Transmission of hepatitis C virus (HCV) was studied in a colony of 85 chimpanzees using assays for anti-HCV and HCV-RNA. Thirteen of the 85 sera were positive for anti-HCV, and 12 of the 13 were also positive for HCV-RNA. All of the anti-HCV positive sera except one were obtained from chimpanzees which had been inoculated with non-A, non-B hepatitis virus. On the other hand, only one of 63 sera of chimpanzees without history of experimental infection of the virus was positive for anti-HCV. Transmission to this chimpanzee was thought to be a needle contaminated with HCV. All 39 samples of chimpanzees born in the center were negative for both anti-HCV and HCV-RNA. Sixteen of their mothers had undergone experimental infection, and 6 of them were positive for both anti-HCV and HCV-RNA. These results suggest that nonpercutaneous transmission, including sexual and mother-to-infant transmissions, is not an important mode of transmission. If these findings apply to humans, definition of inapparent sources of the infection is needed.