Neonates, especially those in a neonatal intensive care unit (NICU), are at high risk for infection. Neonates are susceptible to infection by microorganisms that can be easily transmitted between neonates via caregiver hands or equipment. Sources of infection include contaminated patient care supplies, and infected personnel, family, and other visitors. The risk increases with decreased birth weight and/or gestational age, and with exposure to invasive procedures and indwelling devices. In a nursery for newborns, infections of the skin, mouth, and eye are most frequent; in a neonatal intensive care unit, BSIs predominate. Coagulase-negative Staphylococcus, Staphylococcus aureus, Enterococcus, bacteria in the Enterobacteriaceae family, and Candida are the most frequently isolated organisms. Methicillin-resistant S. aureus (MRSA) is a significant presence in neonatal intensive care units. Outbreaks of respiratory and gastrointestinal viral infections also occur. This chapter presents the essential elements of infection prevention in neonatal environments.