The emergence of metallo-β-lactamase (MBL)-producing Enterobacteriaceae is a serious public health concern. Producers have been repeatedly isolated from patients and long-term care facility (LTCF) residents around Bolzano, and we sought to assess their prevalence and clinical impact. All routine Enterobacteriaceae isolates from a Bolzano tertiary-care hospital and associated long-term care facilities in 2008 (n=5500) were screened for MBLs, with case details reviewed for the source patients. In total, 36 producers were obtained from 29 patients, comprising 14 Escherichia coli, six Klebsiella pneumoniae, four Klebsiella oxytoca, four Citrobacter freundii, two Enterobacter cloacae and two Morganella morganii, as well as single Citrobacter amalonaticus, Enterobacter aerogenes, Providencia stuartii and Proteus mirabilis isolates. All were PCR-positive for blaVIM and 25 were PCR-positive for qnrS; 19 non-K. pneumoniae had blaSHV and one had blaCTX-M-group1; 13 were from 12 LTCF residents and 23 were from 17 acute-care patients. All these patients had serious underlying diseases with prolonged hospitalization or LTCF stay; only seven had infections due to the MBL producers, comprising four urinary tract infections, two catheter-related bloodstream infections and one patient with both a surgical site infection and pneumonia. Five patients had more than one MBL-producing organism. Pulsed-field gel electrophoresis identified a cluster of six related E. coli, whereas pairs of K. pneumoniae and C. freundii isolates had >85% profile similarity. Transformants prepared from two isolates were shown to be PCR-positive for blaVIM, qnrS and blaSHV; their plasmids gave similar restriction fragment length polymorphism patterns, and blaVIM-1, qnrS1 and blaSHV-12 were detected by sequencing.