Fig 1.
Tracking and behavior classification of flies with Flyworld and Flytracker.
(A) Schematics of Flyworld, with camera, illumination system, and walking arena. (B-D) Example trajectories of different numbers (1, 16 and 50) of male flies tracked by Flytracker. (E) A touch event where one fly’s head approaches another’s tail in darkness, recorded with an infrared camera. (F) Adjacency matrix of touch behaviors. Colors represent frequencies of touch behavior. Mathematically, no self-interaction was defined.
Table 1.
Tracking results of Ctrax and Flytracker with different numbers of flies and videos at different frame rates.
Fig 2.
Effects of social isolation on the parameters of average networks.
(A) The Total Walking Distance in the isolated group is significantly higher than in the control group (p<0.0001). (B) No significant difference is found between the Weighted Total Interaction of the control and isolation groups. (C and D) A significant increase of Weighted Degree and Clustering Coefficient was observed between average networks of control and isolation groups (p<0.0001). (E and F) No significant difference was found between the global parameters Global Efficiency and Assortativity of the control and isolation groups. Average networks representing 10 repeats of experiment on control (G) and isolation (H) groups. All plots use 2.5th to 97.5th percentiles as whiskers and 25th to 75th percentiles as box, with the median value in the middle of the box (n = 10).
Fig 3.
The change of average networks of the control and isolated groups with changing isolation time interval.
Ctr_1 to Ctr_6 are averaged networks from flies in the control group from 1 to 6 days of preparation. Iso_1 to Iso_6 are averaged networks from flies in the isolation group kept in isolation from 1 to 6 days. Each average network is the mean graph of the networks from 10 biological repeats. Nodes represent “average flies” and edges represent the average number of interactions from one “average fly” to another. Isolated flies generally display more interactions than control flies.
Fig 4.
Changes of locomotor activity and social network parameters with changing isolation time interval.
(A) The Total Walking Distance, represented by its median with an interquartile range, shows significant differences between control and isolation groups (p<0.0001) except after 1 day of isolation. Different parameters reveal differences between control (blue) and isolated (red) groups over time. Significance is denoted with asterisks, the number of which represents the negative order of magnitude of the p value. Data are shown as median values with interquartile ranges, with 10 repeats on each isolation day (n = 10). (B) Individual parameter Weighted Degree of control and isolated flies shows a significant difference from day 1 which increases significantly after day 2. (C) Difference in the local parameter Clustering Coefficient also shows from day 1 and increases over 6 days. (D) Global parameter Weighted Total Interaction significantly differs only on day3 and day5 and not on other days. (E) Global parameter Global Efficiency significantly differs on day3 and day5 only. (F) No significant difference was found on the global parameter Assortativity.
Fig 5.
Genome-wide gene analysis of expression and physiological changes.
(A, B) Scatter plots of log2 of fold changes versus log10 of gene counts on day 1 and day 6. Red up triangles show genes whose expression levels are significantly increased more than 1.5 folds. Blue down triangles show genes whose expression levels are significantly decreased more than 1.5 folds. Blue lines show the ±log2 1.5 thresholds. (C) Plot of the number of genes whose expression is upregulated (blue), downregulated (red), or changed either way (green) specifically between isolated and control groups on Days 1, 2, 3 and 6. All experiments were repeated 3 times. (D, E) Gene Ontology analysis of differentially expressed genes on day one. Most changed genes on day one has been clustered into metabolism related categories, in both up- and down-regulated genes. (F) Lipid/dry-weight changes between control and isolation groups in male and female flies. All groups are paired and paired t-test show significant increase in lipid/dry-weight proportion in isolated flies than control male flies (n = 25, P = 0.0063). Isolated female flies also have increased lipid proportion to dry-weight than control flies (n = 22, P = 0.0233).