Survival of Coxiella burnetii within free-living amoeba Acanthamoeba castellanii.
Coxiella burnetii was co-cultured with the free-living amoeba A. castellanii. C. burnetii within amoebae was observed using confocal and electron microscopy. Detection of surviving C. burnetii within amoebae and the effect of infection on the amoebae was assessed by titration of surviving bacteria on shell vials and counting the amoeba. Results were expressed as association index (the product of the mean number of bacteria per infected amoeba and the percentage of infected amoebae). The association index decreased in a time-dependent manner. At day 1, all amoebae were infected by a mean of 5.25 bacteria per amoeba. A decrease of the association index was mostly observed until day 7, when the amoeba population steadily increased. After day 7, both remained stable. After 30 days, 52% of the amoebae remained infected with an average of 2 bacteria per amoeba. After 3 days of cultivation, bacteria were found in vacuoles within the amoebae, some of which appeared to be in spore-like forms. The number of viable C. burnetii increased slightly until day 6 and decreased thereafter. The decrease was significantly more rapid in bacteria cultivated in axenic medium and in amoebic lysate than with live amoebae. Furthermore, no viable bacteria were detected after day 15 in flasks where bacteria were cultivated alone or with amoebic lysate, whereas in flasks where bacteria were co-cultivated with amoebae at day 18 numerous viable bacteria remained detectable. It is concluded that free-living amoebae could support the survival and resistance of C. burnetii in an environmental setting.