Forested areas adjacent to Milwaukee, Wis., and Chicago, Ill., were investigated for rodents and ticks infected with Borrelia burgdorferi, the causative agent of Lyme disease. White-footed mice (Peromyscus leucopus or Peromyscus maniculatus), meadow voles (Microtus pennsylvanicus), and eastern chipmunks (Tamias striatus) were captured; and specimens from these animals were cultured for B. burgdorferi to define whether the midwestern Lyme disease area currently encompasses these large metropolitan centers. During 1988, B. burgdorferi was successfully cultured from the tissues of two M. pennyslvanicus voles captured from the Chicago area. However, no Ixodes spp. ticks were captured. None of 274 animals captured from sites I3 and 12 additional sites in Wisconsin and Illinois during the summer of 1989 were infected with B. burgdorferi or Ixodes spp. In addition, no ticks were recovered when the underbrush in 11 contiguous areas was flagged. Apparently, B. burgdorferi is rarely found in these areas because of the absence of the appropriate tick vectors. Further studies are needed to monitor the dispersal of B. burgdorferi-infected Ixodes dammini into this heavily populated midwestern region.