A 2 years old child living in an area of the State of São Paulo, known in the past as endemic for rickettsiosis developed clinical evidences of spotted fever after a tick bite. Rickettsiae were isolated from guinea pigs inoculated with a skin homogenate. In sera tested by indirect immunofluorescence with Rickettsia rickettsii standard antigen, IgG specific antibody titers raised from 1:512 in the first sample to 1:2048 in the third one; IgM specific antibody titer was 1:128 in the three samples. Also positive were sera obtained from the inoculated guinea pigs. In the last 20 years no other case of rickettsial spotted fever has been confirmed by isolation of the agent in Brasil. To our knowlwdge, there are no previous reports of isolation of Rickettsiae through inoculation of skin biopsy homogenates.