In Saudi Arabia, blood donation is performed in hospitals and blood collected from replacement and volunteer donors. Blood transfusion carries risks, but also has life-saving benefits. In the past several years there was an increasing in the concern of safety of blood transfusion, which helps for the remarkable improvements in the safety of the blood supply. The aim of this study was to evaluate the prevalence of many transfusion-transmitted diseases (TTDs), including hepatitis B (HBV), hepatitis C virus (HCV), human immunodeficiency virus (HIV), venereal disease research laboratory (VDRL) and human T-lymphotropic virus (HDLV) infections among voluntary and replacement Saudi and non-Saudi, male and female blood donors in the Makkah region of Saudi Arabia. The result of this study showed that, 8820 of the blood donors whose records were examined, 1.7% had transfusion-transmitted diseases (TTDs). The majority of positively tested blood units were obtained from male donors. In addition, the overall prevalence of HBsAg was the highest 47% compared to the other infection viruses; HCV (30.2%), HIV (3.4%), VDRL (16.1%) and HDLV (2.7%). This study also revealed that the prevalence of HBsAg positivity was significantly (p < 0.05) higher among Saudi donors compared to non-Saudi. While, the VDRL infection was higher among non-Saudi donors.