Predator selection on multicomponent warning signals in an aposematic moth.

Aposematic prey advertise their unprofitability with conspicuous warning signals that are often composed of multiple color patterns. Many species show intraspecific variation in these patterns even though selection is expected to favor invariable warning signals that enhance predator learning. However, if predators acquire avoidance to specific signal components, this might relax selection on other aposematic traits and explain variability. Here, we investigated this idea in the aposematic moth Amata nigriceps that has conspicuous black and orange coloration. The size of the orange spots in the wings is highly variable between individuals, whereas the number and width of orange abdominal stripes remains consistent. We produced artificial moths that varied in the proportion of orange in the wings or the presence of abdominal stripes. We presented these to a natural avian predator, the noisy miner (Manorina melanocephala), and recorded how different warning signal components influenced their attack decisions. When moth models had orange stripes on the abdomen, birds did not discriminate between different wing signals. However, when the stripes on the abdomen were removed, birds chose the model with smaller wing spots. In addition, we found that birds were more likely to attack moths with a smaller number of abdominal stripes. Together, our results suggest that bird predators primarily pay attention to the abdominal stripes of A. nigriceps, and this could relax selection on wing coloration. Our study highlights the importance of considering individual warning signal components if we are to understand how predation shapes selection on prey warning coloration.

Predator selection on phenotypic variability of cryptic and aposematic moths.

Natural selection generally favours phenotypic variability in camouflaged organisms, whereas aposematic organisms are expected to evolve a more uniform warning coloration. However, no comprehensive analysis of the phenotypic consequences of predator selection in aposematic and cryptic species exists. Using state-of-the-art image analysis, we examine 2800 wing images of 82 moth species accessed via three online museum databases. We test whether anti-predator strategy (i.e., camouflage or aposematism) explains intraspecific variation in wing colour and pattern across northern hemisphere moths. In addition, we test two mutually non-exclusive, ecological hypotheses to explain variation in colour pattern: diel-activity or dietary-niche. In this work, taking into account phylogenetic relationships, moth phenotypic variability is best explained by anti-predator strategy with camouflaged moths being more variable in wing patterning than aposematic species.

Diet influences resource allocation in chemical defence but not melanin synthesis in an aposematic moth.

For animals that synthesise their chemical compounds de novo, resources, particularly proteins, can influence investment in chemical defences and nitrogen-based wing colouration such as melanin. Competing for the same resources often leads to trade-offs in resource allocation. We manipulated protein availability in the larval diet of the wood tiger moth, Arctia plantaginis, to test how early life resource availability influences relevant life history traits, melanin production and chemical defences. We expected higher dietary protein to result in more effective chemical defences in adult moths and a higher amount of melanin in the wings. According to the resource allocation hypothesis, we also expected individuals with less melanin to have more resources to allocate to chemical defences. We found that protein-deprived moths had a slower larval development, and their chemical defences were less unpalatable for bird predators, but the expression of melanin in their wings did not differ from that of moths raised on a high-protein diet. The amount of melanin in the wings, however, unexpectedly correlated positively with chemical defences. Our findings demonstrate that the resources available in early life have an important role in the efficacy of chemical defences, but melanin-based warning colours are less sensitive to resource variability than other fitness-related traits.

Colour polymorphism associated with a gene duplication in male wood tiger moths.

Colour is often used as an aposematic warning signal, with predator learning expected to lead to a single colour pattern within a population. However, there are many puzzling cases where aposematic signals are also polymorphic. The wood tiger moth, Arctia plantaginis, displays bright hindwing colours associated with unpalatability, and males have discrete colour morphs which vary in frequency between localities. In Finland, both white and yellow morphs can be found, and these colour morphs also differ in behavioural and life-history traits. Here, we show that male colour is linked to an extra copy of a yellow family gene that is only present in the white morphs. This white-specific duplication, which we name valkea, is highly upregulated during wing development. CRISPR targeting valkea resulted in editing of both valkea and its paralog, yellow-e, and led to the production of yellow wings. We also characterise the pigments responsible for yellow, white, and black colouration, showing that yellow is partly produced by pheomelanins, while black is dopamine-derived eumelanin. Our results add to a growing number of studies on the genetic architecture of complex and seemingly paradoxical polymorphisms, and the role of gene duplications and structural variation in adaptive evolution.

Linking Predator Responses to Alkaloid Variability in Poison Frogs.

Many chemically-defended/aposematic species rely on diet for sequestering the toxins with which they defend themselves. This dietary acquisition can lead to variable chemical defenses across space, as the community composition of chemical sources is likely to vary across the range of (an aposematic) species. We characterized the alkaloid content of two populations of the Dyeing Poison Frog (Dendrobates tinctorius) in northeastern French Guiana. Additionally, we conducted unpalatability experiments with naive predators, Blue Tits (Cyanistes caeruleus), using whole-skin secretion cocktails to assess how a model predator would respond to the defense of individuals from each population. While there was some overlap between the two D. tinctorius populations in terms of alkaloid content, our analysis revealed that these two populations are markedly distinct in terms of overall alkaloid profiles. Predator responses to skin secretions differed between the populations. We identified 15 candidate alkaloids (including three previously undescribed) in seven classes that are correlated with predator response in one frog population. We describe alkaloid profile differences between populations for D. tinctorius and provide a novel method for assessing unpalatability of skin secretions and identifying which toxins may contribute to the predator response. In one population, our results suggest 15 alkaloids that are implicated in predator aversive response. This method is the first step in identifying the causal link between alkaloids and behavioral responses of predators, and thus makes sense of how varying alkaloid combinations are capable of eliciting consistent behavioral responses, and eventually driving evolutionary change in aposematic characters (or characteristics).

Not just the sum of its parts: Geographic variation and nonadditive effects of pyrazines in the chemical defence of an aposematic moth.

Chemical defences often vary within and between populations both in quantity and quality, which is puzzling if prey survival is dependent on the strength of the defence. We investigated the within- and between-population variability in chemical defence of the wood tiger moth (Arctia plantaginis). The major components of its defences, SBMP (2-sec-butyl-3-methoxypyrazine) and IBMP (2-isobutyl-3-methoxypyrazine), are volatiles that deter bird attacks. We hypothesized that (1) variation in the chemical defences of male wood tiger moths reflects the local predation pressure; (2) observed differences in quantity and quality of defence among populations have a genetic basis; and (3) increasing concentrations of SBMP and IBMP will elicit greater aversive reactions in predators, with the two pyrazines having an additive effect on predators' avoidance. We found that (1) the chemical defence of wild moths partly reflects local predator selection: high predation pressure populations (Scotland and Georgia) had stronger chemical defences, but not lower variance, than the low-predation populations (Estonia and Finland). (2) Based on the common garden results, both genetic and environmental components seem to influence the strength of chemical defence in moth populations; and (3) IBMP alone did not provide protection against bird predators but worked against bird attacks only when combined with SBMP, and while SBMP was more effective at higher concentrations, IBMP was not. Altogether this suggests that, when it comes to pyrazine concentration, more is not always better, highlighting the importance of testing the efficacy of chemical defence and its components with relevant predators, as extrapolating from chemical data may be less than straightforward.

Host's genetic background determines the outcome of reciprocal faecal transplantation on life-history traits and microbiome composition.

BACKGROUND: Microbes play a role in their host's fundamental ecological, chemical, and physiological processes. Host life-history traits from defence to growth are therefore determined not only by the abiotic environment and genotype but also by microbiota composition. However, the relative importance and interactive effects of these factors may vary between organisms. Such connections remain particularly elusive in Lepidoptera, which have been argued to lack a permanent microbiome and have microbiota primarily determined by their diet and environment. We tested the microbiome specificity and its influence on life-history traits of two colour genotypes of the wood tiger moth (Arctia plantaginis) that differ in several traits, including growth. All individuals were grown in the laboratory for several generations with standardized conditions. We analyzed the bacterial community of the genotypes before and after a reciprocal frass (i.e., larval faeces) transplantation and followed growth rate, pupal mass, and the production of defensive secretion. RESULTS: After transplantation, the fast-growing genotype grew significantly slower compared to the controls, but the slow-growing genotype did not change its growth rate. The frass transplant also increased the volume of defensive secretions in the fast-growing genotype but did not affect pupal mass. Overall, the fast-growing genotype appeared more susceptible to the transplantation than the slow-growing genotype. Microbiome differences between the genotypes strongly suggest genotype-based selective filtering of bacteria from the diet and environment. A novel cluster of insect-associated Erysipelotrichaceae was exclusive to the fast-growing genotype, and specific Enterococcaceae were characteristic to the slow-growing genotype. These Enterococcaceae became more prevalent in the fast-growing genotype after the transplant, which suggests that a slower growth rate is potentially related to their presence. CONCLUSIONS: We show that reciprocal frass transplantation can reverse some genotype-specific life-history traits in a lepidopteran host. The results indicate that genotype-specific selective filtering can fine-tune the bacterial community at specific life stages and tissues like the larval frass, even against a background of a highly variable community with stochastic assembly. Altogether, our findings suggest that the host's genotype can influence its susceptibility to being colonized by microbiota, impacting key life-history traits.

Heterozygote advantage and pleiotropy contribute to intraspecific color trait variability.

The persistence of intrapopulation phenotypic variation typically requires some form of balancing selection because drift and directional selection eventually erode genetic variation. Heterozygote advantage remains a classic explanation for the maintenance of genetic variation in the face of selection. However, examples of heterozygote advantage, other than those associated with disease resistance, are rather uncommon. Across most of its distribution, males of the aposematic moth Arctia plantaginis have two hindwing phenotypes determined by a heritable one locus-two allele polymorphism (genotypes: WW/Wy = white morph, yy = yellow morph). Using genotyped moths, we show that the presence of one or two copies of the yellow allele affects several life-history traits. Reproductive output of both males and females and female mating success are negatively affected by two copies of the yellow allele. Females carrying one yellow allele (i.e., Wy) have higher fertility, hatching success, and offspring survival than either homozygote, thus leading to strong heterozygote advantage. Our results indicate strong female contribution especially at the postcopulatory stage in maintaining the color polymorphism. The interplay between heterozygote advantage, yellow allele pleiotropic effect, and morph-specific predation pressure may exert balancing selection on the color locus, suggesting that color polymorphism may be maintained through complex interactions between natural and sexual selection.

Rich resource environment of fish farms facilitates phenotypic variation and virulence in an opportunistic fish pathogen.

Phenotypic variation is suggested to facilitate the persistence of environmentally growing pathogens under environmental change. Here, we hypothesized that the intensive farming environment induces higher phenotypic variation in microbial pathogens than natural environment, because of high stochasticity for growth and stronger survival selection compared to the natural environment. We tested the hypothesis with an opportunistic fish pathogen Flavobacterium columnare isolated either from fish farms or from natural waters. We measured growth parameters of two morphotypes from all isolates in different resource concentrations and two temperatures relevant for the occurrence of disease epidemics at farms and tested their virulence using a zebrafish (Danio rerio) infection model. According to our hypothesis, isolates originating from the fish farms had higher phenotypic variation in growth between the morphotypes than the isolates from natural waters. The difference was more pronounced in higher resource concentrations and the higher temperature, suggesting that phenotypic variation is driven by the exploitation of increased outside-host resources at farms. Phenotypic variation of virulence was not observed based on isolate origin but only based on morphotype. However, when in contact with the larger fish, the less virulent morphotype of some of the isolates also had high virulence. As the less virulent morphotype also had higher growth rate in outside-host resources, the results suggest that both morphotypes can contribute to F. columnare epidemics at fish farms, especially with current prospects of warming temperatures. Our results suggest that higher phenotypic variation per se does not lead to higher virulence, but that environmental conditions at fish farms could select isolates with high phenotypic variation in bacterial population and hence affect evolution in F. columnare at fish farms. Our results highlight the multifaceted effects of human-induced environmental alterations in shaping epidemiology and evolution in microbial pathogens.

Genetic colour variation visible for predators and conspecifics is concealed from humans in a polymorphic moth.

The definition of colour polymorphism is intuitive: genetic variants express discretely coloured phenotypes. This classification is, however, elusive as humans form subjective categories or ignore differences that cannot be seen by human eyes. We demonstrate an example of a 'cryptic morph' in a polymorphic wood tiger moth (Arctia plantaginis), a phenomenon that may be common among well-studied species. We used pedigree data from nearly 20,000 individuals to infer the inheritance of hindwing colouration. The evidence supports a single Mendelian locus with two alleles in males: WW and Wy produce the white and yy the yellow hindwing colour. The inheritance could not be resolved in females as their hindwing colour varies continuously with no clear link with male genotypes. Next, we investigated if the male genotype can be predicted from their phenotype by machine learning algorithms and by human observers. Linear discriminant analysis grouped male genotypes with 97% accuracy, whereas humans could only group the yy genotype. Using vision modelling, we also tested whether the genotypes have differential discriminability to humans, moth conspecifics and their bird predators. The human perception was poor separating the genotypes, but avian and moth vision models with ultraviolet sensitivity could separate white WW and Wy males. We emphasize the importance of objective methodology when studying colour polymorphism. Our findings indicate that by-eye categorization methods may be problematic, because humans fail to see differences that can be visible for relevant receivers. Ultimately, receivers equipped with different perception than ours may impose selection to morphs hidden from human sight.

The Effect of Predator Population Dynamics on Batesian Mimicry Complexes.

AbstractUnderstanding Batesian mimicry is a classic problem in evolutionary biology. In Batesian mimicry, a defended species (the model) is mimicked by an undefended species (the mimic). Prior theories have emphasized the role of predator behavior and learning as well as evolution in model-mimic complexes but have not examined the role of population dynamics in potentially governing the relative abundances and even persistence of model-mimic systems. Here, we examined the effect of the population dynamics of predators and alternative prey on the prevalence of warning-signaling prey composed of models and mimics. Using optimal foraging theory and signal detection theory, we found that the inclusion of predator and alternative prey population dynamics could reverse traditional theoretical predictions: as alternative prey increase in numbers, mimics suffer because larger populations of predators are maintained, resulting in apparent competition. Under some circumstances, apparent competition affects model populations as well, although not as severely as it affects mimics. Our results bear on the intriguing puzzle that in nature warning signals are relatively scarce, yet experiments suggest that such signals can be highly advantageous. The availability of alternative prey and numerical responses by predators can overwhelm advantages observed in experiments to keep warning signals in model-mimic systems relatively scarce.

Context-dependent coloration of prey and predator decision making in contrasting light environments.

A big question in behavioral ecology is what drives diversity of color signals. One possible explanation is that environmental conditions, such as light environment, may alter visual signaling of prey, which could affect predator decision-making. Here, we tested the context-dependent predator selection on prey coloration. In the first experiment, we tested detectability of artificial visual stimuli to blue tits (Cyanistes caeruleus) by manipulating stimulus luminance and chromatic context of the background. We expected the presence of the chromatic context to facilitate faster target detection. As expected, blue tits found targets on chromatic yellow background faster than on achromatic grey background whereas in the latter, targets were found with smaller contrast differences to the background. In the second experiment, we tested the effect of two light environments on the survival of aposematic, color polymorphic wood tiger moth (Arctia plantaginis). As luminance contrast should be more detectable than chromatic contrast in low light intensities, we expected birds, if they find the moths aversive, to avoid the white morph which is more conspicuous than the yellow morph in low light (and vice versa in bright light). Alternatively, birds may attack first moths that are more detectable. We found birds to attack yellow moths first in low light conditions, whereas white moths were attacked first more frequently in bright light conditions. Our results show that light environments affect predator foraging decisions, which may facilitate context-dependent selection on visual signals and diversity of prey phenotypes in the wild.

Complex plant quality-microbiota-population interactions modulate the response of a specialist herbivore to the defence of its host plant.

Many specialist herbivores have evolved strategies to cope with plant defences, with gut microbiota potentially participating to such adaptations.In this study, we assessed whether the history of plant use (population origin) and microbiota may interact with plant defence adaptation.We tested whether microbiota enhance the performance of Melitaea cinxia larvae on their host plant, Plantago lanceolata and increase their ability to cope the defensive compounds, iridoid glycosides (IGs).The gut microbiota were significantly affected by both larval population origin and host plant IG level. Contrary to our prediction, impoverishing the microbiota with antibiotic treatment did not reduce larval performance.As expected for this specialized insect herbivore, sequestration of one of IGs was higher in larvae fed with plants producing higher concentration of IGs. These larvae also showed metabolic signature of intoxication (i.e. decrease in Lysine levels). However, intoxication on highly defended plants was only observed when larvae with a history of poorly defended plants were simultaneously treated with antibiotics.Our results suggest that both adaptation and microbiota contribute to the metabolic response of herbivores to plant defence though complex interactions. Read the free Plain Language Summary for this article on the Journal blog.

De nombreux herbivores spécialistes ont évolué vers des stratégies qui leurs permettent de contourner les défenses de leur plantes hôtes. Le microbiote pourrait potentiellement participer à certaines de ces adaptations.Dans cette étude, nous avons essayé de déterminer si l'adaptation d'un herbivore est influencée par son microbiote et l'historique d'utilisation de sa plante hôte (origine de la population).Nous avons testé en quoi le microbiote contribue à la performance de chenilles Melitaea cinxia sur leur plante hôte Plantago lanceolata ainsi que leur capacité à faire face aux glucosides d'iridoïdes (GI), des molécules de défenses produites par P. lanceolata.Comme attendu, la concentration de GI stockée était plus importante chez les chenilles qui étaient nourries avec des plantes produisant de fortes concentrations de GI. Ces chenilles présentaient par ailleurs des signes d'intoxications (i.e. diminution de la concentration de Lysine). Cependant cela n'était visible que lorsque les chenilles étaient issues de populations qui se nourrissaient historiquement sur des plantes peu défendues et lorsqu'elles étaient simultanément traitées par des antibiotiques.Nos résultats suggèrent donc que des processus complexes d'adaptation couplés à l'activité du microbiote contribuent à la réponse des herbivores aux défenses de leurs plante hôtes.

Antibiotics accelerate growth at the expense of immunity.

Antibiotics have long been used in the raising of animals for agricultural, industrial or laboratory use. The use of subtherapeutic doses in diets of terrestrial and aquatic animals to promote growth is common and highly debated. Despite their vast application in animal husbandry, knowledge about the mechanisms behind growth promotion is minimal, particularly at the molecular level. Evidence from evolutionary research shows that immunocompetence is resource-limited, and hence expected to trade off with other resource-demanding processes, such as growth. Here, we ask if accelerated growth caused by antibiotics can be explained by genome-wide trade-offs between growth and costly immunocompetence. We explored this idea by injecting broad-spectrum antibiotics into wood tiger moth (Arctia plantaginis) larvae during development. We follow several life-history traits and analyse gene expression (RNA-seq) and bacterial (r16S) profiles. Moths treated with antibiotics show a substantial depletion of bacterial taxa, faster growth rate, a significant downregulation of genes involved in immunity and significant upregulation of growth-related genes. These results suggest that the presence of antibiotics may aid in up-keeping the immune system. Hence, by reducing the resource load of this costly process, bodily resources may be reallocated to other key processes such as growth.

The complete mitochondrial genome of the wood tiger moth (Arctia plantaginis) and phylogenetic analyses within Arctiinae.

We report the assembly and annotation of the complete mitochondrial genome of the warningly-coloured wood tiger moth (Arctia plantaginis) and investigate its phylogenetic position within Arctiinae. The A.plantaginis mitogenome is 15,479 bp long with 13 protein-coding genes, 22 transfer RNAs, 2 ribosomal RNA genes, and an A + T-rich region (D-loop). The phylogenetic analyses based on 13 protein-coding genes showed A.plantaginis clustering within a clade of species with white wings and yellow or red bodies. This result can be useful in understanding the evolution of coloration in Arctiid moths.

Safety in Numbers: How Color Morph Frequency Affects Predation Risk in an Aposematic Moth.

AbstractPolymorphic warning signals in aposematic systems are enigmatic because predator learning should favor the most common form, creating positive frequency-dependent survival. However, many populations exhibit variation in warning signals. There are various selective mechanisms that can counter positive frequency-dependent selection and lead to temporal or spatial warning signal diversification. Examining these mechanisms and their effects requires first confirming whether the most common morphs are favored at both local and regional scales. Empirical examples of this are uncommon and often include potentially confounding factors, such as a lack of knowledge of predator identity and behavior. We tested how bird behavior influences the survival of three coexisting morphs of the aposematic wood tiger moth Arctia plantaginis offered to a sympatric predator (great tit Parus major) at different frequencies. We found that although positive frequency-dependent selection is present, its strength is affected by predator characteristics and varying prey profitability. These results highlight the need to understand predator foraging in natural communities with variable prey defenses in order to better examine how behavioral interactions shape evolutionary outcomes.

Evolutionary importance of intraspecific variation in sex pheromones.

Sex pheromones in many insect species are important species-recognition signals that attract conspecifics and inhibit attraction between heterospecifics; therefore, sex pheromones have predominantly been considered to evolve due to interactions between species. Recent research, however, is uncovering roles for these signals in mate choice, and that variation within and between populations can be drivers of species evolution. Variation in pheromone communication channels arises from a combination of context-dependent, condition-dependent, or genetic mechanisms in both signalers and receivers. Variation can affect mate choice and thus gene flow between individuals and populations, affecting species' evolution. The complex interactions between intraspecific and interspecific selection forces calls for more integrative studies to understand the evolution of sex pheromone communication.

Social transmission in the wild can reduce predation pressure on novel prey signals.

Social transmission of information is taxonomically widespread and could have profound effects on the ecological and evolutionary dynamics of animal communities. Demonstrating this in the wild, however, has been challenging. Here we show by field experiment that social transmission among predators can shape how selection acts on prey defences. Using artificial prey and a novel approach in statistical analyses of social networks, we find that blue tit (Cyanistes caeruleus) and great tit (Parus major) predators learn about prey defences by watching others. This shifts population preferences rapidly to match changes in prey profitability, and reduces predation pressure from naïve predators. Our results may help resolve how costly prey defences are maintained despite influxes of naïve juvenile predators, and suggest that accounting for social transmission is essential if we are to understand coevolutionary processes.