Iridescence is a striking and taxonomically widespread form of animal coloration [], but that its intense and varying hues could function as concealment [] rather than signaling seems completely counterintuitive. Here, we show that the color changeability of biological iridescence, produced by multilayer cuticle reflectors in jewel beetle (Sternocera aequisignata) wing cases, provides effective protection against predation by birds. Importantly, we also show that the most likely mechanism to explain this increase in survival is camouflage and not some other protective function, such as aposematism. In two field experiments using wild birds and humans, we measured both the “survival” and direct detectability of iridescent and non-iridescent beetle models and demonstrated that the iridescent treatment fared best in both experiments. We also show that an increased level of specular reflection (gloss) of the leaf background leads to an increase in the survival of all targets and, for detectability by humans, enhances the camouflage effect of iridescence. The latter suggests that some prey, particularly iridescent ones, can increase their chance of survival against visually hunting predators even further by choosing glossier backgrounds. Our study is the first to present direct empirical evidence that biological iridescence can work as a form of camouflage, providing an adaptive explanation for its taxonomically widespread occurrence.

Concealing-Coloration in the Animal Kingdom: an Exposition of the Laws of Disguise through Color and Pattern: Being a Summary of Abbott H. Thayer’s Discoveries.

Results and Discussion

3 Stevens M.

Merilaita S. Animal Camouflage: Mechanisms and Function. 4 Cuthill I.C.

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et al. The biology of color. 5 Ruxton G.D.

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Speed M.P. Avoiding Attack: the Evolutionary Ecology of Crypsis, Aposematism, and Mimicry. 1 Doucet S.M.

Meadows M.G. Iridescence: a functional perspective. 1 Doucet S.M.

Meadows M.G. Iridescence: a functional perspective. 6 Stavenga D.G.

Wilts B.D.

Leertouwer H.L.

Hariyama T. Polarized iridescence of the multilayered elytra of the Japanese jewel beetle, Chrysochroa fulgidissima. 7 Barrows F.P.

Bartl M.H. Photonic structures in biology: a possible blueprint for nanotechnology. 8 Moyroud E.

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et al. Disorder in convergent floral nanostructures enhances signalling to bees. 9 Whitney H.M.

Kolle M.

Andrew P.

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Steiner U.

Glover B.J. Floral iridescence, produced by diffractive optics, acts as a cue for animal pollinators. 10 Wilts B.D.

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Stavenga D.G. Sparkling feather reflections of a bird-of-paradise explained by finite-difference time-domain modeling. 1 Doucet S.M.

Meadows M.G. Iridescence: a functional perspective. 11 Komdeur J.

Oorebeek M.

van Overveld T.

Cuthill I.C. Mutual ornamentation, age, and reproductive performance in the European starling. 12 Rutowski R.L.

Kemp D.J. Female iridescent colour ornamentation in a butterfly that displays mutual ornamentation: is it a sexual signal?. 13 Steinbrecht R.A. Fine structure and development of the silver and golden cuticle in butterfly pupae. 14 Seago A.E.

Brady P.

Vigneron J.P.

Schultz T.D. Gold bugs and beyond: a review of iridescence and structural colour mechanisms in beetles (Coleoptera). 15 Fabricant S.A.

Exnerová A.

Ježová D.

Štys P. Scared by shiny? The value of iridescence in aposematic signalling of the hibiscus harlequin bug. 16 Waldron S.J.

Endler J.A.

Valkonen J.K.

Honma A.

Dobler S.

Mappes J. Experimental evidence suggests that specular reflectance and glossy appearance help amplify warning signals. 2 Thayer G.H. Concealing-Coloration in the Animal Kingdom: an Exposition of the Laws of Disguise through Color and Pattern: Being a Summary of Abbott H. Thayer’s Discoveries. Protective coloration in animals, including various forms of camouflage, aposematism, and mimicry, provides multiple ways for prey to escape predation []. However, the adaptive function of a vivid type of biological coloration, iridescence, is not fully understood []. Iridescence is generated by nanostructures that produce intensely chromatic colors that shift with changing angle of view or illumination []. This variability can produce a striking visual appearance and make objects more conspicuous. Due to this, iridescence is often coupled with a signaling function [], frequently driven by sexual selection []. However, iridescence is also common in many monomorphic species. Although mutual ornamentation could be explained by sexual selection in some cases [], and correlated selection remains a viable explanation for the female ornament in others [], natural selection remains a possibility. Importantly, iridescence is found in non-reproductive stages, such as caterpillars and butterfly chrysalises []. Here, sexual selection seems unlikely, although a warning role (aposematism) is certainly possible []. Instead, the “father of camouflage,” Abbott Thayer [], suggested that iridescence in many animals is actually camouflage, because the directionality of color in iridescent animals makes them appear “dissolved into many depths and distances” (p. 87). Thus, one of the first functional hypotheses for biological iridescence was that it conceals rather than reveals.

17 Pike T.W. Interference coloration as an anti-predator defence. 18 Kjernsmo K.

Hall J.R.

Doyle C.

Khuzayim N.

Cuthill I.C.

Scott-Samuel N.E.

Whitney H.M. Iridescence impairs object recognition in bumblebees. 17 Pike T.W. Interference coloration as an anti-predator defence. 18 Kjernsmo K.

Hall J.R.

Doyle C.

Khuzayim N.

Cuthill I.C.

Scott-Samuel N.E.

Whitney H.M. Iridescence impairs object recognition in bumblebees. Despite being proposed more than a century ago, empirical support for Thayer’s theory of iridescence as camouflage has only appeared very recently []. These studies confirmed that iridescence appeared to interfere with the ability of birds to successfully strike at simulated virtual prey [] and with the ability of bees to identify a target shape []. Impeding prey recognition may therefore explain the evolution of iridescence in many monomorphic species. However, does biological iridescence actually provide a survival advantage against birds, likely to be one of the most important predators of iridescent insects, and, if so, what is the underlying mechanism: camouflage or aposematism?

Figure 1 Prey Target Designs for All Six Treatments Show full caption From left to right: iridescent on privet, static rainbow on bramble, green on beech, black on holly, purple on English ivy, and blue on bramble. Also illustrated in these images is the varying level of specular highlights (gloss) between the different backgrounds. Using real, multilayer iridescent wing cases of the Asian jewel beetle (Sternocera aequisignata) and non-iridescent beetle wing case models as prey ( Figure 1 ), we investigated these fundamental questions about the adaptive function of iridescence. In two separate field experiments, we tested the effects of iridescence on both survival and detectability of the prey targets. In experiment 1, we studied the survival of iridescent and non-iridescent targets against predation by wild birds in a natural setting. In experiment 2, we used humans as surrogate predators who searched for these targets in the same woodland location, enabling us to measure directly the detectability of iridescent and non-iridescent targets (see STAR Methods for full details). Controls included targets with the same reflectance peaks as seen in the iridescent targets (green, blue, and purple) and the same base color (black) and, to distinguish the benefits of having changeable colors from being multicolored but non-iridescent, targets wrapped with calibrated photos of the iridescent beetle cuticle (henceforth “static rainbow”; Figure 1 ).

19 Whitney H.M.

Rands S.A.

Elton N.J.

Ellis A.G. A technique for measuring petal gloss, with examples from the Namaqualand flora. 20 Merilaita S.

Scott-Samuel N.E.

Cuthill I.C. How camouflage works. For the bird experiment, we predicted that, if iridescence provides a survival advantage for prey, iridescent prey should have a higher probability of surviving than non-iridescent prey. For the human experiment, we predicted that, if the mechanism providing a survival benefit for iridescent prey was camouflage, the iridescent prey should be harder to detect. If, on the other hand, survival from the bird experiment is due to aposematism or neophobia, we predicted that the iridescent prey should be easy to detect in the human experiment. Finally, as the surface of these iridescent beetles also produces specular reflection of white light (hereafter, gloss, e.g., []), we also predicted an interactive effect of background gloss: iridescent beetles on glossy leaves should have a lower signal-to-noise ratio and thus be less detectable [].

2 = 95.302; degrees of freedom [df] = 5; p < 0.001), with iridescent targets surviving better than all others except black (versus static rainbow, z = 4.05, p < 0.001; versus green, z = 2.26, p = 0.024; versus violet, z = 7.64, p < 0.001; versus blue, z = 6.87, p < 0.001; versus black, z = 0.93, p = 0.350). The human experiment mirrored the bird experiment (2 = 699.13; df = 5; p < 0.001) and iridescent targets being less detectable than all others except black (versus static rainbow, z = 7.86, p < 0.001; versus green, z = 6.50, p < 0.001; versus purple, z = 17.00, p < 0.001; versus blue, z = 17.29, p < 0.001; versus black, z = 1.57, p = 0.118). For those targets that were detected, detection distance varied with treatment (GLMM with log-normal errors; F 5,1265 = 27.54; p < 0.001). Iridescent targets were detected further away than black (t 1266 = −2.72; p = 0.007) and at a similar distance to static rainbow targets (t 1266 = 0.90; p = 0.367), but participants needed to be closer to them than green, purple, or blue to detect them (t 1265 = 2.12, p = 0.033; t 1266 = 3.61, p < 0.001; t 1266 = 6.11, p < 0.001, respectively). Figure 2 Avian Predation and Human Detection Data Show full caption (A) Odds ratios (±95% confidence intervals [CIs]) from Cox mixed-model survival analysis comparing all treatments to the iridescent in the avian predation experiment. (B) Mean (±95% CI) probability of detecting targets for each treatment in the human detection experiment. In both experiments, the iridescent targets survived significantly better than all except the black treatment. Bla, black; Blu, blue; Gre, green; Irid, iridescent; Pur, purple; Stat, static rainbow. See also Figure S1 A. In the bird experiment, a total of 646 out of 886 targets (73%) showed evidence of avian predation; the rest were treated as censored in the survival analysis. Treatment affected relative mortality ( Figure 2 A; mixed-model Cox regression χ= 95.302; degrees of freedom [df] = 5; p < 0.001), with iridescent targets surviving better than all others except black (versus static rainbow, z = 4.05, p < 0.001; versus green, z = 2.26, p = 0.024; versus violet, z = 7.64, p < 0.001; versus blue, z = 6.87, p < 0.001; versus black, z = 0.93, p = 0.350). The human experiment mirrored the bird experiment ( Figure 2 B), with treatment affecting detection probability (generalized linear mixed model [GLMM] with binomial errors; χ= 699.13; df = 5; p < 0.001) and iridescent targets being less detectable than all others except black (versus static rainbow, z = 7.86, p < 0.001; versus green, z = 6.50, p < 0.001; versus purple, z = 17.00, p < 0.001; versus blue, z = 17.29, p < 0.001; versus black, z = 1.57, p = 0.118). For those targets that were detected, detection distance varied with treatment (GLMM with log-normal errors; F= 27.54; p < 0.001). Iridescent targets were detected further away than black (t= −2.72; p = 0.007) and at a similar distance to static rainbow targets (t= 0.90; p = 0.367), but participants needed to be closer to them than green, purple, or blue to detect them (t= 2.12, p = 0.033; t= 3.61, p < 0.001; t= 6.11, p < 0.001, respectively).

We next performed secondary analyses, including natural variation in background gloss as a covariate. In the bird experiment, there was no significant interaction between treatment and gloss (χ2 = 5.29; df = 5; p = 0.381), but average survival increased with the glossiness of the substrate (χ2 = 18.80; df = 1; p < 0.001) and again differed between treatments (χ2 = 96.70; df = 5; p < 0.001). Planned comparisons between the iridescent and five other treatments confirmed our primary analysis that the iridescent targets survived better than all treatments except black (versus static rainbow, z = 4.01, p < 0.001; versus green, z = 2.30, p = 0.022; versus violet, z = 7.68, p < 0.001; versus blue, z = 6.96, p < 0.001; versus black, z = 0.95, p = 0.340).

2 = 16.58; df = 5; p = 0.005). The decrease in detection probability with increasing substrate gloss was significantly steeper for iridescent targets than green, purple, and black targets, but not static rainbow or blue (2 = 16.40; df = 5; p = 0.006). The decrease in detection distance with increasing substrate gloss was significantly steeper for iridescent targets than all others ( Figure 3 Results from the Human Detection Experiment Show full caption Mean probability of detecting targets (A) and mean detection distance (B) as a function of gloss for each treatment. Lines are best fits from GLMMs. The iridescent targets became significantly more difficult to detect as substrate gloss increased, more so than other treatments. See also Figures S1 C and S1D. Table 1 Human Experiment: Effect of Substrate Gloss on the Probability and Distance of Target Detection Probability of Detection Slope t p Slope versus Irid (t) Slope versus Irid (p) Intercept Intercept versus Irid (t) Intercept versus Irid (p) Iridescent −0.15 −2.63 0.009 – – −1.09 – – Static −0.05 −1.68 0.093 1.43 0.154 −0.34 3.11 0.002 Green 0.00 0.01 0.994 2.64 0.008 −0.88 1.13 0.258 Purple 0.00 0.05 0.958 2.04 0.042 0.88 6.26 <0.001 Blue −0.07 −1.52 0.128 1.45 0.148 1.40 7.59 <0.001 Black 0.06 1.69 0.090 3.21 0.001 −1.70 −1.97 0.048 Distance Iridescent −0.06 −2.25 0.027 – – 1.44 – – Static −0.01 −0.93 0.352 2.32 0.021 1.36 −1.05 0.293 Green −0.03 −2.37 0.019 2.27 0.024 1.49 −0.17 0.863 Purple −0.02 −1.08 0.280 2.11 0.035 1.44 −0.09 0.929 Blue 0.01 1.45 0.148 3.50 0.000 1.47 −0.06 0.956 Black 0.00 −0.28 0.784 2.87 0.004 0.97 −3.56 <0.001 Parameter estimates from GLMMs (binomial distribution, logit link) for detection probability and LMMs for detection distance (log transformed). All treatment slopes and intercepts are tested against the corresponding estimates for the iridescent treatment. Testing significance of individual intercepts is not of interest, only differences between treatment intercepts. In the human experiment, there was a significant interaction between gloss and treatment for the probability of detecting a target ( Figure 3 A; χ= 16.58; df = 5; p = 0.005). The decrease in detection probability with increasing substrate gloss was significantly steeper for iridescent targets than green, purple, and black targets, but not static rainbow or blue ( Table 1 ). When testing the relationship between gloss and treatment using GLMMs in which the iridescent treatment was contrasted against all the other treatments, the negative relationship with gloss was only significant for the iridescent treatment ( Table 1 ). There was also a significant interaction between treatment and gloss for detection distance ( Figure 3 B; χ= 16.40; df = 5; p = 0.006). The decrease in detection distance with increasing substrate gloss was significantly steeper for iridescent targets than all others ( Table 1 ). Indeed, the negative relationship with gloss was only significant for the iridescent and green treatments ( Table 1 ).

∗target interaction was non-significant, but the trends were similar ( 20 Merilaita S.

Scott-Samuel N.E.

Cuthill I.C. How camouflage works. 21 Dimitrova M.

Merilaita S. Prey concealment: visual background complexity and prey contrast distribution. 22 Xiao F.

Cuthill I.C. Background complexity and the detectability of camouflaged targets by birds and humans. 23 Kjernsmo K.

Merilaita S. Background choice as an anti-predator strategy: the roles of background matching and visual complexity in the habitat choice of the least killifish. For humans, iridescent and black targets were least detectable on matte leaves, but on glossy leaves, iridescent targets outperformed black ( Figure 3 ). In the bird experiment, the glosstarget interaction was non-significant, but the trends were similar ( Figure S1 A), and the main effect of background gloss was still to reduce mortality. Taken together, the results suggest that an increase in the level of background specularity reduces detectability. This is plausibly because high specular reflectance acts as “visual noise,” decreasing the signal-to-noise ratio for target detection []. However, the disproportionate benefit to iridescent targets was not simply because they were glossier than other treatments ( Figure S4 B) or lighter in terms of diffuse reflectance ( Figure S4 A). The iridescent and static rainbow treatments had (by design) very similar lightness (and color) under diffuse illumination and yet very different survival. The black treatment was most different in lightness from the types of plant used in the study and much darker than any other treatment yet survived as well, and was as hard to detect, as the iridescent treatment on matte leaves. Previous studies on humans and birds have found that an increase of the visual complexity of the background makes objects harder to find [] and that some prey, in the presence of a predator, actively choose more visually complex backgrounds over those they simply match []. Specularity of the background is an under-researched factor affecting visual search that, given the effects we have demonstrated, merits attention. We predict that rainfall may have similar effects to leaf gloss.

24 Hegna R.H.

Nokelainen O.

Hegna J.R.

Mappes J. To quiver or to shiver: increased melanization benefits thermoregulation, but reduces warning signal efficacy in the wood tiger moth. With these experiments, we have clearly demonstrated an anti-predator function of iridescence: in both experiments, with wild birds or using humans as surrogate predators, the iridescent treatment fared best in terms of survival and probability of remaining undetected, particularly on glossy leaves. Importantly, the results from the human experiment clearly demonstrate that iridescence significantly lowers the probability of being detected, strongly suggesting camouflage as the underlying mechanism for the anti-predator function of iridescence. Non-visual factors that vary between plants and affect bird foraging (e.g., insect abundance) cannot account for the human detection results, and we note, the treatment differences in the bird experiment remain the same even when we limit the analysis to the single, commonest plant substrate, common ivy (Hedera helix; Figures S4 B and S4C and accompanying analyses). It is noteworthy that the black treatment also fared well in both experiments, providing an adaptive explanation as to why so many insects in nature are black, in addition to any thermoregulatory benefits of melanization []. This is an important result, because it demonstrates that, in terms of visual perception of predators, there is no evident cost for prey to be iridescent compared to being black. Indeed, when adding the effect of background gloss, the iridescent treatment fared better than the black.