Overall, our results demonstrate that different pygmy grasshopper colour morphs vary in susceptibility to visually oriented predators, that different morphs are favoured in different visual environments, and indicate that predation may contribute to selection and spatial and temporal micro-evolutionary shifts of colour morph frequencies in natural populations. We first analyzed encounters between human ‘predators’ and images of natural black, grey and striped colour morphs of the polymorphic Tetrix subulata pygmy grasshoppers presented on background images of unburnt, intermediate or completely burnt natural habitats. We found that the proportion of pygmy grasshoppers that were detected and the time to detection depend both on colour patterns of the grasshoppers and on the visual properties of the background, as well as on their interaction. Our findings thus allow us to reject the null-hypothesis that detection rates of pygmy grasshoppers by visually oriented predators are independent of colour pattern. Additionally, that no single colour pattern offered superior protection against all visual backgrounds, and that the detection rate of each morph changed across backgrounds supports the notion that crypsis is context dependent . The backgrounds used here were chosen such that they could be considered as different types of environments, different succession stages within temporally changing habitats, or different microhabitat patches within heterogeneous environments, and our results therefore have broad applicability.
Tsurui et al.  reported different detection rates by humans of Tetrix japonica pygmy grasshopper colour morphs against grass and sand backgrounds, but there was no indication in their study that morphs were more common in those habitat types in which they were most cryptic. Our present finding that the protective value of the black morph increases with increasing proportion burnt substrate in the visual background corroborates the results of a previous experiment that used a similar approach , and is consistent with the demonstration that the spatiotemporal variation in the incidence of the melanistic morph in natural T. subulata pygmy grasshopper populations is correlated with habitat modifications associated with succession in post-fire environments. Specifically, very high proportions of melanistic individuals in sooty environments the first year after fire are followed by rapid declines associated with recovery of vegetation and the gradual disappearance of burnt substrates . That rates of detection were associated with differences in the frequencies of striped, black and grey morphs both within habitat types and across populations that occupy different types of environments (Table 1, Figure 5) adds further support to the notion [7, 28] that selection for camouflage, particularly background matching is an important driver of the evolutionary dynamics of the exuberant colour polymorphism in pygmy grasshoppers.
Presumably, a high degree of background matching in one background often comes with the cost of low degree of matching in other, visually different backgrounds . The consequences of such a trade-off depend on the scale of spatiotemporal variation relative to the mobility, lifespan and reproductive life history of the organism [34–38]. For instance, it has been proposed that selection in variable environments may promote the maintenance of polymorphism [35, 36]. Under other conditions, environmental heterogeneity may instead select for a ‘compromise’ colour pattern that may not offer superior protection in any one habitat type but provides the best solution across a range of different habitats . The striped T. subulata morph might represent such a compromise. It was detected at the lowest rate both in the non-burnt and in the semi-burnt treatments, and it was only against the completely burnt background that it was more easily detected than the black morph. A possible explanation for this may be that the striped pattern provides protection not only via camouflage by means of background matching, it may also impair detection or recognition by disrupting the body outline of the prey [39, 40], or by closely resembling inedible objects in the environment, such as dry spruce needles, twigs or straws, i.e., via masquerade .
In heterogeneous environments, individuals may reduce the risk of predation further by adopting a matching habitat choice and utilize to disproportionate degrees substrate patches in the environment that offer the highest concealment . Accordingly, Gillis  showed that green and red morphs of the grasshopper Circotettix rabula rabula each prefer to rest on different but matching backgrounds. Similarly, colour morphs of our current study species T. subulata differently utilize alternative microhabitats and surface substrates .
We found that the grey morph was detected at the highest rate against all three visual backgrounds. This finding is in good agreement with the observation that the frequency of grey individuals in samples from natural populations was relatively low, ca. 5%, across all three environments (Table 1), but it does not provide an explanation for the continued persistence of the grey morph. However, our three habitat types did not include all the available backgrounds influencing selection on crypsis in the pygmy grasshopper. Furthermore, visual predation is only one of many selective agents that influence the evolutionary dynamics of colour morph frequencies. Pygmy grasshopper colour morphs represent integrated phenotypes that differ in a suite of ecologically important traits such as preferred body temperatures, thermal physiology, reproductive life-history (egg and clutch size, inter-clutch interval), body size, predator avoidance behaviour, microhabitat utilization and diet e.g., [7, 45, 46] and references therein. Fitness differences among individuals that belong to different morphs are thus influenced not only by how difficult they are to detect, but also by selection that operates on characters and aspects of performance that are directly influenced by (such as warming up rates) or associated with colour pattern.
To use humans as ‘predators’ in detection tasks is an increasingly used approach in studies of function and evolution of protective coloration (Figure 1), but few attempts have been made to assess whether such experiments allow for reliable inferences about the influence of selection imposed by natural predators in the wild. A striking similarity has been reported between humans and birds with regard to the ability to recognize and discriminate conspicuous colour patterns presented against homogeneous backgrounds under artificial laboratory conditions [25, 26]. We carried out a series of comparisons that enabled us to evaluate and validate the approach for detection of camouflaged prey in a more natural setting. First, we showed that differences in rates of detection among morphs presented on computer screens mirrored the previously reported  differences in rates of capture by humans of free-ranging grasshoppers in the wild. The effect of colour pattern on the relative ease with which humans can detect images of (motionless) grasshoppers in a two-dimensional representation is thus similar to the effect of colour pattern on the ability of humans to detect and capture live grasshoppers in their natural environment. Second, and more importantly, rates of detection on computer screens were correlated with estimates of survival (adjusted for differences in capture probabilities) of striped, black and grey female (but not male) T. subulata in the wild, thus demonstrating that detection rates by humans may reliably predict selection imposed by natural visual predators, such as birds e.g., . That colour pattern differently influences survival of male and female pygmy grasshoppers in the wild can be attributed to females being larger, utilizing different microhabitats, and being more active (longer average daily movement distances) than males , since the protective value of a given colour pattern may depend on body size , behaviours and movement patterns , and visual backgrounds ; this study. In the current study, however, body size and behaviours did not differ between images that represented different colour morphs and body size corresponded to that of large females. Third and finally, detection rates of grasshopper images were correlated with relative frequencies of striped, black and grey morphs in 27 samples of more than 4,000 T. subulata individuals collected from natural populations that occupied the same habitat types that were used as visual backgrounds in the detection experiment. These findings implicate visual predation as an important driver of evolutionary modifications of colour polymorphism in pygmy grasshoppers, and demonstrate that, in our system, using humans as ‘predators’ in detection experiments contributes reliable and relevant information that enhances our understanding of natural selection and evolution.