Salmonella is one of genus under the family Enterobacteriaceae and recorded as an important zoonotic pathogen. Salmonella infection ( salmonellosis ) is a serious problem to the chicken farms in different areas because the broiler represented its important host. S. Enteritidis strain affected mainly internal organs of the chicken, resulting in elevation of the morbidity and mortality rates, consequently a high economic loses in the broiler production. The aim of this work was the detection of S. Enteritidis(sefA) gene, isolated from internal organs of healthy slaughtered broiler Chickens. The total samples 129, were determined as follow: (Liver, kidneys, small intestine, gizzard and heart blood) are ( 20, 19, 37, 30, 23) respectively. The samples were collected under microbiological examination. The incidence of Salmonella Species was tabulated as flow: (20%, 10.52%, 37.83%, 6.66% and 8.69%) from liver, kidneys, small intestine, gizzard and heart blood respectively. Only 8 isolates from 24 isolated strains were under go serological tests, recording one serotype of Salmonella Enteritidis. The high prevalence occurred in small intestine and liver of apparently healthy broilers, and the lower prevalence occurred in heart blood, kidneys and gizzard of internal organ of broilers. The data resulted from antimicrobial sensitivity test application; high resistant rate to vancomycin (64.6%); gentamicin (64.6%); tetracycline (92.8%); chloramphenicol, (85.7%); ciprofloxacin (35.7%); levofloxacin, (64.28%); amoxicillin+clavulanic, (100%); streptomycin, (92.85); trimethoprim+sulfamethoxaole (85.7%). while the sensitivity to florfenicol was 100%. Confirmatory PCR technique for detection of (sefA) gene amplification at ( 310 bp) in 8 isolated strain.