Antibiotic use in animals has been identified as a major contributing factor for antibiotic resistance. Given the limited access to veterinary services, the choice of antibiotic and the dosage are determined by the farmers. By analyzing prescription practices and their appropriateness, we can compare performance in antibiotic stewardship across regions to drive change toward national goals and to guide policy interventions. This study used an integrated approach to assess antibiotic use at the grassroots in a rural poultry-keeping community in Wakiso District, Uganda. Medical prescriptions of 347 outpatients were captured between September 2021 and November 2021 via systematic random sampling of patients attending a local health center and were reviewed to decode antibiotic use characteristics and patterns. In the same period, structured questionnaires were issued to 36 poultry farmers in the same area to obtain information on their antibiotic use practices. Of the 347 human outpatient prescriptions reviewed, 250 (72%) recommended at least one antibiotic at a mean of 1.7 antibiotics per prescription. Conversely, all the poultry farms reported antibiotic use. There was significantly more use of macrolides and tetracyclines on poultry farms than at the local health centers and more use of penicillins, nitroimidazoles, and cephalosporins in humans than in poultry. There is unregulated use of critically important antibiotics in poultry and a misuse of antibiotics in the health center due to the lack of adequate diagnostic services. There is a need to improve point-of-care diagnostics of both human and poultry diseases in this community to guide antibiotic use.