FIG 7.

Δash4 parasites have dispersion defects following egress. (A) Graph of average percentages of vacuoles that contain 2, 4, 8, >8, or “other” parasites (i.e., vacuoles with a non-power of 2 number of parasites or vacuoles with <8 parasites that could not be confidently counted). Error bars indicate standard deviations from the means. Results represent three independent experimental replicates with at least 80 vacuoles counted per experiment. Statistical significance was determined initially by chi-square analysis of counts, and data were significant (P < 0.05). To identify where the statistical significance lay, unpaired 2-tailed student’s T tests were performed. All statistically significant comparisons are shown (*, P < 0.05). (B). Graph of the average percentages of invasion of wild-type and Δash4 parasites normalized to average percentage of wild-type invasion. Error bars represent standard deviations from the means. No statistical significance was found by an unpaired 2-tailed Student's t test. (C) Scatter plot of average root mean squared displacement (RMSD) of wild-type, Δash4, and ASH4 rescue (Δash4 ASH4) vacuoles at 50 s postegress. At least 30 vacuoles with 8 parasites were analyzed for each strain. Red lines indicate means ± standard errors of the means (SEM). One-way ANOVA was performed with a subsequent Tukey’s multiple-comparison test to determine significance, and all statistically significant comparisons are indicated (*, P <0.05; **, P < 0.01. (D) Scatter plot of average RMSD for wild-type and ASH4 rescue parasites based on whether they were derived from ordered rosettes or nonrosetted vacuoles at 50 s postegress. Red lines indicate means ± SEM. In total, 46 ordered vacuoles and 19 disordered vacuoles were used in the analysis. Statistical significance was determined by an unpaired 2-tailed Student's t test. **, P < 0.01. (E) Model for why Δash4 parasites form small plaques. Wild-type parasites (green) successfully divide and have an ordered architecture and normal dispersion after egress. Δash4 parasites (blue) have endodyogeny defects leading to fewer viable parasites overall and to parasites that organize as pairs, resulting in a disordered intravacuolar architecture, which leads to a reduction in the ability to efficiently disperse from host cells following egress.