We conducted this study to investigate the isolation frequency and phenotypic antibiotic resistance pattern of Staphylococcus aureus isolated from rodents, chickens, humans, and household soils. Specimens were plated onto mannitol salt agar (Oxoid, Basingstoke, UK) and incubated aerobically at 37 °C for 24 h. Presumptive colonies of S. aureus were subjected to Gram staining, as well as catalase, deoxyribonuclease (DNAse), and coagulase tests for identification. Antibiotic susceptibility testing was performed by using the Kirby–Bauer disc diffusion method on Mueller–Hinton agar (Oxoid, Basingstoke, UK). The antibiotics tested were tetracycline (30 μg), erythromycin (15 μg), gentamicin (10 μg), ciprofloxacin (5 μg), clindamycin (2 μg), and amoxicillin-clavulanate (20 μg/10 μg). The S. aureus strain American Type Culture Collection (ATCC) 25,923 was used as the standard organism. We found that 483 out of 956 (50.2%) samples were positive for S. aureus. The isolation frequencies varied significantly between samples sources, being 52.1%, 66.5%, 74.3%, and 24.5%, respectively, in chickens, humans, rodents, and soil samples (p < 0.001). S. aureus isolates had high resistance against clindamycin (51.0%), erythromycin (50.9%), and tetracycline (62.5%). The overall prevalence of multidrug-resistant (MDR) S. aureus isolates was 30.2%, with 8.7% resistant to at least four different classes of antibiotics.