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This pilot study was performed to determine the safety and size of effect of antibiotic cycling to reduce colonization and infection with antibiotic-resistant bacteria.Open, observational study.The study was performed in a 16-bed pediatric medical-surgical intensive care unit.Critically ill children requiring antibiotic therapy.Three antibiotic classes were systematically cycled for 3-month intervals over 18 months. Antibiotic regimens were used for all empirical therapy and continued if the bacterial isolate was susceptible.The primary outcome was colonization with antibiotic-resistant bacteria, determined by surveillance cultures obtained twice monthly from all patients in the unit. Rates of antibiotic-resistant, nosocomial blood stream infections, and risks of colonization over calendar time in the intensive care unit were also evaluated.The cycling of broad-spectrum, empirical antibiotics was safe and did not generate increased antibiotic resistance nor select for new organisms. Over the study period, the trend in prevalence of children colonized with antibiotic-resistant bacteria was from 29% to 24% (p = .41). The effect on prevalence of resistant blood stream infections was similar (p = .29). Changes in individual risks of colonization with resistant bacteria over calendar time were consistent with the ecologic effect in size and direction.Results of this pilot intervention suggest that cycling antibiotics may be a safe and viable strategy to minimize the emergence of antibiotic resistance in intensive care units. A definitive study will require a randomized and controlled trial of only four pediatric intensive care units over an 18-month period.