Pseudomonas and micrococcus (Pseudomonas fluorescens and M. luteus) are an emerging opportunistic fish pathogens that results in considerable economic losses among the infected fish. In the present study, a total number of 150 cultured Nile tilapia, Oreochromis niloticus were collected from concrete ponds farms in Alkharga oases, New Valley governorate during the period from March to July 2019 and subject to clinical and bacteriological examination. including excessive black colouration of skin, scales loss, skin ulcer and focal hemorrhages, exophthalmia, fin congestion and rot. Internally, the diseased fish had congested internal organs including liver, spleen, and kidney., The prevalence of Pseudomonas fluorescens and M. luteus infection was 6.7 % withe a total number of 10 isolates and 11.3% withe a total number of 17 isolates  respectively. Were recovered from infected fish. The suspected Pseudomonas fluorescens isolates were biochemically similar while The suspected M. luteus isolates were biochemically similar, except for Arginine hydrolysis, Vogus-Proskauer and Urease tests. The isolation and biochemical characterizations suggested that the isolates belong to the genus Pseudomonas. And genus micrococcus. The recovered M. luteus isolates were pathogenic to O. niloticus, this is the first report of M. luteus infection in O. niloticus in Egypt.