Chemical Signature and Reproductive Status in the Facultatively Polygynous ant Pachycondyla Verenae

In insects, cuticular hydrocarbons (CHCs) generally are used as cues and signals for within colony processes, such as signaling reproductive status, and between colony processes, such as colony membership. We examined CHC profiles of the facultatively polygynous ant Pachycondyla verenae in order to identify chemical signals of reproductive queens within colonies containing many gynes. Colonies of P. verenae, belonging to two different members of a complex of morphospecies, were collected from three geographic localities within South America. We also tested whether CHC profiles differed between geographic localities and morphospecies. We found three alkenes, two isomers of pentacosene and heptacosene, which were more abundant in CHC profiles of reproductive queens of this morphospecies complex. When we tested whether these differences were consistent across geographic localities, we found the abundance of these alkenes differed according to morphospecies, with the isomers of pentacosene being more abundant in queens from morph one, and heptacosene being more abundant in queens from morph two. Our study has given further insight into the mechanisms behind maintenance of reproductive dominance, and has demonstrated that chemical signatures associated with reproductive status in Pachycondyla verenae are not conserved within this species complex.     

Conservation of Queen Pheromones Across Two Species of Vespine Wasps

Social insects are known for their reproductive division of labor between queens and workers, whereby queens lay the majority of the colony’s eggs, and workers engage mostly in non-reproductive tasks. Queens produce pheromones that signal their presence and fertility to workers, which in turn generally remain sterile. Recently, it has been discovered that specific queen-characteristic cuticular hydrocarbons (CHCs) function as queen pheromones across multiple lineages of social insects. In the common wasp, Vespula vulgaris, several long-chain linear alkanes and 3-methylalkanes were shown to act as queen signals. Here, we describe similar bioassays with a related species of highly eusocial vespine wasp, the Saxon wasp, Dolichovespula saxonica. We show that a blend of queen-characteristic hydrocarbons that are structurally related to those of the common wasp inhibit worker reproduction, suggesting conservation of queen pheromones across social wasps. Overall, our results highlight the central importance of CHCs in chemical communication among social insects in general, and as conserved queen pheromones in these social wasps in particular.     

Functional subcaste discrimination (foragers and brood-tenders) in the antCamponotus vagus scop.: polymorphism of cuticular hydrocarbon patterns

In the antCamponotus vagus, when selected foragers that had been earlier removed from the foraging arena and brood-tenders that had been earlier removed from the nest were placed together in a foraging arena, most of the brood-tenders and only a few of the selected foragers were carried back to the nest by nonselected foragers. We hypothesize that cuticular hydrocarbons serve as a cue that allows foragers to discriminate between members of their own subcaste and brood-tenders. It has been established that the proportions of certain hydrocarbons, which are the same regardless of the colony studied, vary from one worker subcaste to another and thus constitute a specific chemical signature. These hydrocarbons belong to a wide range of chemical families (alkanes, monomethylalkanes, and dimethylalkanes). The greatest differences between the two subcastes were observed on the thorax of workers. Principal component analyses performed on the hydrocarbons (or hydrocarbon combinations) corresponding to the 45 main peaks in the cuticular profiles of the head and thorax of brood-tenders and foragers of several colonies show that there exist quantitative differences between the various signatures that characterize the colony, the worker subcastes, and the various body parts within the same species, which can be classified in a hierarchy where the differences between worker subcastes are less pronounced than those between body parts or between colonies.     

Nestmate Recognition and the Role of Cuticular Hydrocarbons in the African Termite Raiding Ant <Emphasis Type="Italic">Pachycondyla analis</Emphasis>

Cuticular hydrocarbons (CHCs) are used for chemical communication among nestmates in many ant species, and they may play a role in the discrimination of nestmates and non-nestmates. Using the mandible opening response (MOR) bioassay, we tested the response of the African termite raiding ant, Pachycondyla analis, to CHC extracts of nestmates and non-nestmates. The ants were able to distinguish control chemical cues, from nestmate CHCs, and from non-nestmate CHCs, and, based on a CHC recognition threshold, aggression was demonstrated toward non-nestmates. Gas chromatography (GC) and GC-mass spectrometric analyses showed that CHC components of different ant colonies had chain lengths ranging from C₈ to C₃₁, comprising mainly n-alkanes, alkenes, and methyl branched alkanes, with the n-alkanes occurring in the same proportions among all colonies. The ants were grouped successfully according to their colonies of origin by using discriminant analysis of CHCs. We demonstrate that nestmate recognition occurs in P. analis, and that some of the cues involved are evidently alkenes and methyl-branched alkanes.     

Hydrocarbons in the Ant <Emphasis Type="Italic">Lasius niger</Emphasis>: From the Cuticle to the Nest and Home Range Marking

The cuticular hydrocarbons (CHCs) of the ant Lasius niger are described. We observe a high local colony specificity of the body cuticular profile as predicted for a monogynous and multicolonial species. The CHCs show a low geographical variation among different locations in France. The CHCs on the legs also are colony specific, but their relative quantities are slightly different from those on the main body. For the first time, we demonstrate that the inner walls of the ant nest are coated with the same hydrocarbons as those found on the cuticle but in different proportions. The high amount of inner-nest marking and its lack of colony-specificity may explain why alien ants are not rejected once they succeed in entering the nest. The cuticular hydrocarbons also are deposited in front of the nest entrance and on the foraging arena, with a progressive increase in n-alkanes relative amounts. Chemical marks laid over the substrate are colony specific only when we consider methyl-branched alkanes. Our data confirm that these “footprint hydrocarbons” are probably deposited passively by the contact of ant tarsae with the substrate. These results suggest that the CHCs chemical profiles used by ants in colony recognition are much more complex than a single template: ants have to learn and memorize odors that vary depending on their context of perception.     

Variations in Worker Cuticular Hydrocarbons and Soldier Isoprenoid Defensive Secretions Within and Among Introduced and Native Populations of the Subterranean Termite, Reticulitermes flavipes

In social insects, cuticular hydrocarbons (CHCs) play a central role in nestmate recognition. CHCs have proved to be useful for identifying species and differentiating populations. In combination with CHCs, isoprenoid soldier defensive secretions (SDSs) have been previously used in some termite species for chemotaxonomic analyses. This study compared the levels of chemical variation within and among introduced (French) and native (U.S.) populations of the subterranean termite, Reticulitermes flavipes. Worker CHCs and soldier SDSs from termites collected from colonies in nine populations in Florida, Louisiana, and France were analyzed. Discriminant analyses revealed that both localities and populations can be distinguished by using the variation in CHC profiles. Principal component analyses of CHC profiles as well as the calculation of two distance parameters (Nei and Euclidean) revealed remarkable chemical homogeneity within and among French populations. These analyses also showed that the CHC profiles of French populations were closer to termite populations from Louisiana than to those from Florida. Of the six distinct SDS chemotypes, one was common to populations in France and Louisiana. The possibility that populations in France originated from Louisiana, and the potential causes and consequences of chemical homogeneity within introduced populations are discussed.     

Epicuticular Compounds of Drosophila subquinaria and D. recens: Identification, Quantification, and Their Role in Female Mate Choice

The epicuticle of various Drosophila species consists of long-chain cuticular hydrocarbons (CHCs) and their derivatives that play a role in waterproofing and a dynamic means of chemical communication. Here, via gas chromatography and mass spectrometry, we identified and quantified the epicuticular composition of D. recens and D. subquinaria, two closely related species that show a pattern of reproductive character displacement in nature. Twenty-four compounds were identified with the most abundant, 11-cis-Vaccenyl acetate, present only in males of each species. Also exclusive to males were five tri-acylglycerides. The 18 remaining compounds were CHCs, all shared between the sexes and species. These CHCs were composed of odd carbon numbers (C₂₉, C₃₁, C_33, and C₃₅), with an increase in structural isomers in the C₃₃ and C₃₅ groups. Saturated hydrocarbons comprise only methyl-branched alkanes and were found only in the C₂₉ and C₃₁ groups. Alkenes were the least prevalent, with alkadienes dominating the chromatographic landscape in the longer chain lengths. Sexual dimorphism was extensive with 6/8 of the logcontrast CHCs differing significantly in relative concentration between males and females in D. recens and D. subquinaria, respectively. Males of the two species also differed significantly in relative concentration of six CHCs, while females differed in none. Female-choice mating trials revealed directional sexual selection on male CHCs in a population of each species, consistent with female mate preferences for these traits. The sexual selection vectors differed significantly in multivariate trait space, suggesting that different pheromone blends determine male attractiveness in each species.     

Chemical Characterization of Young Virgin Queens and Mated Egg-Laying Queens in the Ant <Emphasis Type="Italic">Cataglyphis cursor</Emphasis>: Random Forest Classification Analysis for Multivariate Datasets

Social insects are well known for their extremely rich chemical communication, yet their sex pheromones remain poorly studied. In the thermophilic and thelytokous ant, Cataglyphis cursor, we analyzed the cuticular hydrocarbon profiles and Dufour’s gland contents of queens of different age and reproductive status (sexually immature gynes, sexually mature gynes, mated and egg-laying queens) and of workers. Random forest classification analyses showed that the four groups of individuals were well separated for both chemical sources, except mature gynes that clustered with queens for cuticular hydrocarbons and with immature gynes for Dufour’s gland secretions. Analyses carried out with two groups of females only allowed identification of candidate chemicals for queen signal and for sexual attractant. In particular, gynes produced more undecane in the Dufour’s gland. This chemical is both the sex pheromone and the alarm pheromone of the ant Formica lugubris. It may therefore act as sex pheromone in C. cursor, and/or be involved in the restoration of monogyny that occurs rapidly following colony fission. Indeed, new colonies often start with several gynes and all but one are rapidly culled by workers, and this process likely involves chemical signals between gynes and workers. These findings open novel opportunities for experimental studies of inclusive mate choice and queen choice in C. cursor.     

Chemical Fertility Signaling in Termites: Idiosyncrasies and Commonalities in Comparison with Ants

Termites evolved eusociality independently from social Hymenoptera. As a common trait, reproductive monopoly is maintained through chemical communication. The queen (and in termites also a king) prevents workers from reproduction by conveying their reproductive status. In termites all soldiers are sterile, but workers’ potential to reproduce differs between species. It ranges from totipotency in wood-dwelling lower termites where workers are a transient stage from which all other castes develop, to sterile workers in some higher termites. Intermediate are species in which workers can develop into replacement sexuals within the nest but not into winged sexuals. I summarize the patchy picture about fertility signaling that we currently have for termites, pointing also to potential conflicts over reproduction that differ from those in social Hymenoptera. Recent findings imply that, similar to many social Hymenoptera, wood-dwelling termites that live in confined nests use long-chain cuticular hydrocarbons (CHCs) as fertility signals. Yet other compounds are important as well, comprising proteinaceous secretions and especially volatiles. For a subterranean termite, two volatiles have been identified as primer pheromones that prevent reproductive differentiation of workers. It requires more data to test whether wood-dwelling termites use CHCs, while species with larger colonies and less confined nests use volatiles, or whether all species rely on multicomponent signals. Ultimately, we need more effort to model and test potential conflicts over reproduction between queens, kings and workers. Here results from social Hymenoptera cannot be transferred to termites as the latter are diploid and commonly inbred. This review illustrates promising future research avenues.     

Do Aphid Carcasses on the Backs of Larvae of Green Lacewing Work as Chemical Mimicry against Aphid-Tending Ants?

Ants attack and exclude natural enemies of aphids in ant–aphid mutualisms. However, larvae of the green lacewing, Mallada desjardinsi, prey on the cowpea aphid, Aphis craccivora, without exclusion by aphid-tending ants. Lacewing larvae are protected from ants by carrying aphid carcasses on their backs. Here, we tested whether cuticular hydrocarbons (CHCs) of aphid carcasses affected the aggressiveness of aphid-tending ants. Aphid carcasses were washed with n-hexane to remove lipids. Lacewing larvae with washed aphid carcasses were attacked by aphid-tending ants more frequently than those with untreated aphid carcasses. We measured the aggressiveness of aphid-tending ants to lacewing larvae that were either carrying a piece of cotton wool (a dummy aphid carcass) treated with CHCs from aphids or lacewing larvae, or carrying aphid carcasses. The rates of attack by ants on lacewing larvae carrying CHCs of aphids or aphid carcasses were lower than that of attack on lacewing larvae with conspecific CHCs. Chemical analysis by gas chromatography/mass spectrometry showed similarity of CHCs between aphids and aphid carcasses. These results suggest that aphid carcasses on the backs of lacewing larvae function via chemical camouflage to limit attacks by aphid-tending ants.     

Venom Alkaloid and Cuticular Hydrocarbon Profiles Are Associated with Social Organization,Queen Fertility Status,and Queen Genotype in the Fire Ant <Emphasis Type="Italic">Solenopsis invicta</Emphasis>

Queens in social insect colonies advertise their presence in the colony to: a) attract workers’ attention and care; b) gain acceptance by workers as replacement or supplemental reproductives; c) prevent reproductive development in nestmates. We analyzed the chemical content of whole body surface extracts of adult queens of different developmental and reproductive stages, and of adult workers from monogyne (single colony queen) and polygyne (multiple colony queens) forms of the fire ant Solenopsis invicta. We found that the composition of the most abundant components, venom alkaloids, differed between queens and workers, as well as between reproductive and non-reproductive queens. Additionally, workers of the two forms could be distinguished by alkaloid composition. Finally, sexually mature, non-reproductive queens from polygyne colonies differed in their proportions of cis-piperidine alkaloids, depending on their Gp-9 genotype, although the difference disappeared once they became functional reproductives. Among the unsaturated cuticular hydrocarbons characteristic of queens, there were differences in amounts of alkenes/alkadienes between non-reproductive polygyne queens of different Gp-9 genotypes, between non-reproductive and reproductive queens, and between polygyne and monogyne reproductive queens, with the amounts increasing at a relatively higher rate through reproductive ontogeny in queens bearing the Gp-9 b allele. Given that the genotype-specific piperidine differences reflect differences in rates of reproductive maturation between queens, we speculate that these abundant and unique compounds have been co-opted to serve in fertility signaling, while the cuticular hydrocarbons now play a complementary role in regulation of social organization by signaling queen Gp-9 genotype.     

The Social Integration of a Myrmecophilous Spider Does Not Depend Exclusively on Chemical Mimicry

Numerous animals have evolved effective mechanisms to integrate into and exploit ant societies. Chemical integration strategies are particularly widespread among ant symbionts (myrmecophiles), probably because social insect nestmate recognition is predominantly mediated by cuticular hydrocarbons (CHCs). The importance of an accurate chemical mimicry of host CHCs for social acceptance recently has been demonstrated in a myrmecophilous silverfish. In the present study, we investigated the role of chemical mimicry in the myrmecophilous spider Gamasomorpha maschwitzi that co-occurs in the same host, Leptogenys distinguenda, as the silverfish. To test whether spiders acquire mimetic CHCs from their host or not, we transferred a stable isotope-labeled hydrocarbon to the cuticle of workers and analyzed the adoption of this label by the spiders. We also isolated spiders from hosts in order to study whether this affects: 1) their chemical host resemblance, and 2) their social integration. If spiders acquired host CHCs, rather than biosynthesizing them, they would be expected to lose these compounds during isolation. Spiders acquired the labeled CHC from their host, suggesting that they also acquire mimetic CHCs, most likely through physical contact. Furthermore, isolated spiders lost considerable quantities of their CHCs, indicating that they do not biosynthesize them. However, spiders remained socially well integrated despite significantly reduced chemical host similarity. We conclude that G. maschwitzi depends less on chemical mimicry to avoid recognition and aggressive rejection than the silverfish previously studied, suggesting that the two myrmecophiles possess different adaptations to achieve social integration.     

Cuticular Chemistry of Males and Females in the Ant Formica fusca

Communication between organisms involves visual, auditory, and olfactory pathways. In solitary insects, chemical recognition cues are influenced mainly by selection regimes related to species recognition and sexual selection. In social insects, chemical recognition cues have an additional role in mediating recognition of society members and, thereby, allowing kin selection to operate. Here, we examined whether cuticular hydrocarbon profiles are sex-specific and whether males and young queens of the ant Formica fusca have colony-specific profiles. We also investigated whether there is a relationship between genetic relatedness and chemical diversity within colonies. We demonstrated that female and male sexuals do not have unique sex-specific compounds, but that there are quantitative chemical differences between the sexes. Out of the 51 cuticular hydrocarbon compounds identified, 10 showed a significant quantitative difference between males and females. We also showed that both males and females have a significant colony-specific component in their profiles. Finally, we found a negative correlation between within-colony relatedness and within-colony chemical diversity of branched, but not linear compounds. This suggests that colonies with multiple matri- or patrilines also have a significantly greater chemical diversity.     

Cuticular Hydrocarbons Provide Reliable Cues of Fertility in the Ant Gnamptogenys striatula

In ca. 150 species of queenless ants, a specialized queen caste is rare or absent, and mated workers take over the role of the queen in some or all of the colonies. Previously, it has been shown that reproduction in queenless ants is regulated by a combination of dominance behavior and chemical fertility signaling. It is unknown, however, whether chemical signals alone can sufficiently regulate reproduction. To investigate this possibility, we studied reproductive regulation in the facultatively queenless ant Gnamptogenys striatula, a species where dominance behavior is rare or absent. Active egg layers and infertile workers showed qualitative and quantitative differences in their cuticular hydrocarbon profile. Five long-chain methyl alkanes, 3,13- and 3,15-dimethyl pentriacontane, 3,13- and 3,15-dimethyl heptentriacontane, and 3,11,15-trimethyl heptentriacontane occurred only on the cuticles of virgin and mated egg layers. Pronounced quantitative differences were found in a further 27 alkenes; alkanes; and mono-, di-, and trimethyl alkanes. Workers that had recently stopped laying eggs had profiles similar to infertile workers, and mating status did not affect this chemical pattern. We conclude that the cuticular hydrocarbon profiles of G. striatula workers provide reliable information about their current fertility. In the interest of colony productivity, this allows reproduction to be regulated without the use of aggression.     

Cuticular Hydrocarbon Cues Are Used for Host Acceptance by <Emphasis Type="Italic">Pseudacteon</Emphasis> spp. Phorid Flies that Attack <Emphasis Type="Italic">Azteca sericeasur</Emphasis> Ants

Parasitoids often use complex cues to identify suitable hosts in their environment. Phorid fly parasitoids that develop on one or a few host species often use multiple cues, ranging from general to highly specific, to home in on an appropriate host. Here, we describe the hierarchy of cues that Pseudacteon phorid flies use to identify Azteca ant hosts. We show, through behavioral observations in the field, that phorid flies are attracted to two cryptic Azteca species, but only attack Azteca sericeasur (Hymenoptera: Formicidae: Dolichoderinae). To test whether the phorid flies use cuticular hydrocarbons (CHCs) to distinguish between the two Azteca taxa, we first documented and compared cuticular hydrocarbons of the two Azteca taxa using gas chromatography/mass spectrometry. Then, using cuticular hydrocarbon-transfer experiments with live ants, we characterized the cuticular hydrocarbons of A. sericeasur as a short-range, host location cue used by P. lasciniosus (Diptera: Phoridae) to locate the ants.     

Do Cuticular Hydrocarbons Provide Sufficient Information for Optimal Sex Allocation in the Ant <Emphasis Type="Italic">Formica exsecta</Emphasis>?

Split sex ratio theory predicts that when kin structure varies among colonies of social insects, in order to maximize the inclusive fitness, colonies with relatively high sister-sister relatedness should specialize in producing reproductive females, whereas in those with relatively low sister-sister relatedness workers should bias their sex ratio towards males. However, in order to achieve this, workers need to be able to reliably assess the type of colony in which they live. The information on colony kin structure may be encoded in cuticular hydrocarbons (CHCs), assuming that genetic variability translates accurately into chemical variability. However, in genetically heterogeneous colonies, too accurate information may encourage the pursuit of individual interests through nepotistic behavior and reduce colony efficiency or cause social disruption. In this study, we estimated how well variability of CHC recognition cues reflects colony kin structure in the ant Formica exsecta. Our results show that CHC variability does not covary with kin structure or the overall genetic diversity of the colony, and that patrilines and matrilines can have distinct CHC profiles in some but not all colonies. However, within-colony relatedness remains the key determinant of colony sex ratios. Based on our results, CHC variability cannot serve as accurate information on within-colony relatedness, kin structure, or full-sib affiliation, nor do workers seem to use colony CHC variability as a proxy for sex-ratio adjustment. The use of this type of information thus could lead workers to make mistakes, and it remains unclear how colonies of Formica exsecta adjust offspring sex ratio to their optimal value.     

Colony-Specific Cuticular Hydrocarbon Profile in Formica argentea Ants

The cuticular hydrocarbons of the ant Formica argentea were identified by gas chromatography/mass spectrometry. Behavioral bioassays tested the role of each class of cuticular hydrocarbon in nestmate recognition, and statistical analyses looked for potential colony-specific signatures. The cuticular hydrocarbons of F. argentea consist of n-alkanes, alkenes, and methyl-branched alkanes. Behavioral bioassays demonstrated that changes in the alkene and methyl-branched alkane signature of F. argentea increased aggression, but changes in alkanes did not. Statistical analyses demonstrated that F. argentea workers present a colony-specific hydrocarbon profile based on their methyl-branched C₂₉ alkane signature. Using this signature alone, it is possible to group worker ants statistically by nest, suggesting that methyl-branched C₂₉ alkanes may be important in nestmate recognition for this species. These results support the idea that variation in positional isomers of cuticular hydrocarbons of the same carbon chain length may provide enough information for nestmate recognition. Although the addition of alkenes increased aggression in F. argentea, alkenes did not provide a colony-specific signature. This study reinforces the idea that investigators studying nestmate recognition should not examine cuticular hydrocarbon profiles as a whole but rather, should look for colony-specific signatures embedded in parts of the profile.     

<Emphasis Type="Italic">Tetramorium tsushimae</Emphasis> Ants Use Methyl Branched Hydrocarbons of Aphids for Partner Recognition

In mutualisms, partner discrimination is often the most important challenge for interacting organisms. The interaction between ants and aphids is a model system for studying mutualisms; ants are provided with honeydew by aphids and, in turn, the ants offer beneficial services to the aphids. To establish and maintain this system, ants must discriminate mutualistic aphid species correctly. Although recent studies have shown that ants recognize aphids as mutualistic partners based on their cuticular hydrocarbons (CHCs), it was unclear which CHCs are involved in recognition. Here, we tested whether the n-alkane or methylalkane fraction, or both, of aphid CHCs were utilized as partner recognition cues by measuring ant aggressiveness toward these fractions. When workers of Tetramorium tsushimae ants were presented with dummies coated with n-alkanes of their mutualistic aphid Aphis craccivora, ants displayed higher levels of aggression than to dummies treated with total CHCs or methyl alkanes of A. craccivora; responses to dummies treated with n-alkanes of A. craccivora were similar to those to control dummies or dummies treated with the CHCs of the non-mutualistic aphid Acyrthosiphon pisum. By contrast, ants exhibited lower aggression to dummies treated with either total CHCs or the methylalkane fraction of the mutualistic aphid than to control dummies or dummies treated with CHCs of the non-mutualistic aphid. These results suggest that T. tsushimae ants use methylalkanes of the mutualistic aphid’s CHCs to recognize partners, and that these ants do not recognize aphids as partners on the basis of n-alkanes.     

Species-Specific Cuticular Hydrocarbon Stability within European <Emphasis Type="Italic">Myrmica</Emphasis> Ants

Recognition is a fundamental process on which all subsequent behaviors are based at every organizational level, from the gene up to the super-organism. At the whole organism level, visual recognition is the best understood. However, chemical communication is far more widespread than visual communication, but despite its importance is much less understood. Ants provide an excellent model system for chemical ecology studies as it is well established that compounds known as cuticular hydrocarbons (CHCs) are used as recognition cues in ants. Therefore, stable species-specific odors should exist, irrespective of geographic locality. We tested this hypothesis by comparing the CHC profiles of workers of twelve species of Myrmica ants from four countries across Europe, from Iberia to the Balkans and from the Mediterranean to Fennoscandia. CHCs remained qualitatively stable within each species, right down to the isomer level. Despite the morphological similarity that occurs within the genus Myrmica, their CHCs were highly diverse but remarkably species-specific and stable across wide geographical areas. This indicates a genetic mechanism under strong selection that produces these species-specific chemical profiles, despite each species encountering different environmental conditions across its range.     

Fine Tuning of Social Integration by Two Myrmecophiles of the Ponerine Army Ant, <Emphasis Type="Italic">Leptogenys distinguenda</Emphasis>

Myrmecophiles are animals that live in close association with ants and that frequently develop elaborate mechanisms to infiltrate their well-defended host societies. We compare the social integration strategies of two myrmecophilic species, the spider, Gamasomorpha maschwitzi, and the newly described silverfish, Malayatelura ponerophila gen. n. sp. n., into colonies of the ponerine army ant, Leptogenys distinguenda (Emery) (Hymenoptera: Formicidae). Both symbionts use chemical mimicry through adoption of host cuticular hydrocarbons. Exchange experiments between L. distinguenda and an undetermined Leptogenys species demonstrate that reduced aggression toward alien ants and increased social acceptance occurred with individuals of higher chemical similarity in their cuticular hydrocarbon profiles. We found striking differences in chemical and behavioral strategies between the two myrmecophiles. Spider cuticular hydrocarbon profiles were chemically less similar to the host than silverfish profiles were. Nevertheless, spiders received significantly fewer attacks from host ants and survived longer in laboratory colonies, whereas silverfish were treated with high aggression and were killed more frequently. When discovered and confronted by the host, silverfish tended to escape and were chased aggressively, whereas spiders remained in contact with the confronting host ant until aggression ceased. Thus, spiders relied less on chemical mimicry but were nevertheless accepted more frequently by the host on the basis of behavioral mechanisms. These findings give insights into the fine tuning of social integration mechanisms and show the significance of qualitative differences among strategies. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.