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To monitor trends in HIV seroprevalence in the United States, HIV testing was included in the National Health and Nutrition Examination Survey (NHANES) conducted from 1999 to 2006.From 1999 to 2006, 11,928 participants aged 18-49 years were tested for HIV antibody. Prevalence estimates were weighted to account for oversampling and nonresponse.There were 67 HIV antibody-reactive individuals for a seroprevalence of 0.5% [95% confidence interval (CI) 0.3-0.6]. In the only age subgroup directly comparable between surveys (18-39 years), HIV seroprevalence remained constant from NHANES III (1988-1994) to NHANES 1999-2002 and 2003-2006. In NHANES 1999-2006, non-Hispanic blacks had significantly higher HIV seroprevalence (2.0%, 95% CI 1.5-2.7) compared with individuals in all other race/ethnic groups combined. Seroprevalence was also higher in each race/ethnic group among men who have sex with men (9.4% 95% CI 5.0-17.1), among persons who had detectable antibody to herpes simplex type-two (1.9% 95% CI 1.4-2.8), among those who had 50 or more lifetime sex partners (3.4%, 95% CI 1.7-6.7), and among those who never married (0.8%, 95% CI 0.5-1.3).In this household-based population, seroprevalence did not significantly change from NHANES III to NHANES 1999-2006. Non-Hispanic blacks had significantly higher prevalence of infection compared with other race/ethnic groups. Male-to-male sex and the presence of HSV-2 antibody were the strongest predictors of HIV infection.