Healthcare workers (HCWs) have an occupational risk of infection with hepatitis C virus (HCV). However, data regarding the magnitude of this risk are limited. We conducted a prospective study on a cohort of 24 HCWs who were exposed to HCV by needlestick injuries involving 25 patients. All source patients were viremic with a mean HCV-RNA level of 1.65 megagenomic equivalents per milliliter. At least 64% of patients were infected with HCV serotype 4 (Simmond's classification). After a follow up period of at least 6 months, none of the exposed HCWs acquired HCV. Thus, HCV does not seem to be easily transmitted by needlestick injuries. However, further large-scale studies are needed for a more accurate estimation of the risk of transmission.