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Background and aims: In compared to other domestic field animals, rare studies have conducted in buffaloes to detect the prevalence rate of different diseases. Anaplasmosis, caused by Anaplasma marginale, is one of the most prevalent and important haemoparasitic diseases which result in severe economic losses in particular in milk and weight. Hence, aim of the current study is investigating the prevalence rate of subclinical persistent form of anaplasmosis in buffaloes of Wasit province using of serological method (ELISA) with estimating the association of positivity to some important risk factors. Materials and methods: Totally, 184 buffaloes of different ages and sexes were selected randomly from different regions in Wasit province (Iraq) during April to August (2021). All study animals were and subjected to sampling of venous blood that used to collect of sera. Results: Examination of sera by ELISA revealed that 38.59% were positive to anaplasmosis. According to severity of infection, there were 60.56%, 22.54% and 16.9% mild, moderate and severe infections, respectively. Furthermore analysis of positive results in association with risk factors revealed a significant variation in their values. Significantly, the higher prevalence rate of anaplasmosis was seen in buffaloes aged >4-8 years while the lowest in those aged <1 year when compared to other age groups. Regarding study regions, positive infections were increased significantly in Al-Kut when compared to other study regions; Al-Aziziyah, Al-Hay and Shaykh Sa’d. However, no significant differences were observed between females and males, but males appear to be at a higher risk of infection than females. Conclusion: This study represents the first one in Wasit province which revealed the relative high prevalence rate of anaplasmosis in buffaloes. ELISA can apply as a test of choice for detection of carrier infections. However, furthermore studies are needed to discover the distribution of infection in different provinces of Iraq. A study of vaccine preparation and vector control programs should be applied periodically to prevent moreover infections and to eliminate ticks and flies. Sanitary precaution is necessary during vaccination, docking, castration and surgical intervention to prevent the iatrogenic transmission of anaplasmosis.