In 2003, a Leptospira survey was performed on Yoroshima Island of the Amami Islands located in the southwestern part of Japan. Seven Leptospira strains were isolated from the field rat Rattus rattus, which were identified as L. borgpetersenii by flaB sequencing, 16S rDNA sequencing and gyrB sequencing, and serovar Javanica was determined by a microscopic agglutination test. NotI-long restriction fragment analysis indicated that these isolates were genetically indistinguishable from an isolate from the Okinawa Islands. The present results suggest that L. borgpetersenii is migrating into the Amami Islands in Japan.