Twenty-nine strains of Escherichia coli that adhere to HEp-2 cells with a 'stacked brick' pattern (EAggEC), and four nonadherent control strains, were examined for the ability to hybridize with gene probes for aggregative (AA) and diffuse (DA) HEp-2 cell adhesion phenotypes. These strains were also tested for the ability to express an 18 kDa membrane-associated outer-membrane protein (MAP), to agglutinate erythrocytes, and to produce a pellicle during broth culture. Thirteen of the 29 HEp-2 adherent strains of E. coli hybridized with the gene probes for both AA and DA, and expressed an 18 kDa outer membrane protein (OMP) which was antigenically related to the MAP expressed by strains of E. coli O126:H27. The strains that did not carry the additional DA genes did not express an 18 kDa OMP. Although strains of EAggEC share the ability to adhere to HEp-2 cells with a stacked brick pattern, these strains exhibit a diverse range of physical and biochemical properties. From the results of this study, it was concluded that currently, the possession of EAggEC genes or the ability to adhere to HEp-2 cells in a stacked brick formation, remain the only reliable means of identifying EAggEC.