Cat fleas (Ctenocephalides felis) from eight commercial flea colonies from various regions of the USA were examined by selective PCR amplification, and subsequent restriction digest analysis and Southern hybridization of PCR products, for the presence of a rickettsia-like organism (ELB agent). These flea colonies were either started with fleas from one supplier (EL Labs), in which ELB agent was first identified, or were started with fleas from stray cats and dogs and later came into contact with ELB-infected fleas. Infection rates in the colonies ranged from 43% to 93%. The successful propagation of ELB agent in these colonies may be due to efficient trans-stadial and transovarial transmission. While ELB agent has recently been identified in blood from human murine typhus cases, attempts to infect mammalian cells and SCID mice with flea isolates were unsuccessful.