The primary objective of this study was to ascertain the prevalence, antimicrobial susceptibility, and biofilm production of Staphylococcus aureus strains isolated from sheep and goats affected with mastitis and endometritis in Beni Suef governorate, Egypt. In sheep and goats mastitic milk samples, the prevalence rate of S. aureus was found to be 26% (32 isolates out of 122 samples), while it was 7.75% (9 isolates out of 116 samples) in endometritis samples. The antibiogram for S. aureus isolates from sheep and goats mastitis cases revealed resistance to amoxicillin-clavulanic acid (78.1%), cefotaxime (75%), cephalexin (71.8%), amoxicillin (68.7%), and ceftriaxone (66.7%). On the other hand, the recovered S. aureus isolates from sheep and goats with endometritis revealed resistance to amoxicillin-clavulanic acid (88.9%), cefotaxime (77.8%), and ceftriaxone (66.7%). Phenotypic detection of biofilm on Congo Red agar revealed that 20 Staphylococci isolates (66.67%) formed biofilms, out of them, 16 isolates (53.33%) from mastitis and 4 isolates (13.34%) from endometritis. The results of molecular characterization showed that all the four tested S. aureus isolates were found to possess the mecA gene, while the tetK gene was found in three out of four isolates. In relation to genes associated with virulence, all four isolates were tested positive for coa and three out of four isolates tested positive for icaA. In conclusion, S. aureus stands as a prominent pathogen in cases of mastitis and endometritis in both sheep and goats.