This study confirms the systemic position of Dermocystidium sp. in Nile tilapia (Oreochromis niloticus) internal organs (liver, spleen, kidneys, stomach and intestine). Our work reveals an entire approach for Dermocystidium sp. including prevalence, diagnosis, spores multiplication, experimental infection and histopathological examination. A total of 1388 Nile tilapia (766 cultured fish and 622 wild fish) were collected from May 2012 to May 2014 and screened for Dermocystidium infection. The total prevalence of infection was 35.5% and was higher in wild (45%) than cultured fish (27.8%). The highest rates of infection were recorded in the largest fish and in winter. Several different techniques were employed to detect and identify Dermocystidium sp. from the internal organs of infected hosts including macroscopic examination, microscopic observation of tissue squashes, histological examination and in vitro culture. Infected fish were sluggish and suffered from severe ulceration and black discoloration of the skin. The post-mortem lesion included dark-grayish nodules on internal organs, particularly the liver. Squash preparations from the internal organs revealed spherical spores (signet-ring shape), which is characteristic for Dermocystidium sp. The spores showed multiplication in the freshly dead host tissue. Our study demonstrated new successful attempts for in vitro culture of Dermocystidium either on Eagle’s Minimum Essential Medium after adjusting pH of media to 3.5 or on Sabouraud's dextrose agar media enriched with duck decoction 10%. Experimental infection revealed that intra-gastric route was more pathogenic than the immersion route. In histopathological sections from infected fish tissue following natural and experimental infection, spherical spores of Dermocystidium were embedded in the affected tissue. To the best of our knowledge, this study represents the first report of Dermocystidium infection in wild Nile tilapia in Egypt