Background:This study aimed at analyzing the microbiological profile of neonatal sepsis in Egypt and to determine the antibiotic susceptibility patterns of the isolated microorganisms. Methods: Two thousand and four hundred blood samples were collected from neonates showing symptoms suggestive of septicemia, inoculated into BACT/ALERT culture bottles. Positive blood culture samples were identified and tested for antimicrobial susceptibility. Results: Among the neonates included in our study, 457 (19%) neonates were positively diagnosed with sepsis. early onset neonatal sepsis (EOS) was detected in 181 (39.6%), while late-onset neonatal sepsis (LOS) in 276 (60.4%) cases. Klebsiella pneumoniae was the most commonly isolated microorganism. Staphylococcus aureus and Enterococcus spp. were the most common isolated Gram-positive bacteria. Candida spp. was more encountered in LOS.An alarming feature of the present study is the high incidence of multidrug resistant microorganisms (65%). Among Gram negative isolates (56%) of were extended spectrum beta-lactamase (ESBL) producers and (70.5%) were carbapenem resistant. In Gram positive bacteria, resistance to methicillin in S. aureus and coagulase negative staphylococci were detected in (50%) and (41%) of isolates respectively. Additionally, 17% of Enterococcus isolates were vancomycin resistant. Coclusion: Our bacteriological profile of neonatal sepsis showed that Gram negative bacteria represented the majority of isolates. Klebsiella pneumoniae was the predominant isolate. In our study, both EOS & LOS share a nosocomial infection profile, as high antimicrobial resistance was observed among our isolates. The susceptibility profiles of the isolates may urge for the change of the current used empirical therapies.