Diabetic patients, as compared to non-diabetic subjects, run an increased risk of acquiring Gram-negative bacteraemia. We therefore studied the prevalence and coexpression of seven bacterial virulence markers of 69 Escherichia coli strains isolated from 64 bacteraemic patients with diabetes mellitus and 67 E. coli strains from faeces of healthy controls. The strains were analyzed for haemolysin (HLY) production, aerobactin-mediated iron uptake (AMI), cytotoxic necrotizing factor (CNF) production, expression of cell surface hydrophobicity, P-fimbriae, mannose-resistant haemagglutination (MRHA) and mannose-sensitive haemagglutination (MSHA). All bacterial properties were significantly more common among the bacteraemic strains (P < 0.02 vs. controls). Correlations between HLY and CNF (P < 0.0004) and between P-fimbriae and MRHA (P < 0.0001), MSHA (P < 0.0002) or AMI (P < 0.05), as well as between MRHA and MSHA (P < 0.0005) were observed. In patients with proteinuria, as sign of diabetic complications in the urinary tract, HLY-negative strains, P-fimbriae-negative strains, and strains which were both HLY-/CNF-negative, were more common (P = 0.04, P < 0.01 and P = 0.048, respectively). Using a multivariate statistical analysis, production of HLY and the expression of P-fimbriae were the two virulence factors with the highest discrimination between bacteraemic and control strains. In conclusion, all virulence factors studied were more prevalent in bacteraemic than in control strains, although HLY and P-fimbriae were shown to be of greatest and independent importance. Low virulent strains (P-fimbriae-, HLY- and CNF-negative) were more prevalent in diabetic patients with signs of renal complications.