Tropical theileriosis, which is caused by Theileria annulata, is considered as a serious illness that impairs animal fertility and production in the world's tropical and subtropical regions. This study investigates bovine tropical theileriosis in Blue Nile State, Sudan. A total of 472 blood samples were collected from six localities namely Al Damazine, Geissan, Alrosires, Altadamon, Baw, and Wadalmahi from Kenana and Umbararo cattle breeds of different age groups: calves under two years old, heifers and steers between two and four years old, and cows and bulls above four years old. Out of 472 peripheral blood smears, 20 (4.2%, 95% CI: 2.6-6.5%) were Theileria piroplasm's positive with the highest prevalence in Al Damazine and the lowest prevalence in Altadamon. The prevalence reported by PCR was 8.2% (4/49 blood spots, 95% CI: 2.3-19.6%), where heifers and steers showed a higher prevalence rate compared with other age categories. Microscopic analysis and PCR revealed that Kenana cattle had a high prevalence when compared to Umbararo animals. T. annulata seroprevalence was 42.8% (202/472, 95% CI: 38.3-47.4%) by immunofluorescence antibody test (IFAT). Alrosires had the highest seroprevalence, and Baw had the lowest. Heifers and steers had a significantly high association with the T. annulata seroprevalence compared to other age groups. The odds ratio of T. annulata seropositive in Kenana cattle was 1.6 times more likely than Umbararo cattle. In Blue Nile State, where the management program needs to be strengthened, T. annulata is rapidly expanding and poses a threat to the health of cattle.