Antibiotic resistance in 3 common pathogenic types of Enterobacteriaceae was examined in a pointprevalence study in 2004. Strains of Escherichia coli, Klebsiello and Enterobacter species were collected prospectively in 25 institutions in Australian capital cities and tested by broth microdilution to 12 13-lactams and 3 other antibiotics. Almost 22% of isolates tested were from blood cultures. In E. coli, acquired resistance to ampicillin and piperacillin was common (> 40%), and clinically significant percentages of intermediate susceptibility and resistance (> 8%) were observed to amoxycillin-clavulanate, cefazolin and trimethoprim. In Klebsiella species, clinically important acquired resistance (> 8%) was seen to piperacillin, cephalothin and trimethoprim, while in Enterobacter species, this was found with piperacillin, ceftriaxone, ceftazidime and trimethoprim. Blood culture isolates had similar rates of resistance to isolates from other specimen sources. New South Wales/Australian Capital Territory (combined) tended to have higher percentages of resistance than the other states, which were otherwise comparable across the agents and species tested. Multi-resistance, defined as more than 3 acquired resistances to antibiotic classes, was found in 6.5% of E. coli, 8.3% in Klebsiella species and 16.9% of Enterobacter species. Co-resistance to ciprofloxacin, gentamicin and/or trimethoprim was common in isolates presumptively harbouring extended-spectrum beta-lactamases. Strains with extended-spectrum beta-lactamases, although common in other countries, appear to be at fairly low levels in Australia; less than 4% in E. coli and less than 9% in Klebsiella species. Rates in Enterobacter species were not able to be determined. Presumptive plasmid-borne AmpC beta-lactamases were seen at low levels across the country and carbapenemases have now been found for the first time in Australia in Enterobacteriaceae. Both of these types of resistance represent a significant threat to major last-line antibiotics.