A mutant (Ty21a) of S. typhi, which lacks the enzyme uridine 5'-diphosphate-glucose-4-epimerase, was evaluated in volunteers for use as a live attenuated oral typhoid vaccine. Five to eight doses of vaccine (containing 3-1010 viable organisms per dose) were given to 155 men without significant side effects. The rate of excretion of the vaccine strain in stools was low, and the majority of isolations occurred on day 1 after vaccination. Revertants able to ferment galactose were not found in any of 958 stool isolates tested. The mutant, strain Ty21a, grown in brain-heart infusion broth (BHIB) with 0.1% galactose, produces more O side chain than the same vaccine strain cultivated without galactose. Volunteers vaccinated with strain Ty21a grown in galactose and then challenged with 105 virulent S. typhi were significantly protected from disease and also had decreased stool carriage of S. typhi as compared with controls. Strain Ty21a grown without galactose did not provide vaccinees significant protection nor decrease fecal excretion of S. typhi as compared with controls. Strain Ty21a when grown in BHIB with 0.1% galactose, results in a safe, stable, and protective oral vaccine that warrants further study in field trials.