Aims: Millions of lives are saved each year through blood transfusion but a safe blood supply is a critical component of health care to prevent the spread of blood-borne infectious diseases. Therefore, it is essential to assess the prevalence and identify the most common culprit and risk in transfusion services. Methods A cross-sectional study was conducted from April 2015 to May 2015 at the Hawassa blood bank center. Blood donors who donated blood for transfusion purposes were considered for the study. Whole blood was collected and serum was separated from each donor. The sera were used for examination of human immunodeficiency virus (HIV), hepatitis B virus (HBV), hepatitis C virus (HCV), and syphilis. Thick film was prepared and stained using Giemsa for malaria detection. Results: A total of 384 blood donors were screened during the study period. Among these donors, 67.2% (258) were males and 32.8% (126) were females. The overall prevalence of transfusion transmitted infections (TTI) was 28 out of 384 (7.29%) apparently healthy donors. and malaria was 6 (1.6%), 16 (4.2%), 2 (0.5%), 3 (0.8%) and 1 (0.3%) respectively. Two out of 384 (0.5%) had co-infections with HIV-HBV 1 (0.26%) and HBV-HCV 1 (0.26%). A 1:14 ratio (7.14%) of the blood collected was discarded only due to the presence of TTI. The highest discard rate was recorded from HBV infected units 57.1%, followed by HIV 21.40%, syphilis 10.7%, HCV 7.10%, and malaria 3.60%. Overall, TTIs were found in males (7.8%), married (2.5%), rural (8%), private/NGO employed donors (28.6%), 45-54 age group (20%), and in replacement/family donors (16.1%) (p = 0.039). Conclusion: The prevalence of TTIs in the Hawassa blood bank center was significantly high. HBV infection was the major cause for TTIs. Recommendations: Overall, effective clinical and laboratory screening of blood donors is essential. Proper treatment of TTI positive cases and a good donor selection program will help to reduce the prevalence of TTIs in donor units. Blood bank programs should give much more consideration in screening and diagnosis of HBV which is the main TTI that causes a high discard rate of blood.