Fig. 4.

The 3D trajectories of TKO parasites are more linear than those of parental parasites. (A) Frequency distribution and statistical analysis of motion-related parameters, including net displacement, run length (distance between pauses >5.4 s), and run speed of RHΔhx parental and TKO parasites without or with cold treatment (CT), and RHΔku80 parental and Δdcx parasites. For cold-treated TKO parasites, intracellular parasite cultures were placed at 7°C for 2 hours before harvesting. For the other samples/conditions, the parasites were harvested as described previously in Munera Lopez et al. (2022). All motility assays were carried out at 37°C. (B) Representative tracks projected on the (x, y) plane with superimposed origins (red circles) for RHΔhx parental and TKO parasites without or with cold -treatment (CT), and RHΔku80 parental and Δdcx parasites. Enlarged insets (4×) include the segments indicated by the dashed boxes in the trajectories. Scale bars: 10 µm. (C) Frequency distribution and statistical analysis of helicity-related parameters including turning angle, dispersal ratio and the pitch of RHΔhx parent and TKO parasites without or with cold-treatment (CT). An illustration defining pitch is included. Data for all parameters are also displayed for RHΔku80 parental and Δdcx parasites except for the pitch. The mean values for individual histograms in A and C are shown at the top (in red) and also marked by the dashed line (see Table S1 for the number of data points for each analysis). P-values (in brackets under the mean values) were calculated by the Kruskal–Wallis test. ns, not significant.