To test whether rabbits can serve as a model for Lyme neuroborreliosis (LNB), we injected New Zealand white rabbits with Borrelia burgdorferi. We obtained samples of blood and CSF for culture, polymerase chain reaction/hybridization (PCR/H), serology, and Western blot. We then sacrificed the rabbits 2 to 3 months after infection and removed organs for culture and PCR/H analysis. The rabbits developed an antibody response, characterized by ELISA and Western blot analysis, consistent with a systemic infection. PCR/H and blood cultures were negative, but PCR/H analysis of CSF in one rabbit was positive. Positive PCR/H analysis of organs from the rabbits was consistent with widespread dissemination of the organism. The most consistently infected organ was the CNS. To determine whether antigenic variation occurred at the genetic level, we compared the sequences of the OspA gene of both the infecting strain and the brain PCR product. The differences between the two were minimal, indicating that variation in the OspA gene did not occur during in vivo infection in the rabbit. Thus, rabbits, after intradermal inoculation with B burgdorferi, develop a chronic systemic infection in which the CNS is a consistent target. These data indicate that rabbits may be an excellent model of LNB.