Background and Objectives: Non-viral bloodborne diseases are a group of infections that are a public health problem worldwide. The incidence of diseases such as brucellosis and syphilis is increasing in the Americas and Europe. Chagas disease is an endemic problem in Latin America, the United States and Europe. This study aims to determine the prevalence of non-viral bloodborne diseases in blood donors and to discuss some issues related to federal regulations for the control and prevention of these infectious diseases in Mexico. Material and methods: A cross-sectional study was conducted in the Western National Medical Center Blood Bank, including 228,328 blood donors (2018–2023). Frequencies, percentages, means, standard deviation and confidence intervals (CI) were calculated for demographic data. Prevalences were expressed as rates per 100,000 with 95% CI. Results: Of 3949 seroreactive or undetermined blood donors at the first screening, a total of 682 (0.299%) completed their follow-up test and were positive for Treponema pallidum (478), Trypanosoma cruzi (83), or Brucella spp. (121). The overall prevalence for non-viral bloodborne diseases was 299 per 100,000 blood donors. The prevalence for syphilis, Chagas disease, and Brucella was 209, 36, and 53 per 100,000 respectively. Conclusion: Federal regulations should be reviewed to formulate specific public health policies focused on controlling and preventing nonviral bloodborne diseases.