Free-living larvae from natural areas in five regions in Switzerland were collected to determine the infection rate of Ixodes ricinus L. larvae by Borrelia burgdorferi Johnson. Larvae were examined for the spirochetes using direct fluorescent antibody assay. Of 652 larvae examined, spirochetes were detected in 20 (3.1%). No differences in infection rate among sites were detected. The low prevalence of B. burgdorferi in larvae, compared with higher infection rate in nymphs (12.8%) and adults (14.5%), suggests that transovarial transmission is inefficient. These results suggest that /. ricinus can serve as a reservoir for B. burgdorferi in nature. Further study is needed on the relative importance of ticks, compared with mammals and birds, as reservoirs for B. burgdorferi.