AcMNPV undergoes actin-based motility within the cell cytoplasm. (A, top) High Five cell 30 min after infection with AcMNPV stained for viral capsids (red; anti-VP39 immunofluorescence) and actin (green; FITC-phalloidin). Bar, 10 µm. (inset) Magnified nucleocapsid (of boxed region) with associated actin tail. Bar, 2 µm. (bottom) Time series in 5-s intervals of AcMNPV 3mC virus (red) and EGFP-actin (green) in a High Five cell. Bar, 2 µm. (B) Mean velocity ± SD for AcMNPV 3mC (WT; n = 61), 3mC in the presence of latA (WT + lat; n = 26), 3mC I358A (I358A; n = 92), and 3mC in the presence of Arp2/3 inhibitor CK-548 (WT + CK-548; n = 29) or CK-636 (WT + CK-636; n = 37). ***, P < 0.0001 (by one-way analysis of variance). (C) Histograms of nucleocapsid velocities for 3mC (WT), 3mC in the presence of latA (WT + lat), and 3mC I358A (I358A). (D) Representative tracks of 3mC (WT; top) and 3mC I358A (I358A; bottom) over 135 s. (E) Displacement (straight line distance from initial location to destination), total distance traveled (along path length), and straightness (displacement/total distance traveled) over 50 s for 3mC (WT; n = 58) and 3mC I358A (I358A; n = 58). ***, P < 0.0001 (by unpaired t test). Error bars indicate mean ± SD.