ABSTRACT The molecular epidemiology of multidrug-resistant Acinetobacter baumannii was investigated in the medical-surgical intensive care unit (ICU) of a university hospital in Italy during two window periods in which two sequential A. baumannii epidemics occurred. Genotype analysis by pulsed-field gel electrophoresis (PFGE) of A. baumannii isolates from 131 patients identified nine distinct PFGE patterns. Of these, PFGE clones B and I predominated and occurred sequentially during the two epidemics. A. baumannii epidemic clones showed a multidrug-resistant antibiotype, being clone B resistant to all antimicrobials tested except the carbapenems and clone I resistant to all antimicrobials except ampicillin-sulbactam and gentamicin. Type 1 integrons of 2.5 and 2.2 kb were amplified from the chromosomal DNA of epidemic PFGE clones B and I, respectively, but not from the chromosomal DNA of the nonepidemic clones. Nucleotide analysis of clone B integron identified four gene cassettes: aacC1 , which confers resistance to gentamicin; two open reading frames (ORFs) coding for unknown products; and aadA1a , which confers resistance to spectinomycin and streptomycin. The integron of clone I contained three gene cassettes: aacA4 , which confers resistance to amikacin, netilmicin, and tobramycin; an unknown ORF; and bla _OXA-20 , which codes for a class D β-lactamase that confers resistance to amoxicillin, ticarcillin, oxacillin, and cloxacillin. Also, the bla _IMP allele was amplified from chromosomal DNA of A. baumannii strains of PFGE type I. Class 1 integrons carrying antimicrobial resistance genes and bla _IMP allele in A. baumannii epidemic strains correlated with the high use rates of broad-spectrum cephalosporins, carbapenems, and aminoglycosides in the ICU during the study period.