A total of 434 Escherichia coli isolated from septicemic calves between 1958 and 1965 and 430 E. coli isolated from diarrheic calves between 1967 and 1970 were studied by colony hybridisation and PCR assays for the presence of the cnf1- and the cnf2-like genes. They were also studied for the presence of genes coding for putative virulence factors associated with the CNF toxins including F17-, Pap- and Sfa-fimbrial adhesins and the recently described CDT-III toxin and AfaVIII-afimbrial adhesin. Thirty (7%) of the 434 septicemic strains were positive for CNF by colony hybridisation. Twenty-six were confirmed as necrotoxigenic E. coli type 2 (NTEC2) and four as NTEC1 by PCR. Thirty-five (8%) of the 430 diarrheic strains were positive for CNF by colony hybridisation. Five of them were studied by PCR and confirmed as NTEC1. The 26 septicemic NTEC2 strains and 20 of the 35 diarrheic NTEC including three of the five NTEC1 were positive for CDT-III. All adhesins studied were present in NTEC as well as in non-NTEC. NTEC1 were mainly Pap-, Sfa- and/or Afa8-positive, whereas NTEC2 were mainly F17- and/or Afa8-positive. This study shows that necrotoxigenic E. coli with their associated adhesins and toxins were present in calves as early as 1958, but their prevalence seems to have increased since that time.