American Indians are believed to be at high risk of gallbladder disease (GBD), but there has been no systematic evaluation of its prevalence among diverse groups of American Indians. Therefore, we determined the prevalence of GBD and associated risk factors among specified American Indian populations using ultrasonography of the gallbladder and standardized diagnostic criteria. Enrolled members, aged 47 years and older, of 13 American Indian tribes or communities in Arizona, Oklahoma, and South and North Dakota who participated in the Strong Heart Study were analyzed. GBD was the sum of gallstones (determined by ultrasound examination) and cholecystectomy (determined by ultrasound and self-report). The proportion of American Indian heritage was based on the heritage of the grandparents of participants. GBD prevalence was determined among 3,296 participants at the 3 sites. Among women, 17.8% had gallstones, and 46.3% had evidence of a cholecystectomy, for a total of 64.1% with GBD. Among men, 17.4% had gallstones, and 12.1% had evidence of a cholecystectomy, for a total of 29.5% with GBD. When figures were adjusted for age and Indian heritage, there was no significant difference in GBD prevalence across the 3 geographical areas. In multivariate logistic regression analysis, age, American Indian heritage, and waist circumference were associated with GBD among men, and age, American Indian heritage, diabetes, and parity were associated with GBD among women. Body mass index was not independently associated with GBD in either sex. In conclusion, GBD was found in epidemic proportions in diverse American Indian populations.