Abstract: Ehrlichia species are obligatory intracellular bacteria that infect monocytes or macrophages, and cause a tick-borne febrile illness (called ehrlichiosis) in mammals worldwide.Two species of Ehrlichia (E.chaffeensis, and E. ewingii) are known to infect humans, and the severity of the infection ranges from asymptomatic status to death.In Japan, although there is no report of human ehrlichiosis (HE) to date, its potentiality is suggested.Probably the epidemiologic study of Ehrlichia species is insufficient.To survey Ehrlichia species in Japan, we investigated a total of 483 unfed Haemaphysalis longicornis ticks collected in Mie prefecture by p28-targeting PCR assay.Nine of 483 ticks tested were PCR-positive.Multilocus sequence typing (MLST) based on 5 housekeeping genes (16s rDNA, groEL, rpoB, ftsZ, and gltA) was further performed for the characterization of genetic diversity in Ehrlichia species obtained from the 9-positive individuals.The phylogenetic analysis based on the MLST results revealed that Ehrlichia species were divided into 2 clusters, and a high level of genetic variability was detected between them, providing a novel insight into the genetic diversity of Ehrlichia species in Japan.The MLST results also showed that 2 of 9-positive samples shared the high similarities with E. chaffeensis (94.0%-94.1% ), suggesting the potential risk of HE in Japan.