Background: Although ethnic differences in disease incidence among African American (AA) men may reflect differences in screening behaviors and behavioral and socioeconomic factors, genetic ancestry may be a factor. This study will investigate the association between West African and Native American genetic ancestry and PCa diagnosis. Methods: We enrolled 40-79 year-old men who were undergoing prostate biopsy or routine PSA screening at outpatient urology clinics in Chicago, IL. Blood was drawn at the time of enrollment for the genotyping of 105 ancestry informative markers. West African (WAA), European (EA), and Native American (NAA) genetic ancestry was estimated using the Bayesian Markov Chain-Monte Carlo method implemented in the program STRUCTURE. Univariate analysis was performed by Wilcoxon rank-sum test for continuous variables. Unconditional binary logistic regression models were used to estimate odds ratios for the likelihood of PCa diagnosis and for the odds of having high-risk PCa relative to controls, adjusting for age, body mass index (BMI), alcohol-use, and family history of PCa. Results: 857 men were enrolled; 34.5 % were diagnosed with PCa, 50.9 % were controls and 14.6 % were men with negative prostate biopsies. AAs comprised 27% of the sample. The median was 63 years (IQR 57 – 68). Among AA men, the median genetic WAA proportion was 0.78 (IQR 0.69 – 0.83) and NAA was 0.04 (IQR 0.02 – 0.09). There was no significant difference in genetic WAA among AA men with and without PCa diagnosis (p = 0.54); however, genetic NAA was significantly lower among PCa cases (median 0.034 vs 0.057,p = 0.03) compared to controls. Among EA men, the media NAA was 0.067 (IQR (0.03 – 0.10). Similarly among EA men, genetic NAA was significantly lower among cancer cases (0.04 vs. 0.08, p < 0.001). On logistic regression, WAA ≥ 88% was associated with increased odds of PCa in AA men (OR 2.6 95% CI 1.1 – 6.7). In both AAs (OR 0.5, 95% CI 0.2 – 1.1) and EAs (OR 0.3, 95 % 0.2 – 0.6), men in the highest genetic NAA quintile had a decreased risk of PCa on logistic regression compared to men in the lowest quintile. Conclusions: Our study reveals that NAA is protective against PCa diagnosis among all men. Additionally, AA men with a high degree of WAA demonstrate an increased risk of PCa. Future work will focus on determining the association of known prostate cancer risk SNPs such as the 8q24 regions and genetic ancestry.