Background: Infection is a frequent and important cause of morbidity and mortality in the neonatal period.Objective: This work was carried out to investigate the prevalence of bacterial infection and the frequency of different pathogens among newborns admitted to the Neonatal Intensive Care Unit (NICU) at Tripoli Medical Center (TMC), Libya.Methods: The case records of all neonates admitted to the NICU of TMC, Libya for the period Sept. 1996 through August 1997, inclusive, were reviewed. Blood and/or CSF cultures were used to establish the diagnosis of bacterial infection. The admissions were categorized as sterile and unsterile. Results: A total of 1123 newborns were admitted to NICU over the period of the study, 129 (11.5%) of them were proved to be bacterially infected, 10.6% and 24% of the sterile and unsterile admissions, respectively, had bacterial infection. Blood culture was positive in 115 (10.2%) of the admitted newborns, while CSF culture was positive in 24 (2.1%) of them. Gram-negative bacteria were the predominantly isolated bacteria. Serratia spp. was isolated from 38.3% and 50% of blood and CSF cultures, respectively. Klebsilla pneumoniae was isolated from about 25% of both blood and CSF cultures. Coagulase negative staphylococcus (CONS) was isolated from 11.3% of blood cultures.Conclusion: It can be concluded from this study that neonatal infection is still a problem facing the country and there is a need for study of bacterial colonization of anogenital tract of Libyan pregnant women and its relation to neonatal infections . Key words: neonatal infection, gram-negative bacteria, Libya