The two-component system (TCS) composed of a pair of a
sensor histidine kinase and a response regulator, allows bacteria to sense signals and respond to changes in their environment through specific gene activation or repression. The present study examined TCS in the obligatory intracellular bacteria Ehrlichia chaffeensis and Anaplasma phagocytophilum, that cause human monocytic
ehrlichiosis (HME) and
human granulocytic anaplasmosis (HGA) respectively. The genomes of E. chaffeensis and A. phagocytophilum were each predicted to encode three pairs of TCSs. All six genes encoding three
histidine kinases and three response regulators were expressed in both E. chaffeensis and A. phagocytophilum cultured in human leukocytes. Pretreatment of host cell-free E. chaffeensis or A. phagocytophilum with
closantel, an inhibitor of
histidine kinases, completely blocked the
infection of host cells. Treatment of infected cells 1 day post
infection with
closantel cleared
infection in dose-dependent manner. All six genes in E. chaffeensis were cloned,
recombinant proteins were expressed, and polyclonal
antibodies were produced. Double immunofluorescence labelling and Western blot analysis revealed that all six
proteins were expressed in cell culture. Autokinase activities of the three recombinant
histidine kinases from E. chaffeensis were inhibited by
closantel in vitro. A number of E. chaffeensis genes, including the six TCS genes, were downregulated within 5-60 min post
closantel treatment. These results suggest that these TCSs play an essential role in
infection and survival of E. chaffeensis and A. phagocytophilum in human leukocytes.