Background: Neonatal sepsis (NS) due to K. pneumoniae is a major cause of morbidity and mortality in neonates. This study aimed to study the risk factors of NS caused by K.pneumoniae in NICU in Menoufia University Hospitals and to detect rmpA and magA virulence genes and CTX-M antibiotic resistance gene. Correlation between compliance of infection control measures and occurrence of NS and its outcome were also evaluated. Methods: K. pneumoniae were isolated from blood of neonates with sepsis and studied for hypermucovicosity by string test and detection of rmpA and magA genes. ESβL production was studied by cephalosporin/clavulanate combination disks and expression of CTX-M gene groups. Hand hygiene and other infection control measures compliance were evaluated by observational and practical methods. Results: K. pneumoniae was the most frequently isolated organism (31.6%) among neonates with confirmed sepsis. Hypermucoviscous phenotype was detected by string test in 39.6% of isolates while rmpA and magA genes were found in 47.9% and 8.3% respectively. ESβL production was confirmed in 75% (by cephalosporin/clavulanate combination disk). The CTX-M gene was found among 77.8% of ESβL-producing K. pneumoniae isolates. There was a negative correlation between hand hygiene and other infection control measures compliance and occurrence of NS. Conclusion: Virulence and antimicrobial resistance genes are common among K. pneumoniae isolated from neonates with sepsis in our locality. Implementation of infection control measures and proper antimicrobial stewardship programs may be helpful to overcome this problem.