Equine infectious anaemia (EIA) is a life-long viral infection affecting equids, transmitted mechanically by biting flies and iatrogenic means. Despite its global distribution, active surveillance is limited, with passive clinical surveillance or control of specific equine sectors prevailing. In Italy, a national surveillance plan in horse, donkey, and mule populations has been established and includes mandatory passive and active surveillance through annual serological tests. During 2007–2010, the agar gel immunodiffusion (AGID) test served as both screening and confirmatory tests. Since 2011, a three-tier diagnostic pathway was introduced, utilizing the ELISA test for screening, AGID as the confirmatory test, and the immunoblot test for cases where ELISA was positive and AGID was negative. From a total equid population of 406,000 animals, 1,337,899 samples were analysed during 2007–2012, with 2,348 (0.18%) testing positive. EIA seroprevalence significantly decreased across all the species/hybrids during the study period. EIA occurrence was higher in mules (IRR = 48.90) and lower in donkeys (IRR = 0.56) compared to horses. The holding seroprevalence was 1.15%. Spatial analysis revealed clusters of infection in central Italy. These findings demonstrate that active systematic surveillance effectively reduces EIA prevalence in equid populations. Mules and working horses in wooded areas appeared to be at higher risk of infection and act as EIA reservoirs. Surveillance and control should be maintained and strengthened in these species/hybrids and in these areas to effectively control EIA. Passive surveillance alone is insufficient to eradicate the disease, and EIA remains a constant threat for the equine industry if active control is not implemented.