Tuberculosis (TB) is a leading cause of death in patients infected with Human Immunodeficiency Virus (HIV), and HIV infection is the most potent risk factor for the development of active TB disease from a latent TB infection. This study aims to determine the seroprevalence of HIV among newly diagnosed TB patients in Kassala state eastern Sudan. This was a descriptive, hospital-based, cross-sectional study of 251 active and newly diagnosed TB patients, selected by simple random sampling. Blood samples and demographic data were collected from each patient. TB was diagnosed by direct ZN smear and molecular detection by Xpert MTB/RIF. The serum samples were tested for HIV using 4th generation enzyme-linked immunosorbent assay (ELISA). The prevalence of HIV was 13.9% (35/251), the infection rate among pulmonary TB was 17%, whereas that in extrapulmonary TB was 4.8%, the prevalence was (18.2%) in the males, and (7.2%) in the females. In conclusion: TB/HIV co-infection in the Eastern part of Sudan was high compared with the global prevalence, all TB patients should therefore be assessed for HIV risk factors and advised to undergo HIV testing.