A) The C. neoformans (Cn) cells were incubated alone in
PBS, with live and heat-killed A. castellanii (Ac),
live and heat-killed J774.14 macrophage-like cells (Mac), and
polystyrene beads. Live amoebae and macrophages were more effective than
heat-killed cells in triggering capsular enlargement. In contrast, there
was no enlargement in the presence of the polystyrene beads, when
compared to incubation in PBS alone. Each assay was performed
independently on 3 separate days and the numbers shown represent the
average of a minimum of 50 C. neoformans cells. B)
Capsule enlargement activity of A. castellanii extracts
as a function of temperature. Activity assays were done at 4°C, room
temperature (RT), 28°C, and 37°C. This experiment was done two
times on separate days and each condition represents a minimum of 50
cells. C) Representative C. neoformans cells suspended
in India ink showing enlargement of the polysaccharide capsule after
incubation with A. castellani (40x). (*) indicates
P<0.001 for capsule enlargement for each condition when compared to
control incubation alone in PBS.