Maedi-visna virus (MVV) causes a significant economic loss through morbidity, mortality and carcass weight loss in sheep worldwide. This cross-sectional study was conducted to determine the prevalence and identify associations with potential risk factors of MVV virus infection in the selected area of the eastern Amhara region, Ethiopia. A total of 494 sheep blood sera were collected during the period from November, 2017 to October, 2018 and examined using Indirect enzyme linked immuno-sorbent assay (I-ELISA) to screen specific antibodies against MVV. From the total tested sample 3.24% (16/494) were positive for the presence of antibodies against MVV in the area. The sero-prevalence of MVV was not significantly different between associated risk factors of breed, sex, age, production system, flock size and body condition score (P> 0.05). Awassi cross sheep distributor farm and ranch were incriminated as a source for Maedi-visna virus infection and effective control measures should be implemented through appropriate way of testing and culling mechanisms of all seroreactor ewes and their progeny. High sensitive screening test should be practiced and implemented during introduction of new flocks from abroad and before distribution of Awassi cross breed rams from ranches and multiplication center to smallholder farms and individual farmers. In addition further epidemiological study (research) should be done in sheep producing areas of the country to know the level of infection at national and country level.