This study was planned to investigate the phenotypic and genotypic characterization of Gram negative bacteria isolated from birds of prey (Raptors).A total of 281 fecal swabs was collected from Raptors in Giza zoo. All isolates were subjected to bacteriological and biochemical examination, and some of them to serological and PCR analysis. The results of bacterial isolation revealed that E. coli was isolated with percentage of 51.1%, SalmonellaTyphimurium (1.8%), Proteus spp.(30.9%), Shigella spp.(5.6%), Enterobacter spp.(4.6%), Citrobacter spp.(3.2%), and Pseudomonas spp. (2.8%). The isolated E. coli strains,  were found belong to O serotypes in order of frequency O26, O55, O63, O27, O151, untypable,O28a, O148 and O112 with percentage of  20.83%,  20.83%, 16.67% 12.5%, 12.5%, 12.5%, 8.33%, 8.33%, and 4.17%  isolates ,respectively. All isolated salmonella strains were found belong to Salmonella Typhimurium serotype. PCR analysis was carried out for all identified serotypes showed that (10/11) 90.9% of tested E. coli strains carried (eaeA) virulence gene, (2/11) 18.18% of the tested E. coli isolates were positive to tsh gene, (3/11) 27.27% of the examined E. coli strains bearing iss gene while, stx1 gene was not found in any examined E. coli strain (0/11) 0%, and stx2 was found in (4/11)36.36% of the examined E. coli strains.