Helicobacter pylori is strongly associated with both chronic gastritis and duodenal ulcer. Certain strains of H. pylori produce a vacuolating cytotoxin in vitro. At New Orleans Charity Hospital, concentrated culture supernatants from 119 of 144 (83%) H. pylori strains isolated from 86 patients at high risk of developing gastric cancer, caused vacuolization in HeLa S3 cells. Cytotoxin activity was neutralized by acid (pH 4) and basic (pH 10) solutions and proteases, and was precipitable by (NH4)2SO4, which suggests that the cytotoxin is a protein. In 66 of 86 (76.7%) patients, the H. pylori strains isolated from single or multiple sequential gastric biopsies had a vacuolating cytotoxin. These cytotoxin-positive H. pylori strains were isolated from 69% of patients with diffuse antral gastritis and 89% of patients with chronic atrophic gastritis. The latter lesion is considered a precursor of gastric cancer. The cytotoxicity persisted in sequential biopsies over an interval of several months, indicating persistence of these strains in the gastric mucosa. Fifty-eight percent (7/12) of the sera from cytotoxin-positive H. pylori-infected patients neutralized cytotoxin activity, whereas 20% (1/5) of sera from patients with H. pylori cytotoxin-negative strains and none of five H. pylori-negative patients' sera neutralized cytotoxin activity. The relevance of this cytotoxin in the pathogenesis of H. pylori-induced gastritis requires further study.