Leptospirosis is a zoonotic infection of worldwide distribution that is commonly endemic in tropical regions. In the United States, the annual incidence rate in the state of Hawaii is approximately 30 times higher than that reported nationally. U.S. troops deployed to disease-endemic tropical environments and those training in the state of Hawaii are potentially at high risk for acquiring leptospirosis. Serum and risk factor data were obtained from 488 blood donors attending military-sponsored blood drives on the island of Oahu. Testing of sera for the presence of Leptospira-specific antibodies was performed with microscopic agglutination testing (MAT). Seven active duty U.S. Army soldiers were found to be seropositive by MAT (seroprevalence, 1.4%), 2 of 76 newly arrived residents (2.6%) and 5 of 412 established residents (1.2%). Positive MAT results were statistically more common among female subjects and those 18 to 30 years old. Seropositivity was not associated with specific exposures or prolonged residence in the state of Hawaii.