This study aims to examine the prevalence and antimicrobial activity of E.faecalis isolated from 100 O. niloticus that showed opaque eyes, bleeding, dark pigmentation in the whole body, distended abdomen, bleeding, and skin ulceration. Gills, skin, kidney, and spleen from each fish were collected for bacteriological findings. The samples were collected randomly from May to October 2022 from private fish farms located in the Ismailia government. The obtained pinpoint white colonies were confirmed biochemically and tested against different antimicrobial agents using disc diffusion assays. Isolates were examined for 16SrRNA sequence by PCR. The prevalence of E.faecalis in all examined organs was 69/100 (69%). The highest prevalence rate of infection was found in the gills among the other collected organs, followed by the liver, kidney, and spleen with a percentage of 28.9%, 26.7%, 24.1%, and 20.3%, respectively. The obtained isolates showed resistance to ampicillin, tetracycline, and erythromycin. It was sensitive to linezolid, imipenem, and levofloxacin