Background: Methicillin-resistant Staphylococcus aureus (MRSA) colonization in Atopic Dermatitis (AD) patients can contribute to worsening their clinical condition. Objective: A cohort study was carried out to determine the incidence of MRSA acquisition and its risk factors in AD children. Methods: Patients with AD (2 months–14 years old) were followed up for about 1 year at a reference center for AD treatment in Rio de Janeiro, Brazil, from September 2011 to February 2014. Nasal swabs from patients and contacts were collected every 2 months. The SCORAD system assessed the severity of the AD. S. aureus isolates were evaluated to determine the methicillin resistance and the clonal lineages. Results: Among 117 AD patients, 97 (82.9%) were already colonized with S. aureus and 26 (22.2%) had MRSA at the first evaluation. The incidence of MRSA acquisition in the cohort study was 27.47% (n = 25). The SCORAD assessments were: mild (46.15%), moderate (37.36%) or severe (16.48%). Risk factors were: colonized MRSA contacts (HR = 2.27; 95% CI: 1.16–7.54), use of cyclosporine (HR = 5.84; 95% CI: 1.70–19.98), moderate or severe AD (HR = 3.26; 95% CI: 1.13–9.37). Protective factors were: availability of running water (HR = 0.21; 95% CI: 0.049–0.96) and use of antihistamines (HR = 0.21; 95% IC: 0.64–0.75). MRSA isolates carried the SCCmec type IV and most of them were typed as USA800/ST5. Conclusions: The high incidence of MRSA acquisition found among AD patients and the risk factors associated show that an effective surveillance of MRSA colonization in these patients is needed.