Objectives: Necrotizing enterocolitis (NEC) has been associated with a wide variety of bacteria and their cytotoxins. The content and the nature of gut bacterial colonization in newborns that require intensive care hospitalization has been demonstrated to be abnormal. In the 25-bed neonatal intensive care unit in Hospital Simon Bolivar, in Bogota, Colombia, cases of NEC are common causes of morbidity and mortality. This article examines the hypothesis that oral administration of prophylactic Lactobacillus acidophilus and Bifidobacterium infantis to all neonates in an intensive care unit, would decrease the incidence of NEC. Methods: Daily doses of 250 million live L. acidophilus and 250 million B. infantis were given to all 1237 newborns (both inpatients and transfer patients) admitted to the unit during 1 year, until they were discharged from the hospital. In this study, 1282 patients hospitalized during the previous year were used as controls. Results: There were no complications attributed to the daily administration of L. acidophilus and B. infantis. The study groups were compared for place of origin, clinical, and demographic variables, and there was no statistically significant difference in those variables. In the historic control group, there were 85 NEC cases compared to 34 cases in the group that received probiotic prophylaxis (P < 0.0002). In the historic control group, there were 35 NEC-associated fatalities compared to 14 fatalities in the group that received probiotic prophylaxis (P < 0.005). Conclusions. The positive results in this study support the need for further investigation of bacterial colonization and its role in NEC.