Bacteriological examination of 300 samples from chicken and O. niloticus fish (150 from each) revealed isolation of 93 isolates of Staph. aureus, 30 isolates from chicken and 63 isolates from O. niloticus fish with an incidence of ( 20% and 42% ) respectively. All the isolates were subjected to different tests to detect some phenotypic characteristics related to virulence: coagulase, DNase, TNase, slime and hemolysin production. In the present study all examined isolates were coagulase positive and the production of DNase, TNase, slime and beta hemolysin was shown by (90 and 96.83), (83.33% -90.48%), (73.33%- 65.8%) and (80% - 92.06%) of chicken and fish isolates respectively. Coagulase gene was detected by PCR and the amplified products of the examined Staph. aureus strains isolated from chicken were analyzed at the band (570 bp), while in case of fish strains the amplified products were analyzed at the band (570 bp and 630 bp ). All Staph. aureus isolated from chicken were (100%) sensitive to vancomycin followed by gentamicin (80%) , (36.67% and 40%) were sensitive to doxycycline and rifampicin respectively, while (36.67%) of the isolates were intermediately sensitive to doxycycline and rifampicin. Staph. aureus isolated from chicken were resistant to penicillin, spectinomycin, cefoxitin and ciprofloxacin at rates of (83.33%, 73.33%, 53.33% and 50%) respectively. Staph. aureus isolated from fresh water O. niloticus samples were (100%) sensitive to vancomycin and gentamycin. The majority of the isolates were (73% and 57.1%) sensitive to doxycycline and ciprofloxacin respectively. While (100%, 76.2% and 58.7%) of Staph. aureus isolates resistant to penicillin, spectinomycin and cefoxitin respectively.