Background: Neonatal Sepsis is a significant cause of neonatal mortality and morbidity due to the attendant naivety of neonate’s immune systems, delays in diagnosis and inappropriate therapy. Objective: To determine the prevalence of Neonatal Sepsis, characterize and identify causative organisms and identify possible risk factors. Setting: The Lagos University Teaching Hospital (LUTH) is a 761 bed tertiary healthcare center with four neonatal wards. Methods: A total of 250 in-patients in the neonatal wards that had signs and symptoms suggestive of sepsis were consecutively recruited into the study. Blood was collected aseptically and dispensed into BACTEC blood culture bottles and incubated for up to 5 days. Isolates were identified and susceptibility testing performed on them using the modified Kirby-Bauer method. Resistance testing was done for Extended Spectrum Beta Lactamase (ESBL) and Methicillin Resistance Staphylococcus Aureus (MRSA) expression using Clinical Laboratory Standards Institute (CLSI) guidelines. Risk factors and outcome were ascertained using data obtained from a structured questionnaire.