Discrimination of nestmate workers and drones in honeybees

Summary Discrimination of nestmates from non-nestmates has mainly been investigated in female social insects. Little is known about discrimination of males. Here we show that under natural conditions at the nest entrance, honeybee workers can discriminate nestmate drones from non-nestmate drones as effectively as they can discriminate nestmate workers from non-nestmate workers.     

Disentangling environmental and heritable nestmate recognition cues in a carpenter ant

Discriminating between group members and strangers is a key feature of social life. Nestmate recognition is very effective in social insects and is manifested by aggression and rejection of alien individuals, which are prohibited to enter the nest. Nestmate recognition is based on the quantitative variation in cuticular hydrocarbons, which can include heritable cues from the workers, as well as acquired cues from the environment or queen-derived cues. We tracked the profile of six colonies of the ant Camponotus aethiops for a year under homogeneous laboratory conditions. We performed chemical and behavioral analyses. We show that nestmate recognition was not impaired by constant environment, even though cuticular hydrocarbon profiles changed over time and were slightly converging among colonies. Linear hydrocarbons increased over time, especially in queenless colonies, but appeared to have weak diagnostic power between colonies. The presence of a queen had little influence on nestmate discrimination abilities. Our results suggest that heritable cues of workers are the dominant factor influencing nestmate discrimination in these carpenter ants and highlight the importance of colony kin structure for the evolution of eusociality.     

A sensitive and reliable assay for queen discrimination ability in laboratory-reared workers of the ant Camponotus japonicus

The queen discrimination abilities of laboratory-reared Camponotus japonicus workers were examined individually by allowing them to carry their nestmate larvae toward either the mother queen or an alien queen. Source colonies had been reared under controlled conditions from founding queens and maintained at small size (< or = 10 workers each). Fifty-two of fifty-four workers raised in these eight different colonies carried nestmate larvae to the mother queen, and never carried them to the alien queen. Most of them attended nestmate larvae but never alien larvae. These results clearly demonstrate that the tested workers discriminate the nestmate queen and larvae from non-nestmate conspecifics. The assay used in this study is novel and sensitive, and may be suitable for neuroethological and molecular studies of social discrimination mechanisms.     

Nestmate recognition in a neotropical polygynous wasp

In social insects, nestmate recognition systems can be dynamic and modulated in response to various kinds of genetic and environmental cues. For example, multiple-queen colonies can possess weak recognition abilities relative to single-queen colonies, due to broader exposure to heritable and environmentally derived nestmate recognition cues.We conducted field experiments to examine nestmate recognition ability in a neotropical polygynous wasp, Polybia paulista. Despite the fact that the effective queen number in P. paulista is the highest ever recorded in polygynous wasps, this species exhibits a well functioning nestmate recognition system, which allows colony entry only to nestmate individuals. Similar to other social Hymenoptera, young wasps express colony specific chemical signatures within several days after emergence. This is the first study to show that the polygynous epiponine wasp is able to distinguish nestmates from non-nestmates. Received 23 May 2006; revised 6 October 2006; accepted 23 October 2006.     

Nestmate recognition in the stingless bee Melipona panamica (Apidae, Meliponini)

Summary: Nestmate recognition was studied in the Neotropical stingless bee Melipona panamica, a species in which workers "sneak" their own reproductive eggs into 1 % of brood cells. We manipulated four factors that could influence individual recognition cues: the mother queen, the environment during the immature stage, the environment during the early adult stage, and worker age. We also simulated the action of natural enemies on colonies tested for discrimination of such worker characteristics. All factors that we tested affected responses of the discriminating workers, which could recognize sisters, nieces and unrelated workers. Previous exposure of unrelated callow bees to the odor of the host nest greatly increased chances of acceptance by the host colony. Probability of acceptance decreased, however, with increasing age of introduced bees or increasing disturbance of the host colony. These complexities in patterns of nestmate recognition and nest defense are adequately explained from the standpoint of inclusive fitness of the discriminating workers. Differences in nestmate recognition and worker egg laying among Meliponini are also discussed.     

Low intraspecific aggression level in the polydomous and facultative polygynous ant Ectatomma tuberculatum

Nestmate recognition is a key feature of social insects, as it preserves colony integrity. However, discrimination of non‐nestmates and nestmate recognition mechanisms are highly variable according to species and social systems. Here, we investigated the intraspecific level of aggression in the facultative polygynous and polydomous ant, Ectatomma tuberculatum Olivier (Hymenoptera: Formicidae: Ectatomminae), in a population with a strong genetic structure. We found that the intraspecific level of aggression was generally low in this population of E. tuberculatum. However, the level of aggression was significantly correlated with the geographical distance, suggesting that both genetic and environmental cues could be involved in nestmate recognition and discrimination mechanisms. Moreover, polydomy was confirmed by the absence of aggression between workers from nests at a distance of 3 m, while the level of aggression was significantly higher between workers from nests separated by a distance of 10 or 1300 m. Field experiments showed that the low level of aggression between neighbouring colonies was associated with shared foraging areas, which could suggest that familiarization processes may occur in this species. We propose that the particular social organization of this species, with secondary polygyny, polydomy, and budding, may have favoured a high acceptance threshold, because of the low probability of interactions with unrelated conspecifics, the high cost of erroneously rejecting nestmates, and the low cost of accepting non‐nestmate workers. The resulting open recognition system can thus allow privileged relationships between neighbouring colonies and promote the ecological dominance of E. tuberculatum in the mosaic of arboreal ants.     

Colony social structure in native and invasive populations of the social wasp Vespula pensylvanica

Social insects rank among the most invasive of terrestrial species. The success of invasive social insects stems, in part, from the flexibility derived from their social behaviors. We used genetic markers to investigate if the social system of the invasive wasp, Vespula pensylvanica, differed in its introduced and native habitats in order to better understand variation in social phenotype in invasive social species. We found that (1) nestmate workers showed lower levels of relatedness in introduced populations than native populations, (2) introduced colonies contained workers produced by multiple queens whereas native colonies contained workers produced by only a single queen, (3) queen mate number did not differ significantly between introduced and native colonies, and (4) workers from introduced colonies were frequently produced by queens that originated from foreign nests. Thus, overall, native and introduced colonies differed substantially in social phenotype because introduced colonies more frequently contained workers produced by multiple, foreign queens. In addition, the similarity in levels of genetic variation in introduced and native habitats, as well as observed variation in colony social phenotype in native populations, suggest that colony structure in invasive populations may be partially associated with social plasticity. Overall, the differences in social structure observed in invasive V. pensylvanica parallel those in other, distantly related invasive social insects, suggesting that insect societies often develop similar social phenotypes upon introduction into new habitats.     

Cuticular hydrocarbon dynamics in young adult Polistes dominulus (Hymenoptera: Vespidae) and the role of linear hydrocarbons in nestmate recognition systems

In social insects, cuticular hydrocarbons (CHCs) play an important role in nestmate discrimination processes, but young individuals are usually not discriminated. We studied CHC changes in young workers of the social wasp Polistes dominulus. A quantitative estimation demonstrated that total quantities of CHCs increased after emergence, with branched alkanes increasing drastically when compared with other classes of hydrocarbons. The relative quantity of longer-chain compounds increased with respect to shorter ones; unsaturated compounds decreased. These changes might reduce the capacity of the cuticle to acquire compounds of environmental origin. We then tested whether individuals acquire hydrocarbons from the environment, and whether this capability equally characterises newly emerged and mature wasps. We exposed wasps of two age classes (adults younger or older than 24 h) to four linear hydrocarbons in turn, and observed how nestmates reacted to their re-introduction into the natal colony. Exposed young wasps elicited significantly more aggressive responses than control sisters; but treated wasps older than 24 h were generally accepted by nestmates. Chemical assays showed that exposed young wasps readily absorbed hydrocarbons; older ones did not incorporate hydrocarbons, suggesting that the chemical profiles of mature wasps are less prone to chemical shifts than those of newly emerged wasps.     

Nestmate recognition by guards of the Asian hive bee <Emphasis Type="Italic">Apis cerana</Emphasis>

When a honey bee colony becomes queenless and broodless its only reproductive option is for some of its workers to produce sons before the colony perishes. However, for this to be possible the policing of worker-laid eggs must be curtailed and this provides the opportunity for queenless colonies to be reproductively parasitized by workers from other nests. Such reproductive parasitism is known to occur in Apis florea and A. cerana. Microsatellite analyses of worker samples have demonstrated that the proportion of non-natal workers present in an A. cerana colony declines after a colony is made queenless. This observation suggests that queenless A. cerana colonies may be more vigilant in repelling potentially parasitic non-natal workers than queenright colonies. We compared rates of nestmate and non-nestmate acceptance in both queenright and queenless A. cerana colonies using standard assays and showed that there is no statistical difference between the proportion of non-nestmate workers that are rejected in queenless and queenright colonies. We also show that, contrary to earlier reports, A. cerana guards are able to discriminate nestmate workers from non-nestmates, and that they reject significantly more non-nestmate workers than nestmate workers. Received 25 February 2008; revised 21 May 2008; accepted 25 June 2008.     

Learning and discrimination of individual cuticular hydrocarbons by honeybees (Apis mellifera)

In social insect colonies, recognition of nestmates, kinship, caste and reproductive status is crucial both for individuals and for the colony. The recognition cues used are thought to be chemical, with the hydrocarbons found on the cuticle of insects often cited as being particularly important. However, in honeybees (Apis mellifera) the role of cuticular hydrocarbons in nestmate recognition is controversial. Here we use the proboscis extension response (PER) conditioning paradigm to determine how well honeybees learn long-chain linear alkanes and (Z)-alkenes present on the cuticle of worker bees, and also how well they can discriminate between them. We found large differences both in learning and discrimination abilities with the different cuticular hydrocarbons. Thus, the tested hydrocarbons could be classified into those which the bees learnt and discriminated well (mostly alkenes) and those which they did not (alkanes and some alkenes). These well-learnt alkenes may constitute important compounds used as cues in the social recognition processes.     

The acceptance rate of young wasps by alien colonies depends on colony developmental stages in the swarm-founding wasp, Polybia paulista von ihering (Hymenoptera: Vespidae)

In social insects, newly emerged individuals learn the colony-specific chemical label from their natal comb shortly after their emergence. These labels help to identify each individual's colony of origin and are used as a recognition template against which individuals can discriminate nestmates from non-nestmates. Our previous studies with Polybia paulista von Ihering support this general pattern, and the acceptance rate of young female and male wasps decreased as a function of their age. Our study also showed in P. paulista that more than 90% of newly emerged female wasps might be accepted by conspecific unrelated colonies. However, it has not been investigated whether the acceptance rate of newly emerged female wasps depends on colony developmental stage of recipient colonies. We introduced newly emerged female wasps of P. paulista into different colony developmental stags of recipient colonies, i.e., worker-producing and male-producing colonies. We found that the acceptance rate of newly emerged female wasps by alien colonies was pretty lower by male-producing colonies than worker-producing colonies. This is the first study to show that the acceptance rate of young female wasps depends on stages of recipient colonies.     

Polistes dominulus (Hymenoptera: Vespidae) larvae possess their own chemical signatures

Social insects use cuticular hydrocarbons (CHCs) as recognition cues in a variety of social contexts, such as species and nestmate recognition. Discrimination of nestmates is an important requisite to avoid exploitation by unrelated individuals. In social wasps, use of CHCs in nestmate recognition has been demonstrated only among adults, whereas very little is known regarding brood recognition. We performed gas chromatography-mass spectrometry analyses of the CHCs of adults and larvae of the social wasp Polistes dominulus and found that larvae possess a characteristic chemical colony-specific pattern distinct from that of adults. Behavioural assays confirmed that these are recognized and discriminated by adults. Larval epicuticular substances are therefore sufficient for recognition of nestmate larvae by adults and demonstrate that wasps are able to discriminate between alien and nestmate larval odours.     

The role of cuticular hydrocarbons as chemical cues for nestmate recognition in the invasive Argentine ant (<Emphasis Type="Italic">Linepithema humile</Emphasis>)

Argentine ants (Linepithema humile) in their native South American range, like most other ant species, form spatially restricted colonies that display high levels of aggression toward other such colonies. In their introduced range, Argentine ants are unicolonial and form massive supercolonies composed of numerous nests among which territorial boundaries are absent. Here we examine the role of cuticular hydrocarbons (CHCs) in nestmate recognition of this highly damaging invasive ant using three supercolonies from its introduced range. We conducted behavioral assays to test the response of Argentine ants to workers treated with colonymate or non-colonymate CHCs. Additionally, we quantified the amount of hydrocarbons transferred to individual ants and performed gas chromatography-mass spectrometry (GC/MS) to qualitatively characterize our manipulation of CHC profiles. The GC/MS data revealed marked differences in the hydrocarbon profiles across supercolonies and indicated that our treatment effectively masked the original chemical profile of the treated ants with the profile belonging to the foreign individuals. We found that individual workers treated with foreign CHCs were aggressively rejected by their colonymates and this behavior appears to be concentration-dependent: larger quantities of foreign CHCs triggered higher levels of aggression. Moreover, this response was not simply due to an increase in the amount of CHCs applied to the cuticle since treatment with high concentrations of nestmate CHCs did not trigger aggression.The results of this study bolster the findings of previous studies on social insects that have implicated CHCs as nestmate recognition cues and provide insight into the mechanisms of nestmate recognition in the invasive Argentine ant. Received 6 February 2007; revised 31 May and 27 July 2007; accepted 16 August 2007.     

Foreign ant queens are accepted but produce fewer offspring

Understanding social evolution requires us to understand the processes regulating the number of breeders within social groups and how they partition reproduction. Queens in polygynous (multiple queens per colony) ants often seek adoption in established colonies instead of founding a new colony independently. This mode of dispersal leads to potential conflicts, as kin selection theory predicts that resident workers should favour nestmate queens over foreign queens. Here we compared the survival of foreign and resident queens as well as their relative reproductive share. We used the ant Formica exsecta to construct colonies consisting of one queen with workers related to this resident queen and introduced a foreign queen. We found that the survival of foreign queens did not differ from that of resident queens over a period of 136 days. However, the genetic analyses revealed that resident queens produced a 1.5-fold higher number of offspring than introduced queens, and had an equal or higher share in 80% of the colonies. These data indicate that some discrimination can occur against dispersing individuals and that dispersal can thus have costs in terms of direct reproduction for dispersing queens.     

Brood adoption in the leaf-cutting ant <Emphasis Type="Italic">Acromyrmex echinatior</Emphasis>: adaptation or recognition noise?

The ability to discriminate between nestmates and non-nestmates is an important prerequisite for the evolution of eusociality. Indeed, social insect workers are typically able to discriminate between nestmate and non-nestmate workers. Adult non-nestmate workers are readily detected and rejected from the colony. Whether social insects can discriminate between nestmate and non-nestmate brood, however, is less clear. Here, we show that workers of the leaf-cutting ant Acromyrmex echinatior discriminate between nestmate and non-nestmate brood, and among brood of different stages. Initially, non-nestmate brood is attacked, but it is adopted after a delay. Adoption could occur due to inefficiency of the recognition system, or it could be adaptive because it is an inexpensive way to increase the workforce. Our results suggest that brood adoption may occur accidentally. We also report how workers replace fungal hyphae on the brood’s surface before transporting the brood into their fungus garden.     

Egg discrimination by workers in <Emphasis Type="Italic">Diacamma</Emphasis> sp. from Japan

In eusocial Hymenoptera, workers usually cannot mate but can lay male-destined haploid eggs. In contrast, in many species, worker reproduction is regulated by means of worker policing, which consists of two behavioral categories: aggression towards workers that have developed ovaries, and the destruction of worker-laid eggs. We focused on the latter behavior in the ant Diacamma sp. from Japan, in which a mated worker (gamergate) serves as the functional queen in each colony. An experiment in which we introduced both worker-laid eggs and gamergate-laid eggs from foreign colonies showed that workers preferentially destroyed the worker-laid eggs. This indicates that workers can discriminate worker-laid eggs from gamergate-laid eggs. In contrast, gamergates destroyed all introduced eggs that they discovered. Our chemical analysis of egg surface substances revealed that the worker-laid eggs clearly differed from gamergate-laid eggs in their surface hydrocarbons (SHCs), with the former containing more long-chain components. However, a bioassay to test whether specific components of the SHCs trigger the observed discrimination behavior by workers remains to be done.     

Nestmate recognition and identification of cuticular hydrocarbons composition in the swarm founding paper wasp Ropalidia opifex

Eusocial polistine wasps comprise species in which new colonies are founded by single wasps or groups of foundresses (independent founding species) or by swarms of queens and workers (swarm founding species). The first ones have relatively small societies generally comprising only one egg-laying queen, while the second ones have larger colonies where several egg-laying queens are usually present. These differences in social organisation are expected to influence the acceptance threshold of conspecifics in the colony and the intra-colony communication modalities. In this paper, we showed by field experiments that Ropalidia opifex (a swarm founding species) presents a good discrimination between alien and nestmate conspecifics. Moreover by GC/MS, we identified 19 cuticular hydrocarbons in this species, and we demonstrated that cuticular signature in this species depends mainly on colony membership, not showing any correlation with fertility.     

Acceptance by Honey Bee Guards of Non‐Nestmates is not Increased by Treatment with Nestmate Odours

Honey bee, Apis mellifera, entrance guards use chemical cues to discriminate nestmates from non‐nestmates. Previous research has shown that when wax combs are reciprocally swapped between two colonies, guards become more accepting of workers from the swap partner. However, when combs were transferred only one way, guards in the comb‐receiver colony became more accepting of bees from the comb‐donor colony, but not vice versa. Hence, the increased acceptance of non‐nestmates caused by reciprocal comb swapping was not because of introduced bees acquiring odours from the transferred combs, which was surprising because comb wax was known to affect the odour of bees. In the current experiment, we caused workers to acquire either nestmate or non‐nestmate odours by holding them for 15 min in a tube, which had previously held nestmates or non‐nestmates and then measured their acceptance by entrance guards of nestmate or non‐nestmate hives. When transferred workers had acquired odours of non‐nestmates, acceptance by their own colony’s guards significantly decreased to 66% from 91%. Conversely, the acceptance of non‐nestmates that had acquired odours of the guards’ own nestmates was unchanged, 25% vs. 25%. These results show that when equivalent changes in the odour of introduced bees are made, guards are more sensitive to changes that cause nestmates to acquire non‐nestmate odours than vice versa. These results are also a likely explanation for the earlier and surprising results from the unidirectional comb swap experiment ( Couvillon et al. 2007 ). We make a hypothesis for the underlying mechanism in terms of a multidimensional recognition cue space.     

Ant workers selfishly bias sex ratios by manipulating female development.

Kin selection theory predicts that social insects should perform selfish manipulations as a function of colony genetic structure. We describe a novel mechanism by which this occurs. First, we use microsatellite analyses to show that, in a population of the ant Leptothorax acervorum, workers' relatedness asymmetry (ratio of relatedness to females and relatedness to males) is significantly higher in monogynous (single-queen) colonies than in polygynous (multiple-queen) colonies. Workers rear mainly queens in monogynous colonies and males in polygynous colonies. Therefore, split sex ratios in this population are correlated with workers' relatedness asymmetry. Together with significant female bias in the population numerical and investment sex ratios, this finding strongly supports kin-selection theory. Second, by determining the primary sex ratio using microsatellite markers to sex eggs, we show that the ratio of male to female eggs is the same in both monogynous and polygynous colonies and equals the overall ratio of haploids (males) to diploids (queens and workers) among adults. In contrast to workers of species with selective destruction of male brood, L. acervorum workers therefore rear eggs randomly with respect to sex and must achieve their favoured sex ratios by selectively biasing the final caste (queen or worker) of developing females.     

Egg Cannibalism in a Facultative Polygynous Ant: Conflict for Reproduction or Strategy to Survive?

Policing, i.e. all behaviours that prevent a nestmate from reproducing, is currently observed in social insects. It is presumed to have evolved to regulate potential conflicts generated by genetic asymmetries or to enhance colony efficiency by avoiding surplus reproductives and brood. In the ant, Ectatomma tuberculatum, individual queen fecundity was similar in monogynous and polygynous colonies issued from a Mexican population. Egg cannibalism, however, occurred in the polygynous colonies. The stealing and destruction of reproductive queen‐laid eggs involved only nestmate queens, even if they were highly related. No queen appeared to monopolize reproduction in the polygynous colonies. But, the observed value of relatedness among workers differed from the expected value, suggesting an unequal sharing of reproduction between queens. We discussed whether the cannibalism of queen‐laid eggs in E. tuberculatum results from a competition for reproduction among queens or if this phenomenon is related to constraints on nutritional resources.