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Background: Animal African trypanosomosis (AAT) is a disease caused by several protozoan parasites transmitted by tsetse fly (genus Glossina). AAT causes massive losses to farmers due to the high cost of control measures and treatment of sick animals. This study aimed to assess the knowledge, attitudes and practices of the local community on tsetse flies and trypanosomosis and evaluate demographic factors associated with adequate knowledge of tsetse flies along Kenya coast. Methods: We conducted a cross-sectional study between November and December 2017 among 404 cattle farmers living near the Arabuko-Sokoke Forest Reserve in Kilifi County. Structured questionnaires were used to collect data from randomly selected cattle rearing households. Descriptive statistics were used to analyze the local community's knowledge, attitudes, and practices regarding tsetse flies and trypanosomosis, and demographic factors associated with knowledge of tsetse flies were investigated using a logistic regression model.Results: Of 404 study participants, 214 (53%) were female, and 123 (30%) were elderly (>55 years old). Majority (N=312, 77%) were married, and 326 (81%) had only primary education or none at all. Although majority of farmers (N=353, 89%) knew the local name for tsetse fly and that they are about the size of a housefly (N=307, 77%), only 65 (16%) knew they transmit diseases to livestock, with 55/65 (85%) identifying they transmit Nagana disease. Overall, only 16% of the study participants had adequate knowledge on tsetse flies and trypanosomosis. Being 15 to 24 years old (aOR 2.86 (95% CI 1.10–7.45)) compared to those aged >55 years, secondary education (aOR 2.45 (95% CI 1.42–4.21)) and tertiary education level (aOR 3.78 (95% CI 1.53–9.31)) compared to no education were significantly associated with higher odds of tsetse fly adequate knowledge. Unemployment (aOR 0.15 (95% CI 0.10–0.23)) was significantly associated with lower odds of tsetse fly adequate knowledge when compared to self-employment.Conclusion: Our findings suggest that farmers have poor understanding of trypanosomosis and its biological transmission by tsetse flies. Thus, training of livestock farmers will contribute to improved livestock health through better understanding of key livestock diseases, their vectors in transmission, and control.