The frequency of coinfection with Borrelia burgdorferi and either Anaplasma phagocytophila or Babesia microti among patients with erythema migrans, the initial skin lesion of Lyme disease, was assessed in 2 mainland locations in Rhode Island and Connecticut in a 4-year prospective study. Of the 93 patients with culture-proven erythema migrans, 2 (2%) patients had coinfection with A. phagocytophila and 2 (2%) had coinfection with B. microti. We concluded that the frequency of coinfection with these agents was low among patients with erythema migrans in the study areas.