Severe infections represent the main cause of neonatal mortality and morbidity. Strategies of proven effectiveness in reducing the incidence of infection in neonatal intensive care units (NICUs) include hand hygiene practices and prevention of central venous catheter-related bloodstream infections. In recent years, new strategies have been developed to prevent infections in NICU including prevention of neonatal sepsis with lactoferrin, the use of heparin for the prevention of CRBSIs, the judicious use of antibiotics and chemoprophylaxis, prevention of invasive fungal infections with fluconazole, the use of specific anti-staphylococcal immunoglobulins, and the early identification of infants at higher risk of infection with the use of specific markers (mannose-binding lectin). This review will focus on these new strategies and on their role in clinical practice in order to further reduce the incidence of infection in NICU.