Intracellular survival and replication of Vibrio cholerae O139 in aquatic free-living amoebae

Vibrio cholerae is a highly infectious bacterium responsible for large outbreaks of cholera among humans at regular intervals. A seasonal distribution of epidemics is known but the role of naturally occurring habitats are virtually unknown. Plankton has been suggested to play a role, because bacteria can attach to such organisms forming a biofilm. Acanthamoebea castellanii is an environmental amoeba that has been shown to be able to ingest and promote growth of several bacteria of different origin. The aim of the present study was to determine whether or not an intra-amoebic behaviour of V. cholerae O139 exists. Interaction between these microorganisms in co-culture was studied by culturable counts, gentamicin assay, electron microscopy, and polymerase chain reaction. The interaction resulted in intra-amoebic growth and survival of V. cholerae in the cytoplasm of trophozoites as well as in the cysts of A. castellanii. These data show symbiosis between these microorganisms, a facultative intracellular behaviour of V. cholerae contradicting the generally held view, and a role of free-living amoebae as hosts for V. cholerae O139. Taken together, this opens new doors to study the ecology, immunity, epidemiology, and treatment of cholera.