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Background and Objectives: South Africa has a high prevalence of human immunodeficiency virus (HIV) and hepatitis B virus (HBV) and to a lesser extent human T-lymphotropic virus (HTLV). Each of these agents is transfusion-transmissible (TT) but deciding whether to implement preventive screening depends upon knowledge of background prevalence in transfused patients. We determined the prevalence of HIV, HBV and HTLV I/II among blood transfusion recipients in South African hospitals., Materials and Methods: We obtained identity-unlinked samples used for blood cross-matching at 634 South African hospitals served by the South African National Blood Service (SANBS). The ABBOTT Alinity S® Immunochemiluminescent system measured HIV, HBV and HTLV I/II antibodies. Repeatedly reactive samples were confirmed using the Roche Cobas® 8000. Logistic regression was performed to investigate the determinants of associations for HIV, HBV and HTLV infections., Results: The overall prevalences of HIV, HBV and HTLV were 37.8%, 7.4% and 0.6%, respectively. The HIV prevalence in blood recipients was twice as high as general population estimates. Public hospital patients had a significantly higher prevalence compared with private hospital patients for HIV and HBV. HIV prevalence was significantly higher in females, and HBV prevalence was significantly higher in males, excluding the unknown gender results., Conclusion: Patients receiving blood transfusions in South Africa have high rates of HIV and HBV infection that should be taken into consideration when determining donor screening strategies for other viral infections. Measurable prevalence of HTLV indicates endemicity of this infection in South Africa., (© 2024 The Author(s). Vox Sanguinis published by John Wiley & Sons Ltd on behalf of International Society of Blood Transfusion.)