Background: Uganda has suffered from a series of epidemics of Human African Trypanosomiasis (HAT), a tsetse transmitted disease, also known as sleeping sickness. The area affected by acute Trypanosoma brucei rhodesiense HAT (rHAT) has been expanding, driven by importation of infected cattle into regions previously free of the disease. These regions are also affected by African Animal Trypanosomiasis (AAT) demanding a strategy for integrated disease control. Methods: In 2008, the Public Private Partnership, Stamp Out Sleeping Sickness (SOS) administered a single dose of trypanocide to 31 486 head of cattle in 29 parishes in Dokolo and Kaberamaido districts. This study examines the impact of this intervention on the prevalence of rHAT and AAT trypanosomes in cattle from villages that had (HAT +ve ) or had not (HAT -ve ) experienced a recent case of rHAT. Cattle herds from 20 villages were sampled and screened by PCR, pre-intervention and 6-months post-intervention, for the presence or absence of: Trypanosoma brucei s.l.; human infective T. b. rhodesiense ; Trypanosoma vivax ; and Trypanosoma congolense savannah. Results: Post-intervention, there was a significant decrease in the prevalence of T. brucei s.l . and the human infective sub-species T. b. rhodesiense in village cattle across all 20 villages. The prevalence of T. b. rhodesiense was reduced from 2.4% to 0.74% ( P