Transfusion transmissible infections (TTIs) remain a major health challenge particularly in developing countries. Here, we present a multicentered hospital‐based retrospective study on the prevalence, distribution, and risk factors of TTIs in Ghana. Data on blood donors from four health facilities, namely Nkwanta South Municipal Hospital (Oti region), Weija‐Gbawe Municipal Hospital (Greater Accra region), SDA Hospital (Northern region) and Wa Municipal Hospital (Upper West region) were extracted and analyzed. Descriptive statistics and multinomial logistic regression were applied to compare sociodemographic data with TTI status. A total of 6094 blood donors were included in this study, and 2% were females. The overall prevalence of TTIs was 21.0% (1232/5868). Specifically, the prevalence of HBV, HCV, HIV, and Syphilis was 6.6% (385/5868), 4.9% (286/5830), 2.9% (168/5867), and 6.8% (393/5739), respectively. Wa dominated in all the viral agents considered in this study, while the Oti region recorded the highest prevalence in T. pallidum. The odds of HBV infection was 3.1 (p = 0.008) among first‐time donors, while that for HCV was 2.8 (p = 0.042). For rural dwellers, donors significantly had T. pallidum (p < 0.001; OR = 2.8), HCV (p < 0.001; OR = 2.9), and HIV (p = 0.028; OR = 1.5) infections. Generally, the recipients of transfused blood were predominantly pregnant mothers, followed by children and accident victims. This study has revealed significant disparities and relatively high prevalence of TTIs in Ghana, specifically HBV, HCV, HIV and T. pallidum infections. The variations suggest the presence of unique health challenges per study area, hence the need for a tailored intervention for each study site.