Bacillus anthracis spore interactions with mammalian cells: Relationship between germination state and the outcome of in vitro

Using an in vitro model of infection, we evaluated the influence of the germination state of B. anthracis spores, as controlled by defined culture conditions, on the outcome of infection. Spores prepared from B. anthracis Sterne 7702 germinated in a variety of common cell culture media supplemented with fetal bovine serum (FBS) while, in the absence of FBS, germination was strictly dependent on medium composition. RAW264.7 macrophage-like cells internalized spores to the same extent in either germinating or non-germinating media. However, significantly more viable, intracellular B. anthracis were recovered from cells infected under non-germinating conditions compared to germinating conditions. At the same time, RAW264.7 cells demonstrated a significant loss in viability when infected under non-germinating conditions.These results suggest that the outcome of host cell infection is sensitive to the germination state of spores at the time of uptake. Moreover, this study demonstrates the efficacy of studying B. anthracis spore infection of host cells within a defined, non-germinating, in vitro environment.Inhalational anthrax commences with the deposition of Bacillus anthracis spores into the bronchioalveolar spaces of the lungs, and culminates with the systemic dissemination of vegetative bacilli within the host [1-3]. Within the lungs, internalization of dormant spores, possibly by several different types of host cells, is believed to be a key step for initiating the transition from the localized to disseminated stages of infection. Alveolar macrophages are reported to transport spores out of the lungs to regional lymph nodes [4-7]. Dendritic cells have also been implicated in the rapid carriage of spores to the draining lymph nodes [8,9]. Finally, alveolar epithelial cells have recently been demonstrated to internalize spores both in vitro and in vivo [10-12], and have been proposed to facilitate the transcytosis of B. anthracis across the epithelial barrier. Taken t