To investigate the contamination by and transmission of MRSA/MRCNS in the old NICU of Hospital A and the relocated NICU (new NICU), we isolated and evaluated staphylococci from nurses' palms, towels under the heads of neonates, infant incubators (including portholes and infant covers), and room air. Detection rates of MRSA/MRCNS isolated from different sample locations were 52.6% in the old NICU and 53.4% in the new NICU, which demonstrates that the nurses' palms in both the old and new NICUs and indoor environment were contaminated with MRSA/MRCNS. In the old NICU, numerous MRSA and MRCNS strains (Log 3.17 ± 0.19 cfu/10 cm²) were identified from towels, and the implementation of improvement plans resulted in a decrease in the number of MRSA/MRCNS isolates (Log 1.95 ± 0.57 cfu/10cm²) from the towels used in the new NICU. A homology study of MRSA/MRCNS strains by PFGE DNA restriction patterns identified genotypes that showed similar patterns in the nurses' palms, towels, infant incubators, and room air.