Animal African Trypanosomiasis also known as nagana is a debilitating vector-borne disease caused by trypanosome parasites and transmitted to human and animals by tsetse fly of the Glossina genus within sub-Saharan Africa. The parasite lives extracellularly and multiplies in the blood and fluid tissues of the mammalian host, leading to morbidity and mortality of the infected animal if untreated. As no known vaccine is currently available to combat the disease, prevention is hinged mainly on vector control and the use of trypanocides. We therefore, carried out a cross-sectional survey to investigate the composition of vectors, parasite diversity and transmission intensity. A total of 229 tsetse flies, 65 pigs and 20 cattle blood samples were collected from two districts in the Eastern region. We performed Cytochrome c (cox1) and internal transcribed space (ITS1) dependent PCR assays to detect and identify tsetse fly species and trypanosome parasites circulating in the study area. More than thrice as many female flies 225 (98%) as male flies 74 (32.3%) were collected using biconical traps. The overall vector density of 4.3 flies/trap/day was observed. Sequence analysis revealed that all collected tsetse flies belonged to Glossina palpalis palpalis and PCR results revealed trypanosome prevalence of 58.9%, 46.2% and 0% in tsetse flies, pigs and cattle respectively. Trypanosoma congolense was the predominant parasite species with prevalence of 80.7% in flies and 60% in pigs. There was evidence of multiple trypanosome infection in both tsetse flies and pigs with T. congolense/T. simiae occurring highest with prevalence of 37.03%. The parasite prevalence in the flies and pigs within the communities was high with significant difference associated (χ2967;2 = 16.42, P= 0.0367; χ2967;2 = 28.06, P=0.0009). Blood meal analysis revealed feeding on both domestic Sus scrofa domesticus (pig) and Phacochoerus africanus (warthog) as the source of tsetse blood meal. Tsetse flies remain highly infected with trypanosomes and may continue to transmit the parasites to livestock and human populations in the communities.