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We demonstrated that different seroprotective groups exist among isolates of Borrelia burgdorferi and Borrelia garinii. The major group was composed of isolates 297, B31, S-1-10, MMTI, IPT, and ATCC 35211 and 21 isolates obtained from California, Illinois, New York, Texas, and Wisconsin. A second group was composed of European isolates PBi and G25. A third group was composed of a single isolate, C-1-11. These groupings were supported by Western immunoblot findings. In addition, the seroprotective groups were confirmed by passive transfer of immune sera and challenge of recipient hamsters with the homologous isolate or other isolates of B. burgdorferi or B. garinii. These studies demonstrate that a monovalent vaccine will not provide complete protection against infection with all isolates of B. burgdorferi.