Abstract introduction: Neonatal sepsis is a global health problem that leads to high mortality and increased hospital costs. Aim and objectives: This study sought to assess the predominance of neonatal sepsis and its associated risk factors, pathogen distribution, and antibiotic susceptibility among hospitalized neonates. Methods: A prospective study was done in the NICU of Fayoum University Hospital from November 2022 to November 2024. The study included 179 neonates with culture-proven sepsis. Clinical and laboratory data were collected, comprising different cultures and antibiotic sensitivity tests. Results: The culture-proven sepsis incidence was 14.8%, classified as early-onset sepsis (39%) and late-onset sepsis (61%). Early-onset sepsis is more common in newborns with maternal disease. Klebsiella was the most prevalent pathogen and increased considerably in late-onset sepsis, followed by CONS. Gram-positive cocci and gram-negative bacilli demonstrated strong resistance to penicillins and cephalosporins. Conclusion: In Egypt, Antibiotic resistance is an increasing challenge in treating neonatal sepsis. Therefore, constant monitoring of changes in microbial flora, their susceptibility to antibiotics, the prudent use of antibiotics, and the antibiotic cycling approach should be implemented to address this problem.