ObjectiveTo investigate colonization of multidrugresistant organisms (MDROs) in neonatal intensive care unit (NICU) newborns on admission. MethodsFrom April to November 2013, 293 newborns who admitted to  NICU of a hospital were screened for methicillinresistant Staphylococcus aureus (MRSA) by nasal and throat swabs and for extendedspectrum βlactamases  (ESBLs) bacteria and vancomycinresistant Enterococcus (VRE) by anal swabs.ResultsOf 293 newborns, 61 were detected MDROs (20.82%).The positive rate of MDROs screening  in newborns aged ＜3 days(5.92%) was lower than those aged ＜3-6 days（37.74%） and  7-28 days (43.66%）,the difference was significant (P=0.000). The major colonized MDROs were ESBLsproducing bacteria（83.60%）, the major colonized site was anus(88.52%). ConclusionNeonatal anus and stool are important sources of MDROs in  NICU; more attention should be paid to colonization screening for MDROs by anal swabs in newborns aged ＞3 days, and appropriate isolation measures should be taken for positive screening patients to prevent the transmission of MDROs.