SUPPLEMENTARY INFORMATION Articles https://doi.org/10.1038/s41559-017-0418-x In the format provided by the authors and unedited. Social transmission of avoidance among predators facilitates the spread of novel prey Rose Thorogood 1,2 *, Hanna Kokko 3 and Johanna Mappes 4 1 Department of Zoology, University of Cambridge, Cambridge, UK. 2 Helsinki Institute of Life Science & Department of Biosciences, University of Helsinki, Helsinki, Finland. 3 Department of Evolutionary Biology and Environmental Studies, University of Zurich, Zurich, Switzerland. 4 Centre of Excellence in Biological Interactions, University of Jyväskylä, Jyväskylä, Finland. *e-mail: rt303@cam.ac.uk Nature Ecology & Evolution www.nature.com/natecolevol 2017 Macmillan Publishers Limited, part of Springer Nature. All rights reserved.
Social transmission of avoidance among predators facilitates the spread of novel prey. Rose Thorogood*, Hanna Kokko, Johanna Mappes. *Corresponding author: rt303@cam.ac.uk Contents Supplementary methods... 2 Validating video playback provides social information for great tits... 2 Supplementary references. 3 Supplementary figures.. 4 Supplementary Figure 1. Validation of video playback... 4 Supplementary Figure 2. Experimental set-up of the Novel World..... 5 Supplementary Figure 3. Relative predation risk for conspicuous versus cryptic prey... 6 Supplementary tables 7 Supplementary Table 1. Summary of best-fit GLMMs of first prey choice 8 Supplementary Table 2. Summary of best-fit GLMMs of predation risk 9 Supplementary videos Supplementary Video 1. Example of social information treatment from validation experiment Supplementary Video 2. Example of control from validation experiment Supplementary Video 3. Example of social information treatment from predation experiment Supplementary Video 4. Example of control from predation experiment 1
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 Supplementary methods Validating video playback provides social information for great tits. Before starting our main experiment, we validated that great tits could respond to social information from conspecifics if this was presented via video playback. Digital representation is becoming a widely used tool for studying animal behavior across taxa, including fish, reptiles, and birds 1. This includes blue tits 1 and great tits 2 where video playback has been used to manipulate foraging and social conditions, respectively. We tested whether a demonstrator s response to an unpalatable food item taken from a specific foraging location altered an observer s preferred foraging location, inspired by experiments conducted previously with real demonstrators 3. In this previous study, great tits demonstrated which coloured cups contained food to observers through a plastic screen 3. In our study, demonstrators were provided with a white plastic feeding cup commonly available in pet stores (90 ml, item #88394, Karlie ), but novel to all birds in our study. The cup contained a mealworm (Tenebrio molitor) that we had made highly unpalatable: mealworms were injected with a saturated chloroquinine phosphate solution, and soaked in this solution for at least 24 h. Demonstrators were filmed as they perched on or next to the cup, before taking the worm in their beak to the perch. Most birds dropped the prey item in seconds, before proceeding to vigorously wipe their beak on the perch, repeatedly, for at least 1 min. See Supplementary movies S1 and S3 for examples of this behavior. Videos were edited using imovie version 10.0 to include 1 min 30 s of the demonstrator taking the worm from the cup, attempting to consume it, and beak wiping. For 30 s before and after the appearance of the demonstrator, we spliced a video clip of an identical cup, but a different color (brown). This ensured that when observers were given a choice of cups to feed from after the video playback, they were not attracted to this alternative simply because of neophilia. See Movies S1 and S2 for example videos used. We next presented videos to 20 observers (5 adult and 5 juvenile females, 6 adult and 4 juvenile males) by playing the video of the appropriate demonstrator of that individual s catch group on a Macbook Pro connected to an LCD computer monitor (Dell Professional P190S 19 ), placed adjacent to the plexiglass front of the test chamber. The video was visible through the plexiglass, and birds clearly reacted when videos began. Cups and birds were approximately life-sized in the videos, when viewed from within the test chamber. Like demonstrators, observers were habituated to these boxes for 2 h before the experiment began. Immediately after the video ended, the screen was removed, and we placed three plastic feeding cups inside the test chamber: the white and brown cups were identical to those seen in the video, and a third (red) cup was provided to ensure that responses were not biased by novelty. Cups were introduced by sliding them inside the test chamber without the birds seeing us or outside the chamber. Observers could not see inside the cups from the perch where they sat. We then recorded which of these cups the observer chose to investigate first (if no choice was made after 45 min we stopped the test, n = 2 birds, both controls). If the 2
44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 61 62 63 64 65 66 67 68 69 70 71 72 73 74 75 76 77 78 video playback conveyed social information about prey to observers, then we predicted avoidance of the white cup. We compared their behavior to a control group that watched video clips of the brown and white cups without a demonstrator present, but for an equal length of time as birds in the socially informed group. Our results (Supplementary Fig. 1) showed that while the control group exhibited a preference to investigate the white cup, the socially-informed birds preferred the alternative brown cup. We could be certain that this was because of the video playback, and not because of an effect of avoiding competition (for example), because prior to beginning our validation experiment, we had tested color preferences of an independent sample of birds. Here we provided 15 birds with the three different colored cups, containing 10 sunflower seeds each. While the food was not novel, the cups were. After 30 min we counted the remaining seeds and if less than 10 had been taken we checked the cups content again after another 30 min. Of these birds, 9 out of 15 took more seeds from the white colored cup, indicating a significant preference for this cup color (Friedman χ 2 = 6.60, d.f. = 2, p = 0.037). Furthermore, with a small sample of additional birds (6 observers, 1 demonstrator), we explored whether observers might copy the foraging preferences of demonstrators if food was palatable (as has been demonstrated both in captivity and in the field previously with great tits 3,4 ). These videos showed a demonstrator discovering a palatable mealworm in the brown cup; subsequently 83% of the observers (5/6, 1 bird visited the red cup) visited this same cup and ignored the preferred white cup. Therefore, we were certain that (i) great tits pay attention to video playback and (ii) they gather information from this to use in their foraging decisions. Supplementary references 1. Hämäläinen, L., Rowland, H. M., Mappes, J. & Thorogood, R. Can video playback provide social information for foraging blue tits? PeerJ 5, e3062 (2017). 2. Snijders, L., Naguib, M. & van Oers, K. Dominance rank and boldness predict social attraction in great tits. Behav. Ecol. 28, 398 406 (2017). 3. Marchetti, C. & Drent, P. J. Individual differences in the use of social information in foraging by captive great tits. Anim. Behav. 60, 131 140 (2000). 4. Aplin, L. M. et al. Experimentally induced innovations lead to persistent culture via conformity in wild birds. Nature 518, 538 541 (2015). 3
79 Supplementary figures 70 * ** NS 60 50 % of choices made 40 30 20 10 80 0 Cup A Social cue of unpalatability Cup B No social cue of unpalatability Preferred feeding location of observers Cup C Novel alternative 81 82 83 84 85 86 87 88 89 90 Supplementary Figure 1. Validation of video playback. Following video playback, we recorded great tits initial foraging choices. Socially-informed birds (grey bars, n = 20) differed from controls (dashed lines, n = 18) overall (G-test: G = 11.89, d.f. = 1, p < 0.001), because they avoided foraging in Cup A (shown in video with a conspecific demonstrator encountering a distasteful prey). Instead, they preferred Cup B (shown in videos but with no demonstrator, i.e. no social information available about the cup s contents). Control birds observed videos of both Cups A and B without a demonstrator, and Cup C was a novel alternative to all birds. Asterisks indicate where cup-choice differed (p < 0.05 using Exact binomial tests) between socially-informed and control birds. 4
One-way observation window Video observation chamber 91 Conspicuous prey Cryptic prey Background signal 92 93 94 95 96 97 98 99 100 101 Supplementary Figure 2. Experimental set-up of Novel World aviary. Following training to open novel artificial prey, great tits were held individually within a video observation chamber, food deprived for 2 hr to ensure they were motivated to forage, and then presented with video playback. Socially-informed birds received a presentation of a cryptic cross prey item for 30 s followed by 1 min of a demonstrator showing a typical disgust response to an unpalatable conspicuous square prey, before another 30 s of the cross prey. Control birds observed the same video, but without a demonstrator present during the 1 min of the square prey item being visible. The video screen was then removed and birds were allowed to enter the aviary room to forage. Drawing not to scale, and great tit image provided under a Creative Commons Deed CC0 licence (from publicdomainvectors.org). 102 103 5
1.5 Socially-informed Control Relative predation risk 1.0 0.5 0.0 1 2 3 104 105 106 107 108 109 110 111 112 113 114 115 Learning trial Supplementary Figure 3. Relative predation risk for novel conspicuous prey versus the cryptic phenotype. The mean (± S.E.) relative predation risk for aposematic prey (number of aposematic prey consumed / number expected by chance) during three learning trials over consecutive days (1 trial/day). Great tits with social information about prey signals (circles) consumed relatively fewer aposematic than cryptic prey, compared to birds with no social information (triangles). Light-colored symbols show individual variation in foraging choices, and the solid reference line indicates equal predation of the cryptic and aposematic prey types. Here we plot raw data values (Fig. 2 in the main manuscript presents results from a model including random effects and a binomial error distribution, see Supplementary Table 2). 6
116 117 118 119 120 121 122 123 124 125 126 127 128 129 Supplementary tables Supplementary Table 1. Summary of best-fit GLMMs testing first prey choice. a,b, Latency to select first prey item and c, symbol type (conspicuous square or cryptic cross) of first prey taken. Socially-informed predators (n = 15) received information via videoplayback that the square-type prey was unpalatable, compared to naïve control predators with no prior information (n = 15). One bird was slow to select the first prey item (644 seconds); results were similar if it was retained (b). Focal birds and a demonstrator were caught in groups of 5 so catching group was included in all models as a random effect. Additional variables added to starting model (~ information treatment) included: latency to enter aviary (birds could move freely from a holding box), whether individual was used in validation experiment (n = 14, 7 in both information groups; see Supplementary methods), age (1 st year vs. adult), and the date during experiment. The best-fit model (with (i) fixed effects and (ii) random effects) was found by (iii) ranking candidate models (with a null model) using AICc. (a) Latency to select first prey item during initial trial: (i) Fixed effects: Estimate Std. Error z p(z) (Intercept) 4.458 0.167 26.677 < 0.001 Information [Social] -1.131 0.244-4.634 < 0.001 (ii) Random effects: Variance Catching group 0 (iii) Alternative models: ~ Information + Latency to enter aviary ~ Information + Used in validation experiment AICc 2.20 2.93 ~ Information + Age 2.94 ~ Information + Date during experiment 2.94 ~ Null model 17.35 (b) Latency to select first prey item during initial trial (outlier included): (i) Fixed effects: Estimate Std. Error z p(z) (Intercept) 5.566 0.458 12.152 < 0.001 Information [Social] -0.669 0.339-1.972 0.049 130 131 Date during experiment* -0.037 0.012-3.081 0.002 7
132 133 Table S1 continued. (ii) Random effects: Variance Catching group 0.012 (iii) Alternative models: AICc ~ Information treatment only 2.88 ~ Information + Used in validation experiment 3.47 ~ Null model 4.02 ~ Information + Age 4.46 ~ Information + Latency to enter aviary 5.76 (c) First prey item taken during initial trial: (i) Fixed effects: Estimate Std. Error z p(z) (Intercept) -0.162 0.569-0.285 0.78 Information [Social] 0.908 0.817 1.111 0.27 (ii) Random effects: Variance Catching group 0.238 (iii) Alternative models: AICc ~ Null model -1.14 ~ Information + Days in experiment -0.52 ~ Information + Age 1.94 ~ Information + Latency to enter aviary 2.59 ~ Information + Used in validation experiment 2.58 * outlier tested on day 3 134 135 8
136 137 138 139 140 141 142 143 144 Supplementary Table 2. Summary of best-fit GLMMs testing if social information affected relative predation risk. a, during initial encounter with the prey population (50% conspicuous signal and unpalatable, 50% cryptic and palatable), and b, across three learning trials held on consecutive days ( Trial ). Sample sizes and additional variables included in models are the same as in Supplementary Table 1. In b, a random slope and intercept were included for each bird to account for repeated testing. The best fit model (with (i) fixed effects and (ii) random effects) was found by (iii) ranking candidate models (with a null model) using AICc. (a) Number of aposematic prey taken (out of total prey choices) during the initial trial: (i) Fixed effects: Estimate Std. Error z p(z) (Intercept) 0.283 0.252 1.123 0.26 Information [Social] -0.649 0.232-2.791 0.005 Latency to enter aviary -0.004 0.002-2.338 0.019 (ii) Random effects: Variance Catching group 0.283 (iii) Alternative models: AICc ~ Information + Used in validation experiment 2.60 ~ Information treatment only 3.04 ~ Information + Age 3.44 ~ Information + Days in experiment 5.58 ~ Null model 7.26 (b) Number of aposematic prey taken (out of total prey choices) during each of three trials: (i) Fixed effects: Estimate Std. Error z p(z) (Intercept) 0.276 0.196 1.412 0.158 Information [social] -0.578 0.176-3.277 0.001 Trial -0.774 0.123-6.285 < 0.001 145 146 Latency to enter aviary -0.004 0.001-2.819 0.005 9
147 148 Table S2 continued. (ii) Random effects: Variance Catching group 0.080 Individual slope 0.119 Individual intercept 0.038 (iii) Alternative models: AICc ~ Information + Trial + Used in validation experiment 5.72 ~ Information + Trial 8.01 ~ Information + Trial + Age 10.05 ~ Information * Trial 10.27 ~ Information + Trial + Days in experiment 10.46 ~ Information only 21.55 149 150 ~ Null model 27.63 10