During a three-year survey of the prevalence in central Tunisia of Escherichia coli producing CNF1 toxin (NTEC), 716 samples have been investigated by PCR for cnf1 gene. All samples were isolated from urine of adult and children patients presenting significant bacteriuria (> 10(5) colony-forming units/mL), independently of the severity of the clinical presentation; 328 strains were found harboring cnf1 gene, they were distributed into three clinical categories: 219 (66.76%) from patients with symptomatic bacteriuria, 76 (23.17%) from patients with uncomplicated cystitis and 33 (10.06%) from patients with acute pyelonephritis and complicated urinary tract infections. 98.78% (324) of CNF1 strains presented hemolytic activity. All 328 CNF1 strains harbored both sfa and pap genes and expressed MRHA activity. They belonged to 16 different serotypes. The most common serotypes, in order of frequency, were O6 (25.91%), O4 (17.98%), O2 (12.5%), O75 (9.14%), O78 (8.35%), and O83 (3.65%). Two strains (0.6%) were O168; a serotype shown to be associated to CNF2 producing bovine strains. The frequency of uropathogenic CNF1 strains in center Tunisia was about 45.81% and increased from 26.08% in 1998 to 58.16% in 2000. We showed that E. coli producing cytotoxic necrotizing factor (CNF1) was implicated in urinary tract infections in center Tunisia but no difference was shown between strains isolated from patients with complicated or uncomplicated urinary tract infections. The presence of CNF1 toxin with various associated virulence factors seemed to increase the risk for severe forms of urinary tract infections.