Rickettsiae are Gram-negative bacteria which multiply only inside host cells and need arthropods either as reservoirs or as vectors. Using the polymerase chain reaction and an automated laser fluorescent DNA sequencer, we amplified and sequenced the 16S rRNA (rDNA) of all available bacteria of the genus Rickettsia. R. tsutsugamushi remained close to the other bacteria of the genus Rickettsia using this technique, contrary to previous conclusions based on the study of the Sta-58 protein antigen. We found that R. canada was not included in the typhus group, as is currently recognized, but was grouped with the rickettsiae of the spotted fever group (SFG). All the SFG rickettsiae tested were grouped in the same cluster (R. conorii, Indian tick typhus rickettsia, Astrakhan fever rickettsia, Israeli tick typhus rickettsia, HA-91, R. sibirica, R. parkeri, "R. africae", "R. slovaca", R. rickettsii, Thai tick typhus rickettsia, R. japonica, R. massiliae, R. rhipicephali, R. montana, two recent isolates GS and Bar 29, R. australis, R. akari, R. bellii and R. helvetica). The recently described ELB bacterium, the agent of the Californian murine typhus, and AB bacterium, a bacterium associated with male killing in the ladybird beetle, were found in this cluster. The sequences of R. conorii Moroccan strain/Indian tick typhus rickettsia, R. massiliae/GS and R. sibirica/HA-91 were identical. All the rickettsiae had a unique ancestor with bacteria also isolated in arthropods (Ehrlichia, Cowdria, Anaplasma, Wolbachia pipientis), eventually pathogenic for mammals and implicated in parthenogenesis and cytoplasmic incompatibility. We conclude that a unique bacterium started a stable association with arthropod ancestors and generated the observed diversity of the currently isolated members of the Rickettsiales.