Phylogenetic Signal.

To determine whether the drosophilid species’ social behavior exhibits phylogenetic signal, we applied two tests: Blomberg’s K (29) and Pagel’s λ (30). Both phylogenetic signal tests evaluate whether the observed average trait data across species adhere to a model of Brownian motion across a phylogenetic tree, which approximates the expectation of evolution due to genetic drift (40). Both the K and λ values typically range on a scale from zero to one, where zero indicates low phylogenetic signal (trait evolution does not adhere to Brownian motion) and one indicates high phylogenetic signal (trait evolution adheres to Brownian motion). A published drosophilid phylogenetic tree was applied to these analyses that we pruned to include our species sample (56). Since D. novamexicana was not included in this tree, its placement and branch length were standardized based on another published phylogenetic tree of the virilis group species (57). The tree was made ultrametric in R through the “chronos” function (ape package) for all subsequent phylogenetic comparative analyses. The K and λ tests were implemented in R using the “phylosig” function (phytools package). We tested phylogenetic signal on the four SIN measures and the four behavioral elements where each species’ mean and SEM were incorporated to represent the species’ average and intraspecific variation for the trait, respectively (32). We also tested phylogenetic signal for the distance, angle, and time social spacing parameters. Since some of these variables were manually altered (SI Appendix), we did not incorporate any measure of intraspecific variation into the phylogenetic signal tests. All phylogenetic signal tests were performed separately on the male and female datasets. The estimated K values were statistically evaluated using R through permutation tests where the observed K value was compared with a null distribution of 1,000 K values, each computed by randomly shuffling the tips of the phylogenetic tree. All estimated λ values were statistically evaluated using R through likelihood ratio tests against the null hypothesis that λ = 0. All measures of phylogenetic signal and the associated P values are present in Table 2. To visualize the phylogenetic signal tests for each variable, trait data were mapped onto the drosophilid phylogeny using the “contMap” function (phytools package) in R. This function maps ancestral states for the internal nodes of the phylogeny using maximum likelihood given the trait data for each species at the tips of the tree (58). These phylogenetic trait maps are present in Figs. 2–4. Due to the repeated tests on 11 variables with two phylogenetic signal tests, a Bonferroni correction was employed, and all P values below 0.002 were considered significant.