Infectious diseases of cultured fish are among the most notable constraints on the expansion of aquaculture and the realization of its full potential. 
E. coli in fish is an indicator of potential pollution levels of it are used to determine whether local beaches should be posted with ”no water contact” advisories and causes severe economic losses among cultured freshwater fish and causes high mortalities.. The objective of this study was 1)Isolation and characterization of E. coli isolates from cultured freshwater fish. 2) Determination of the prevalence rate and virulence character associated with E. coli isolates in cultured freshwater fish (Oreochromis niloticus).3) Characterization of E. coli isolates using pathogenicity test and calculation of LD50 and recording clinical signs, P.M lesions with measuring blood parameters and histopathological finding 4) Evaluation of locally prepared bacterin against isolated strain of E. coli . This study was carried-out on :1) Naturally collected fish for isolation of E. coli : 
Three hundereds and fifty samples of fish species summarized as; Clarias garipinas (95 fish), Oreochromis niloticus (73 fish), Common carp (75 fish), Mugil cephalus (107 fish). These fish were obtained from different farms from different governorates, El-Behera (50), Kafr El-Sheikh (100) and Damiette (200) at different seasons. 2) Fish for experimentally infection :A total number of Four hundereds and ninty apparently healthy O. niloticus, were obtained from private fish farm. The results of this study indicatedthat :1)The results of biochemical tests for detection of of isolated bacteria cleared that, the isolated bacteria gave the same ideal biochemical characters-of E. coli .2) Presence of ammonia and pollution in the water initiate the appearance and activate the E. coli incidence3)The most important fish species susceptible to E. coli infection were Catfish, Common carp, Mugil cephalus and finally the O. niloticus and the intestine is the most organ that isolated E. coli from it , followed by spleen, liver and kidney .4) The results of antibiogram test indicated that, the antibiotic sensitivity test for all isolates of E. coli showed that, the antibiotic sensitivity test for all isolates of E. coli showed that spectinomycin enrofloxacine oxanilic acid were the most susceptible antibiotics for E. coli isolates.