Fig. 3

Chamber height and fly density affect behavior. A Track straightness was higher at 3 mm-height chamber. B Unconnected social network prop. was lower at 3 mm-height chamber. C Network degree was higher at 3 mm-height chamber. D Representative motion trace for female flies in 2 mm-height and 3 mm-height chamber. E Move time prop. at arena centre was lower at 3 mm-height chamber. F Move time prop. at arena edge showed no difference between 3 mm-height or 2 mm-height chambers. G No significant change was observed for males in 2 mm-height chamber given the metric total move time. H, Space distance at arena centre was larger in 2 mm-height chamber. I, Space distance at arena edge was not affected by chamber height. J Female flies showed changed social space distance in the 2 mm-height chamber. K locomotion behavior deviation of female flies was about 1.4 times that of male flies in 2 mm-height chamber. L, social behavior deviation of female flies was roughly same with that of male flies in 2 mm-height chamber. M No significant change on fly speed was identified with different density (4, 6, or 8 flies) in the chamber. N No significant change on fly move time was identified with different density in the chamber. O Angular velocity changed with different density. P The changes of angular velocity happened at arena edge. Q Only a little change of angular velocity was found at arena centre with different density. R fly turning was associated with interaction events. S, Interaction episode count significantly increased with higher fly density. Statistical analysis: Welch Two Sample t-test was used for comparisons between two groups. *P < 0.05, **P < 0.01, ***P < 0.001. Non-significant test was not annotated in the graph for brevity. A modified Cohen’s d was used to measure standardized effect size. The effect size for female flies and male flies was linear regressed to assess the spatial restriction on flies at K and L. For experiment on behavior changes on chamber height, n = 26 videos for male w¹¹¹⁸ flies in 2 mm-height chambers, n = 26 videos for female w¹¹¹⁸ flies in 2 mm-height chambers, n = 26 videos for male w¹¹¹⁸ flies in 3 mm-height chambers, n = 25 videos for female w¹¹¹⁸ flies in 3 mm-height chambers were recorded and analyzed. For experiment on behavior changes on fly density, n = 24 videos for male w¹¹¹⁸ flies in 4-flies chambers, n = 24 videos for male w¹¹¹⁸ flies in 6-flies chambers, n = 23 videos for male w¹¹¹⁸ flies in 8-flies chambers, n = 24 videos for female w¹¹¹⁸ flies in 4-flies chambers, n = 24 videos for female w¹¹¹⁸ flies in 6-flies chambers, n = 24 videos for female w¹¹¹⁸ flies in 8-flies chambers were recorded and analyzed. All videos recorded were from three independent experiment replicates