Cuticular Hydrocarbon Composition Research Articles

Pleiotropic Effects of ebony and tan on Pigmentation and Cuticular Hydrocarbon Composition in Drosophila melanogaster.

Pleiotropic genes are genes that affect more than one trait. For example, many genes required for pigmentation in the fruit fly Drosophila melanogaster also affect traits such as circadian rhythms, vision, and mating behavior. Here, we present evidence that two pigmentation genes, ebony and tan, which encode enzymes catalyzing reciprocal reactions in the melanin biosynthesis pathway, also affect cuticular hydrocarbon (CHC) composition in D. melanogaster females. More specifically, we report that ebony loss-of-function mutants have a CHC profile that is biased toward long (>25C) chain CHCs, whereas tan loss-of-function mutants have a CHC profile that is biased toward short (<25C) chain CHCs. Moreover, pharmacological inhibition of dopamine synthesis, a key step in the melanin synthesis pathway, reversed the changes in CHC composition seen in ebony mutants, making the CHC profiles similar to those seen in tan mutants. These observations suggest that genetic variation affecting ebony and/or tan activity might cause correlated changes in pigmentation and CHC composition in natural populations. We tested this possibility using the Drosophila Genetic Reference Panel (DGRP) and found that CHC composition covaried with pigmentation as well as levels of ebony and tan expression in newly eclosed adults in a manner consistent with the ebony and tan mutant phenotypes. These data suggest that the pleiotropic effects of ebony and tan might contribute to covariation of pigmentation and CHC profiles in Drosophila.

Cuticular hydrocarbon composition does not allow Harmonia axyridis males to identify the mating status of sexual partners

Cuticular hydrocarbon composition does not allow Harmonia axyridis males to identify the mating status of sexual partners

Communication versus waterproofing: the physics of insect cuticular hydrocarbons.

Understanding the evolution of complex traits is among the major challenges in biology. One such trait is the cuticular hydrocarbon (CHC) layer in insects. It protects against desiccation and provides communication signals, especially in social insects. CHC composition is highly diverse within and across species. To understand the adaptive value of this chemical diversity, we must understand how it affects biological functionality. So far, CHCs have received ample research attention, but their physical properties were little studied. We argue that these properties determine their biological functionality, and are vital to understanding how CHC composition affects their adaptive value. We investigated melting behaviour and viscosity of CHCs from 11 ant species using differential scanning calorimetry and a novel microrheological technique. CHCs began melting below -45°C, and often were entirely liquid only above 30°C. Thus, they formed a solid-liquid mixture under ambient conditions, which contrasts to previous assumptions of entirely solid layers in many species. This may be adaptive as only biphasic CHC layers ensure uniform coating of the insect body, which is necessary for waterproofing. CHC viscosity was mostly between 0.1 and 0.2 Pa s-1, thus similar to motor oils. Surprisingly, chemically different CHC profiles had similar viscosities, suggesting that a certain viscosity level is adaptive and ensures that communication signals can be perceived. With this study, we draw attention to the importance of studying the physics of CHC layers. Only by understanding how chemical and physical mechanisms enable CHC functionality can we understand the causes and consequences of CHC diversification.

Determining social and population structures requires multiple approaches: A case study of the desert ant Cataglyphis israelensis.

The remarkable diversity of ant social organization is reflected in both their life history and population kin structure. Different species demonstrate a high variation with respect to both social structure and mating strategies: from the ancestral colony type that is composed of a single queen (monogyny), singly inseminated (monoandry), to the more derived states of colonies headed by a multiply inseminated queen (polyandry), to colonies composed of multiple queens (polygyny) that are either singly or multiply inseminated. Moreover, the population structure of an ant species can range from multicoloniality to polydomy to supercoloniality, and Cataglyphis is considered to be a model genus in regard to such diversity. The present study sought to determine the social and population structure of the recently described C. israelensis species in Israel. For this purpose we employed a multidisciplinary approach, rather than the commonly used single approach that is mostly based on genetics. Our study encompassed behavior (nest insularity/openness), chemistry (composition of nestmate recognition signals and cuticular hydrocarbons), and genetics (microsatellite polymorphism). Each approach has been shown to possess both advantages and disadvantages, depending on the studied species. Our findings reveal that C. israelensis colonies are headed by a single, multiply inseminated queen and that the population structure is polydomous, with each colony comprising one main nest and several additional satellite nests. Moreover, our findings demonstrate that none of the above‐noted approaches, when employed individually, is suitable or sufficient in itself for delineating population structure, thus emphasizing the importance of using multiple approaches when assessing such complex systems.

Chemical profiles in Iberoformica subrufa and Formica frontalis, a new example of temporary host–parasite interaction

The aim of this paper is to describe the relationship between two ant species, Formica frontalis and Iberoformica subrufa, found together in shared nests. Therefore, we obtained data from dug nests and outdoor activity in two sympatric populations and investigated the cuticular hydrocarbons (CHCs) in both sympatric populations and in 10 I. subrufa allopatric populations to unravel whether the relationship becomes tuned between both species. We also determined the CHCs of two sympatric Serviformica species (F. cunicularia and F. lemani). Our results showed that the ant F. frontalis is a temporary parasite of I. subrufa which facultatively forms mixed colonies complying with a loose form of the Emery’s rule. Alkanes and methylalkanes are the most abundant compounds found in F. frontalis and I. subrufa CHCs, respectively, but esters were only abundant in I. subrufa. As far as the CHC similarity is concerned, the sympatric free-living hosts were chemically closer to the parasite, albeit not identical, whereas the allopatric I. subrufa populations always maintained a separate CHC composition. We provide different potential hypotheses to explain this similarity of cuticular profiles only in the two geographically distant sympatric populations.

Diapause affects cuticular hydrocarbon composition and mating behavior of both sexes in Drosophila montana.

Environmental cues, mainly photoperiod and temperature, are known to control female adult reproductive diapause in several insect species. Diapause enhances female survival during adverse conditions and postpones progeny production to the favorable season. Male diapause (a reversible inability to inseminate receptive females) has been studied much less than female diapause. However, if the males maximized their chances to fertilize females while minimizing their energy expenditure, they would be expected to be in diapause at the same time as females. We investigated Drosophila montana male mating behavior under short-day conditions that induce diapause in females and found the males to be reproductively inactive. We also found that males reared under long-day conditions (reproducing individuals) court reproducing postdiapause females, but not diapausing ones. The diapausing flies of both sexes had more long-chain and less short-chain hydrocarbons on their cuticle than the reproducing ones, which presumably increase their survival under stressful conditions, but at the same time decrease their attractiveness. Our study shows that the mating behavior of females and males is well coordinated during and after overwintering and it also gives support to the dual role of insect cuticular hydrocarbons in adaptation and mate choice.

A costly chemical trait: phenotypic condition dependence of cuticular hydrocarbons in a dung beetle.

Chemical traits are increasingly recognized as important cues used in mate choice. For example, the cuticular hydrocarbons (CHCs) of insects have been shown to influence mating success in a range of taxa. Less is known, however, about how CHCs are expressed in proportion to an individual's condition, and consequently whether CHCs can function as condition-dependent signals of quality. We investigated this question using the dung beetle, Onthophagus taurus. CHCs are subject to sexual selection in this species through mate choice. A dietary manipulation revealed condition dependence of CHC expression for both sexes: dietary restriction decreased overall CHC production and altered the composition of CHCs. Furthermore, CHC production was associated with a measure of condition in beetles fed a limited diet but not those fed adlibitum. These results implicate a resource cost to CHC production that is likely to result in trade-offs with other fitness components in this species, as these respond similarly to a dietary restriction. The CHC profiles showed sexual dimorphism: males produced more CHCs and the sexes differed in the blend of compounds they produced. There was evidence for a male dimorphism in the CHC profile, in line with the presence of the alternative reproductive tactics (minor sneaks and major fighters) in this species. However, rather than mimicking a female CHC profile, minor males differed more from females than major males. Our results suggest that CHCs are a costly trait in O.taurus that has the potential to act as a condition-dependent signal of quality.

Cuticular hydrocarbon chemistry, an important factor shaping the current distribution pattern of the imported fire ants in the USA

Two sibling species, Solenopsis richteri and S. invicta, were both introduced into the southern USA from South America in the early 20th century. Today, S. richteri occupies higher latitudes and colder areas, while S. invicta occupies lower latitudes. Between the distributions of the two species, there is a large area of viable hybrid (S. richteri × S. invicta) populations. This study aimed to characterize the forces driving this distribution pattern and the underlying mechanisms. Cuticular hydrocarbons (CHCs) of freshly killed workers of S. invicta, hybrids, and S. richteri were removed using hexane. Both intact and CHCs-extracted workers were subjected to a constant rate of increasing temperature from 10 to 60 °C to obtain relative water loss and the water loss transition temperature (Tc-ant). Mass loss and Tc-ant were both significantly increased with CHCs removal. We then examined the CHC composition of three species. CHC profiles of S. richteri are characterized by significant amounts of short-chain (C23–C27) saturated and unsaturated hydrocarbons. In contrast, profiles of S. invicta consist primarily of long-chain (C27–C29) saturated hydrocarbons; unsaturated alkenes are completely lacking. Hybrid fire ants show intermediate profiles of the two parent species. We measured the melting point (Tm) and water-loss transition temperature of CHC blends (Tc-CHC) of different ant species colonies using differential scanning calorimetry (DSC) and an artificial membrane system, respectively. There were 3–5 Tms of each CHCs sample of different ant colonies due to their complex chemistry. The highest Tms (Tm-maxs) of CHCs samples from S. invicta and the hybrid were significantly higher than that from S. richteri. The correlation between Tc-CHC and Tm-max obtained from the same CHCs sample was highly significant. These results reveal that species having higher Tc and Tm-max retain more water under relatively higher temperature, and consequently are able to occupy warmer environments. We conclude that CHC chemistry plays a role in shaping current distribution patterns of S. richteri, S. invicta and their hybrid in the United States.

New chemical data on the ant Myrmecina graminicola (Formicidae, Myrmicinae): Unusual abundance of alkene hydrocarbons and esters

We investigated the possible origin of the specific odor of the terricolous ant Myrmecina graminicola by elucidating its pattern of volatiles. It appeared to have a very large hypertrophied poison gland reservoir, which contains copious amounts of acetate and propionate esters that are also present on the cuticule. The poison gland was previously reported as the source of sex pheromone, but the finding that queens and workers exhibited the same ester patterns tends to refute their role in sex attractants, and their biological significance remains elusive. The composition of cuticular hydrocarbons in this species is also highly original as it mostly comprises alkadienes and alkatrienes (70%), which is unusual and may be an adaptation to subterranean life. Neither the esters nor the cuticular hydrocarbons vary qualitatively and quantitatively between various localities in France, including Corsica.

Chemical and behavioral integration of army ant-associated rove beetles \u2013 a comparison between specialists and generalists

Host-symbiont interactions are embedded in ecological communities and range from unspecific to highly specific relationships. Army ants and their arthropod guests represent a fascinating example of species-rich host-symbiont associations where host specificity ranges across the entire generalist - specialist continuum. In the present study, we compared the behavioral and chemical integration mechanisms of two extremes of the generalist - specialist continuum: generalist ant-predators in the genus Tetradonia (Staphylinidae: Aleocharinae: Athetini), and specialist ant-mimics in the genera Ecitomorpha and Ecitophya (Staphylinidae: Aleocharinae: Ecitocharini). Similar to a previous study of Tetradonia beetles, we combined DNA barcoding with morphological studies to define species boundaries in ant-mimicking beetles. This approach found four ant-mimicking species at our study site at La Selva Biological Station in Costa Rica. Community sampling of Eciton army ant parasites revealed that ant-mimicking beetles were perfect host specialists, each beetle species being associated with a single Eciton species. These specialists were seamlessly integrated into the host colony, while generalists avoided physical contact to host ants in behavioral assays. Analysis of the ants’ nestmate recognition cues, i.e. cuticular hydrocarbons (CHCs), showed close similarity in CHC composition and CHC concentration between specialists and Eciton burchellii foreli host ants. On the contrary, the chemical profiles of generalists matched host profiles less well, indicating that high accuracy in chemical host resemblance is only accomplished by socially integrated species. Considering the interplay between behavior, morphology, and cuticular chemistry, specialists but not generalists have cracked the ants’ social code with respect to various sensory modalities. Our results support the long-standing idea that the evolution of host-specialization in parasites is a trade-off between the range of potential host species and the level of specialization on any particular host.

Distinct physiological, biochemical and morphometric adjustments in the malaria vectors Anopheles gambiae and A. coluzzii as means to survive dry season conditions in Burkina Faso.

Aestivation and dispersive migration are the two strategies evoked in the literature to explain the way in which malaria vectors Anopheles coluzzii and A. gambiae survive the harsh climatic conditions of the dry season in sub-Saharan Africa. However, the physiological mechanisms regulating these two strategies are unknown. In the present study, mosquito species were exposed to controlled environmental conditions mimicking the rainy and dry seasons of south western Burkina Faso. Survival strategies were studied through morphometric (wing length), ecophysiological (respiratory gas exchanges), biochemical (cuticular hydrocarbons composition) and molecular (AKH mRNA expression levels) parameters, variations of which are usually considered to be hallmarks of aestivation and dispersion mechanisms in various insects. Our results showed that ecophysiological and morphometric adjustments are made in both species to prevent water losses during the dry season. However, the usual metabolic rate modifications expected as signatures of aestivation and migration were not observed, highlighting specific and original physiological mechanisms sustaining survival in malaria mosquitoes during the dry season. Differences in epicuticular hydrocarbon composition and AKH levels of expression were found between the permanent and temporary A. coluzzii populations, illustrating the great phenotypic plasticity of this mosquito species. Altogether, our work underlines the diverse and complex pattern of changes occurring in the two mosquito species and at the population level to cope with the dry season and highlights potential targets of future control tools.

Coping with the climate: cuticular hydrocarbon acclimation of ants under constant and fluctuating conditions.

Terrestrial arthropods achieve waterproofing by a layer of cuticular hydrocarbons (CHCs). At the same time, CHCs also serve as communication signals. To maintain waterproofing under different climate conditions, insects adjust the chemical composition of their CHC layer, but this may affect the communication via CHCs. The detailed acclimatory changes of CHCs and how these influence their physical properties are still unknown. Here, we studied acclimation in two closely related ant species with distinct CHC profiles, Myrmica rubra and Myrmica ruginodis, in response to constant or fluctuating temperature and humidity regimes. We measured how acclimation affected CHC composition and viscosity, and the ants' drought survival. In both species, CHC composition showed strong, predictable responses to temperature regimes. Warm-acclimated individuals had higher proportions of linear alkanes, and less methyl-branched or unsaturated CHCs. These changes coincided with higher solid content and viscosity of CHCs in warm-acclimated ants. Temperature fluctuation caused effects similar to those observed under constant-cool conditions in Mrubra, but led to entirely different profiles in Mruginodis, suggesting that fluctuating and constant conditions pose very different challenges. Acclimation to dry conditions led to higher absolute amounts of CHCs, which increased the ants' drought survival, whereas temperature acclimation did not. Hence, the temperature-induced CHC changes cannot be explained by the need for waterproofing alone. Although these changes could be non-adaptive, we propose that they serve to maintain a constant CHC viscosity, which may be essential for communication and other functions.

Diferencias en el potencial evolutivo de las hormigas hospedadoras del género Proformica y sus parásitas esclavistas Rossomyrmex (Hymenoptera: Formicidae)

Ruano, F., Sanllorente, O., Lenoir, A., Tinaut, A. 2017. Different evolutionary potential in host ants of the genus Proformica and their slave-making ant Rossomyrmex (Hymenoptera: Formicidae). Ecosistemas 26(3): 48-53. Doi.: 10.7818/ECOS.2017.26-3.07 Parasitism is known to be one of the main drivers of evolution. Each organism involved in this relationship has an evolutionary potential that can be measured as its genetic diversity. This feature, in turn, depends on factors such as the dispersal ability, genetic flow and size, structure and composition of its populations. The evolutionary potential also constrains local adaptation between parasite and host. In slave-making ants, this local adaptation can be measured as the similitude in the cuticular hydrocarbon (CHCs) composition. Ants of the genus Rossomyrmex parasitize by raiding and enslaving ants from the genus Proformica. In this study we report new genetic data on two of the three host species studied by us, which together with previous published data enable us to analyze the genetic and biological characteristics of the different host-parasite systems. These systems show variable ranges of distribution (fragmented and continuous), genetic diversity, evolutionary potential and local adaptation. The geographic distribution (fragmented or continuous) strongly influences the population structure of Proformica in comparison with nest composition. We also found differences in the evolutionary potential when comparing host and parasite species, being local adaptation different for each host-parasite system. All these results suggest that the ants of both genera constitute interesting model systems for the comparative study of the processes leading to different evolutionary outcomes in the host-parasite arms-race.

How ants acclimate: Impact of climatic conditions on the cuticular hydrocarbon profile

Abstract Organisms from temperate zones are exposed to seasonal changes and must be able to cope with a wide range of climatic conditions. Especially ectotherms, including insects, are at risk to desiccate under dry and warm conditions, the more so given the changing climate. To adjust to current conditions, organisms acclimate through changes in physiology, morphology and/or behaviour. Insects protect themselves against desiccation through a layer of cuticular hydrocarbons (CHC) on their body surface. Hence, acclimation may also affect the CHC profile, changing their waterproofing capacity under different climatic conditions. Here, we investigated the acclimation response of two Temnothorax ant species to different climatic conditions. We analysed CHC profiles of queens, nurses and foragers that were acclimated to different humidity × temperature regimes, and tested the beneficial acclimation hypothesis by measuring survival of workers under desiccation‐heat stress. Both ants possessed a species‐specific CHC profile. Nevertheless, they showed similar acclimation responses concerning changes of certain CHC classes, and finally similar survival rates under desiccation‐heat stress. Warm‐acclimated individuals generally showed longer n‐alkanes, fewer dimethyl alkanes, and more (workers) or less (queens) monomethyl alkanes. In contrast, dry conditions resulted in more n‐alkanes and fewer mono‐ and dimethyl alkanes, but these acclimatory changes were only observed in workers and not in queens. Warm‐ and dry‐acclimated workers survived desiccation‐heat stress better, but we found no species differences. Our results indicate that both ant species can plastically adjust their cuticular hydrocarbon profile, allowing them to acclimate to different climatic conditions. Although their CHC composition differs in a species‐specific manner, they showed similar chemical adjustments and concomitant changes in survival rate. Hence, chemical plasticity may be critical to determine a species’ climatic range and its survival under changing climatic conditions. A plain language summary is available for this article.

Evolution of queen cuticular hydrocarbons and worker reproduction in stingless bees

Social insect queen pheromones can be interpreted as the queen’s means of sterilizing her workers, or as an honest signal of queen presence that benefits both parties. Co-mapping worker reproductive behaviour and queen cuticular hydrocarbon (CH) composition and quantity on a phylogeny of 21 stingless bee species showed that there are no associations between these traits. Furthermore, three species that have independently evolved facultative worker sterility are unexceptional in their queen CH. Combined, our analysis suggests that the action of stingless bee queen CHs are best interpreted as a signal of queen presence and not as a chemical contraceptive. Co-mapping the incidence of worker reproduction and queen pheromone chemical composition on a phylogeny of stingless bees reveals no association between queen hydrocarbon profiles and worker reproduction.

The evolution of a complex trait: cuticular hydrocarbons in ants evolve independent from phylogenetic constraints

Cuticular hydrocarbons (CHCs) are ubiquitous and highly diverse in insects, serving as communication signal and waterproofing agent. Despite their vital function, the causes, mechanisms and constraints on CHC diversification are still poorly understood. Here, we investigated phylogenetic constraints on the evolution of CHC profiles, using a global data set of the species-rich and chemically diverse ant genus Crematogaster. We decomposed CHC profiles into quantitative (relative abundances, chain length) and qualitative traits (presence/absence of CHC classes). A species-level phylogeny was estimated using newly generated and previously published sequences from five nuclear markers. Moreover, we reconstructed a phylogeny for the chemically diverse Crematogaster levior species group using cytochrome oxidase I. Phylogenetic signal was measured for these traits on genus and clade level and within the chemically diverse C.levior group. For most quantitative CHC traits, phylogenetic signal was low and did not differ from random expectation. This was true on the level of genus, clade and species group, indicating that CHC traits are evolutionary labile. In contrast, the presence or absence of alkenes and alkadienes was highly conserved within the C.levior group. Hence, the presence or absence of biosynthetic pathways may be phylogenetically constrained, especially at lower taxonomic levels. Our study shows that CHC composition can evolve rapidly, allowing insects to quickly adapt their chemical profiles to external selection pressures, whereas the presence of biosynthetic pathways appears more constrained. However, our results stress the importance to consider the taxonomic level when investigating phylogenetic constraints.

The relationship between epicuticular long-chained hydrocarbons and surface area - volume ratios in insects (Diptera, Hymenoptera, Lepidoptera).

Long-chain cuticular hydrocarbons (CHCs) are common components of the epicuticle of terrestrial arthropods. CHC serve as a protective barrier against environmental influences but also act as semiochemicals in animal communication. Regarding the latter aspect, species- or intra-functional group specific CHCs composition and variation are relatively well studied. However, comparative knowledge about the relationship of CHC quantity and their relation to surface area—volume ratios in the context of water loss and protection is fragmentary. Hence, we aim to study the taxon-specific relationship of the CHC amount and surface-area to volume ratio related to their functional role (e.g. in water loss). We focused on flower visiting insects and analyzed the CHC amounts of three insect orders (Hymenoptera, Lepidoptera and Diptera) using gas chromatography—mass spectrometry (GC-MS). We included 113 species from two grassland plots, quantified their CHCs, and measured their body mass and surface area. We found differences in the surface area, CHCs per body mass and the CHC density (= amount of CHCs per surface area) across the three insect taxa. Especially the Hymenoptera had a higher CHC density compared to Diptera and Lepidoptera. CHC density could be explained by surface area-volume ratios in Hymenoptera but not in Diptera and Lepidoptera. Unexpectedly, CHC density decreased with increasing surface area—volume ratios.

How do cuticular hydrocarbons evolve? Physiological constraints and climatic and biotic selection pressures act on a complex functional trait.

Cuticular hydrocarbons (CHCs) cover the cuticles of virtually all insects, serving as a waterproofing agent and as a communication signal. The causes for the high CHC variation between species, and the factors influencing CHC profiles, are scarcely understood. Here, we compare CHC profiles of ant species from seven biogeographic regions, searching for physiological constraints and for climatic and biotic selection pressures. Molecule length constrained CHC composition: long-chain profiles contained fewer linear alkanes, but more hydrocarbons with disruptive features in the molecule. This is probably owing to selection on the physiology to build a semi-fluid cuticular layer, which is necessary for waterproofing and communication. CHC composition also depended on the precipitation in the ants' habitats. Species from wet climates had more alkenes and fewer dimethyl alkanes than those from drier habitats, which can be explained by different waterproofing capacities of these compounds. By contrast, temperature did not affect CHC composition. Mutualistically associated (parabiotic) species possessed profiles highly distinct from non-associated species. Our study is, to our knowledge, the first to show systematic impacts of physiological, climatic and biotic factors on quantitative CHC composition across a global, multi-species dataset. We demonstrate how they jointly shape CHC profiles, and advance our understanding of the evolution of this complex functional trait in insects.

Beyond Cuticular Hydrocarbons: Chemically Mediated Mate Recognition in the Subsocial Burying Beetle Nicrophorus vespilloides.

Burying beetles have fascinated scientists for centuries due to their elaborate form of biparental care that includes the burial and defense of a vertebrate carcass, as well as the subsequent feeding of the larvae. However, besides extensive research on burying beetles, one fundamental question has yet to be answered: what cues do males use to discriminate between the sexes? Here, we show in the burying beetle Nicrophorus vespilloides that cuticular lipids trigger male mating behavior. Previous chemical analyses have revealed sex differences in cuticular hydrocarbon (CHC) composition; however, in the current study, fractionated-guided bioassay showed that cuticular lipids, other than CHCs, elicit copulation. Chemical analyses of the behaviorally active fraction revealed 17 compounds, mainly aldehydes and fatty acid esters, with small quantitative but no qualitative differences between the sexes. Supplementation of males with hexadecanal, the compound contributing most to the statistical separation of the chemical profiles of males and females, did not trigger copulation attempts by males. Therefore, a possible explanation is that the whole profile of polar lipids mediates sex recognition in N. vespilloides.

Cuticular Lipids as a Cross-Talk among Ants, Plants and Butterflies.

Even though insects and plants are distantly related organisms, they developed an integument which is functionally and structurally similar. Besides functioning as a physical barrier to cope with abiotic and biotic stress, this interface, called cuticle, is also a source of chemical signaling. Crucial compounds with this respect are surface lipids and especially cuticular hydrocarbons (CHCs). This review is focused on the role of CHCs in fostering multilevel relationships among ants, plants and Lepidoptera (primarily butterflies). Indeed, particular traits of ants as eusocial organisms allowed the evolution and the maintenance of a variety of associations with both plants and animals. Basic concepts of myrmecophilous interactions and chemical deception strategies together with chemical composition, biosynthetic pathways and functions of CHCs as molecular cues of multitrophic systems are provided. Finally, the need to adopt a multidisciplinary and comprehensive approach in the survey of complex models is discussed.