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Purpose: The aim of this study was to determine the seroprevalence of Lymes disease in a population at risk in south India. Methods: Prospective ongoing study and included screening of forest workers and staff of Nagarahole and Bandipur forest ranges in South India for Lymes disease. Screening included a detailed questionnaire for Lymes disease, complete ocular and systemic examination by an ophthalmologist and infectious disease specialist and blood collection. ELISA for IgM and IgG antibodies for Borrelia burgdorferi were performed on the collected sera samples. Western blot confirmation was done on the seropositive samples. Ticks were also collected from these forest areas for future studies to detect if they harbor B. burgdorferi. Results: Seroprevalence of 19.9% was noted by ELISA. Western blot confirmation was seen in 15.6% of the seropositive samples. There was significant correlation between seropositivity and exposure to tick bites (P = 0.023). Conclusion: There is a high seroprevalence of infection with B. burgdorferi in the forest areas of Nagarahole and Bandipur ranges in south India.