The major secreted virulence factor of Helicobacter pylori, the vacuolating cytotoxin VacA, is known to insert into eukaryotic membranes and has been observed in association with the surface of H. pylori cells that are actively producing it. Here, it is demonstrated that VacA is capable of interacting with the surface of H. pylori and Escherichia coli after secretion. It is shown that this interaction is resistant to disruption of electrostatic and hydrophobic forces, and that it appears to occur despite truncation of LPS and the removal of trypsin-accessible surface proteins. Adsorption to bacterial cell surfaces was independent of the VacA subtype, suggesting that it is not mediated through recognition of a known receptor by the VacA p58 subunit. Similarly, adsorption to bacterial cell surfaces is unlikely to be instigated by the extreme N-terminus of VacA, since a hydrophilic extension at this location that is known to disrupt VacA-induced vacuolation did not interfere with adsorption to H. pylori cells.