The gram-negative fecal floras from 953 infants were studied upon discharge of the infants from 22 neonatal wards. More than 600 distinct phenotypes of Escherichia coli, Klebsiella spp., and Enterobacter spp. were distinguished by high-resolution biotyping. The colonization patterns observed showed considerable local and temporal variation. The major (M) strains (phenotypes), which colonized more than 10% and up to 78% of the infants in a ward (median, 23%), were Klebsiella oxytoca (15 strains), E. coli (4 strains), Klebsiella pneumoniae (1 strain), and Enterobacter cloacae (1 strain). Resistance to beta-lactam antibiotics was more pronounced among M strains than among strains of enteric bacteria colonizing few or single infants only. Local antibiotic policy influenced the colonization patterns. Despite the fact that M strains of Klebsiella spp. were usually resistant to ampicillin as well as to cephalexin and cefuroxime, their local dissemination was associated with the use of ampicillin with or without gentamicin but not with the use of cefuroxime. It thus appeared that in the neonatal setting, ampicillin posed a greater risk of local spread of certain drug-resistant bacterial clones than a newer cephalosporin, such as cefuroxime.