), and good knowledge (AOR = 1.990), 95% CI (1.207-3.282) were factors significantly associated with the practice towards hepatitis B virus infection prevention. Conclusion and Recommendation: The current study showed that about half of the study participants had good practice towards hepatitis B virus infection prevention and more than half weren’t completely vaccinated against HBV. Age and knowledge were factors significantly associated. It’s recommended to give training for students on hepatitis B virus infection prevention. It’s also advisable to Screen and vaccinate students before they start their clinical attachments.