<b><i>Background:</i></b> Methicillin-susceptible <i>Staphylococcus aureus</i> (MSSA) is a major contributor to infectious episodes of very low birth weight infants (VLBWI), resulting in significant morbidity and mortality. <b><i>Objective:</i></b> To examine the efficacy and safety of surveillance cultures and the decolonization of MSSA-colonized VLBWI. <b><i>Methods:</i></b> VLBWI admitted to our neonatal wards in 2011-2016 were retrospectively analyzed. Rates of MSSA-attributable infections were compared before and after the implementation of active surveillance cultures and the decolonization of MSSA-colonized patients. The mupirocin susceptibility of isolated MSSA strains was routinely tested. <b><i>Results:</i></b> A total of 1,056 VLBWI were included in the study, 552 in the pre-intervention period and 504 in the post-intervention period. The implementation of surveillance cultures and decolonization of colonized patients resulted in a 50% reduction of incidence rates per 1,000 patient-days of MSSA-attributable infections (1.63 [95% CI 1.12-2.31] vs. 0.83 [95% CI 0.47-1.35], <i>p</i> = 0.024). No adverse effects were observed from application of the decolonization protocol with mupirocin and octenidin. No mupirocin-resistant MSSA strains were detected during the study period. <b><i>Conclusion:</i></b> Implementation of an active surveillance and decolonization protocol resulted in a reduction of MSSA-attributable infections in VLBWI.