No facultative worker policing in the honey bee (Apis mellifera L.)

Kin selection theory predicts that in colonies of social Hymenoptera with multiply mated queens, workers should mutually inhibit (“police”) worker reproduction, but that in colonies with singly mated queens, workers should favor rearing workers’ sons instead of queens’ sons. In line with these predictions, Mattila et al. (Curr Biol 22:2027–2031, 2012) documented increased ovary development among workers in colonies of honey bees with singly mated queens, suggesting that workers can detect and respond adaptively to queen mating frequency and raising the possibility that they facultative police. In a follow-up experiment, we test and reject the hypothesis that workers in single-patriline colonies prefer worker-derived males and are able to reproduce directly; we show that their eggs are policed as strongly as those of workers in colonies with multiply mated queens. Evidently, workers do not respond facultatively to a kin structure that favors relaxed policing and increased direct reproduction. These workers may instead be responding to a poor queen or preparing for possible queen loss.     

Aggressive reproductive competition among hopelessly queenless honeybee workers triggered by pheromone signaling

In the honeybee, Apis mellifera, the queen monopolizes reproduction, while the sterile workers cooperate harmoniously in nest maintenance. However, under queenless (QL) conditions, cooperation collapses and reproductive competition among workers ensues. This is mediated through aggression and worker oviposition, as well as shifts in pheromones, from worker to queen-like composition. Many studies suggest a dichotomy between conflict resolution through aggression or through pheromonal signaling. In this paper, we demonstrate that both phenomena comprise essential components of reproductive competition and that pheromone signaling actually triggers the onset of aggression. We kept workers as QL groups until first aggression was observed and subsequently determined the contestants' reproductive status and content of the mandibular (MG) and Dufour's glands (DG). In groups in which aggression occurred early, the attacked bee had consistently more queen-like pheromone in both the MG and DG, although both contestants had undeveloped ovaries. In groups with late aggression, the attacked bee had consistently larger oocytes and more queen-like pheromone in the DG, but not the MG. We suggest that at early stages of competition, the MG secretion is utilized to establish dominance and that the DG provides an honest fertility signal. We further argue that it is the higher amount of DG pheromone that triggers aggression.     

Aggressions and size-related fecundity of queenless workers in the ant <Emphasis Type="Italic">Cataglyphis cursor</Emphasis>

In social hymenoptera, the reproductive division of labor is often linked to differences in individual body size with the reproductive caste (the queen) being larger than the workers. Likewise, the reproductive potential may vary with size within the worker caste and could affect the evolution of worker size in social insects. Here, we tested the relationship between worker size and reproductive potential in the facultative parthenogenetic ant Cataglyphis cursor. Colonies are headed by a multiply mated queen, but workers can produce gynes (virgin queens) and workers by thelytokous parthenogenesis after the queen's death. We observed the behaviour of workers (n = 357) until the production of gynes (212 h over 3 months) in an orphaned colony (mated queen not present). The size of workers was measured, and their paternal lineage determined using six microsatellite markers, to control for an effect of patriline. Larger workers were more likely to reproduce and lay more eggs indicating that individual level selection could take place. However, paternal lineage had no effect on the reproductive potential and worker size. From the behavioural and genetic data, we also show for the first time in this species, evidence of aggressive interactions among workers and a potential for nepotism to occur in orphaned colonies, as the five gynes produced belonged to a single paternal lineage.     

Irregular brood patterns and worker reproduction in social wasps

The potential for reproductive conflict among colony members exists in all social insect societies. For example, queens and workers may be in conflict over the production of males within colonies. Kin selection theory predicts that in a colony headed by a multiply-mated queen, worker reproduction is prevented by worker policing in the form of differential oophagy. However, few studies have demonstrated that workers actually lay eggs within queenright colonies. The purpose of this study was to determine if workers laid male eggs within unmanipulated queen-right colonies of the polyandrous social wasps Vespula maculifrons and V. squamosa. We focused our analysis on an unusual brood pattern within colonies, multiple egg cells. We were primarily interested in determining if individuals reared in these irregular circumstances were queen or worker offspring. To address this question, we genotyped 318 eggs from eight V. maculifrons and two V. squamosa colonies. No worker‑reproduction was detected in any of the queenright colonies; all of the eggs found in multiple egg cells were consistent with being queen‑produced. However, the frequency of multiple egg cells differed among colonies, suggesting that queens vary in the frequency of errors they make when laying eggs within cells. Finally, we suggest that workers may not be laying eggs within queenright colonies and that worker reproduction may be controlled through mechanisms other than differential oophagy in polyandrous Vespula wasps.     

Fertility signals in the bumblebee Bombus terrestris (Hymenoptera: Apidae)

In eusocial Hymenoptera, queen control over workers is probably inseparable from the mechanism of queen recognition. In primitively eusocial bumblebees (Bombus), worker reproduction is controlled not only by the presence or absence of a dominant queen but also by other dominant workers. Furthermore, it was shown that the queen dominance is maintained by pheromonal cues. We investigated whether there is a similar odor signal released by egg-laying queens and workers that may have a function as a fertility signal. We collected cuticular surface extracts from nest-searching and breeding Bombus terrestris queens and workers that were characterized by their ovarian stages. In chemical analyses, we identified 61 compounds consisting of aldehydes, alkanes, alkenes, and fatty acid esters. Nest-searching queens and all groups of breeding females differed significantly in their odor bouquets. Furthermore, workers before the competition point (time point of colony development where workers start to develop ovaries and lay eggs) differed largely from queens and all other groups of workers. Breeding queens showed a unique bouquet of chemical compounds and certain queen-specific compounds, and the differences toward workers decrease with an increasing development of the workers' ovaries, hinting the presence of a reliable fertility signal. Among the worker groups, the smallest differences were found after the competition point. Egg-laying females contained higher total amounts of chemical compounds and of relative proportions of wax-type esters and aldehydes than nest-searching queens and workers before the competition point. Therefore, these compounds may have a function as a fertility signal present in queens and workers.     

Daughters inherit colonies from mothers in the 'living-fossil' ant Nothomyrmecia macrops

Newly mated queens of monogynous (single queen) ants usually found their colonies independently, without the assistance of workers. In polygynous (multiple queen) species queens are often adopted back into their natal nest and new colonies are established by budding. We report that the Australian 'living-fossil' ant, Nothomyrmecia macrops, is exceptional in that its single queen can be replaced by one of the colony's daughters. This type of colony founding is an interesting alternative reproductive strategy in monogynous ants, which maximizes fitness under kin selection. Successive queen replacement results in a series of reproductives over time (serial polygyny), making these colonies potentially immortal. Workers raise nieces and nephews (relatedness h 0.375) the year after queen replacement. Although N. macrops is 'primitive' in many other respects, colony inheritance is likely to be a derived specialization resulting from ecological constraints on solitary founding.     

Docile sitters and active fighters in paper wasps: a tale of two queens

Ropalidia marginata and Ropalidia cyathiformis are sympatric, primitively eusocial paper wasps widely distributed in peninsular India. We compare the two species, especially their queens, in an attempt to begin to understand the role of the power of queens over their workers, in social organisation and evolution. Queens of R. marginata have lower levels of activity, rates of interactions and dominance behaviour, compared with queens of R. cyathiformis. For the same variables, R. marginata queens are either indistinguishable from or have lower values than their workers, while R. cyathiformis queens have higher values than their workers. R. marginata queens never occupy the top rank while R. cyathiformis queens are always at the top of the behavioural dominance hierarchies of their colonies. R. marginata queens thus do not appear to use dominance behaviour to suppress reproduction by their workers, while R. cyathiformis queens appear to do so. These different mechanisms used by the two queens to regulate worker reproduction give them different powers over their workers, because R. marginata queens are completely successful in suppressing reproduction by their nestmates while in R. cyathiformis colonies, other individuals also sometimes lay eggs. There is also some evidence that the different powers of the queens result in different mechanisms of regulation of worker foraging in the two species--decentralised, self-regulation in R. marginata and relatively more centralised regulation by the queen in R. cyathiformis. Thus we show here, perhaps for the first time, that the power of the queens over their workers can have important consequences for social organisation and evolution.     

Thelytokous parthenogenesis by queens in the dacetine ant Pyramica membranifera (Hymenoptera: Formicidae)

Thelytokous parthenogenesis in which diploid females are produced from unfertilized eggs, was recently reported for some ant species. Here, we document thelytokous reproduction by queens in the polygynous species Pyramica membranifera. Queens that emerged in the laboratory were kept with or without workers under laboratory conditions. Independent colony founding was successful for a few queens if prey was provided. All artificial colonies, which started with a newly emerged queen and workers produced new workers and some of the colonies also produced female sexuals. Some of the female sexuals shed their wings in the laboratory and started formation of new polygynous colonies. Workers had no ovaries and thus, were obligatorily sterile.     

Scent of a queen—cuticular hydrocarbons specific for female reproductives in lower termites

In social insects, it is assumed that signals of the queen inform nestmates about her reproductive status. Thus, workers forego their own reproduction if the queen signals high fertility. In hemimetabolous termites, little is known about reproductive inhibition, but evidence exists for a royal-pair control. Workers of lower termites exhibit a high developmental flexibility and are potentially able to become reproductives, but the presence of a fertile reproductive restrains them from reaching sexual maturity. The nature of this control, however, remains unknown. Here, we report on qualitative differences in cuticular hydrocarbon profiles between queens and workers of the basal drywood termite Cryptotermes secundus. Queens were characterized by a shift to long-chained and branched hydrocarbons. Most remarkably, similar chemical patterns are regarded as fertility cues of reproductives in social Hymenoptera. This might suggest that both groups of social insects convergently evolved similar chemical signatures. The present study provides deeper insights into how termites might have socially exploited these signatures from sexual communication in their cockroach-like ancestor. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Costs and benefits of subordinate queens in colonies of the ant,<Emphasis Type="Italic"> Leptothorax gredleri</Emphasis>

Queens of some species of social insects form dominance hierarchies in which only the top-ranking individual reproduces. Such unequal partitioning of reproduction can be stable when subordinate queens increase their inclusive fitness indirectly by helping to rear a related dominant queen's offspring. We investigated whether subordinate queens of the ant, Leptothorax gredleri, affect the reproductive success of the dominant queen. In laboratory colonies with two queens, first eggs were laid approximately 20 days later than in colonies with a single queen, presumably due to aggressive competition among nestmate queens. Nevertheless, two-queen colonies produced on average more brood than one-queen colonies, although egg laying was completely or almost completely monopolized by one single queen. The presence of a subordinate queen therefore appears to increase the productivity of the dominant.     

Winged queens replaced by reproductives smaller than workers in <Emphasis Type="Italic">Mystrium</Emphasis> ants

In ants, winged queens that are specialized for independent colony foundation can be replaced by wingless reproductives better adapted for colony fission. We studied this shift in reproductive strategy by comparing two Mystrium species from Madagascar using morphometry, allometry and dissections. Mystrium rogeri has a single dealate queen in each colony with a larger thorax than workers and similar mandibles that allow these queens to hunt during non-claustral foundation. In contrast, Mystrium ‘red’ lacks winged queens and half of the female adults belong to a wingless ‘intermorph’ caste smaller and allometrically distinct from the workers. Intermorphs have functional ovaries and spermatheca while those of workers are degenerate. Intermorphs care for brood and a few mate and reproduce making them an all-purpose caste that takes charge of both work and reproduction. However, their mandibles are reduced and inappropriate for hunting centipedes, unlike the workers’ mandibles. This together with their small thorax disallow them to perform independent colony foundation, and colonies reproduce by fission. M. rogeri workers have mandibles polymorphic in size and shape, which allow for all tasks from brood care to hunting. In M. ‘red’, colonial investment in reproduction has shifted from producing expensive winged queens to more numerous helpers. M. ‘red’ intermorphs are the first case of reproductives smaller than workers in ants and illustrate their potential to diversify their caste system for better colonial economy.     

Sex-specific inhibition and stimulation of worker-reproductive transition in a termite

In social insects, the postembryonic development of individuals exhibits strong phenotypic plasticity in response to the environment, thus generating the caste system. Different from eusocial Hymenoptera, in which queens dominate reproduction and inhibit worker fertility, the primary reproductive caste in termites (kings and queens) can be replaced by neotenic reproductives derived from functionally sterile individuals. Feedback regulation of nestmate differentiation into reproductives has been suggested, but the sex specificity remains inconclusive. In the eastern subterranean termite, Reticulitermes flavipes, we tested the hypothesis that neotenic reproductives regulate worker-reproductive transition in a sex-specific manner. With this R. flavipes system, we demonstrate a sex-specific regulatory mechanism with both inhibitory and stimulatory functions. Neotenics inhibit workers of the same sex from differentiating into additional reproductives but stimulate workers of the opposite sex to undergo this transition. Furthermore, this process is not affected by the presence of soldiers. Our results highlight the reproductive plasticity of termites in response to social cues and provide insights into the regulation of reproductive division of labor in a hemimetabolous social insect.     

Evidence for intra-colonial genetic variance in resistance to American foulbrood of honey bees (<Emphasis Type="Italic">Apis mellifera</Emphasis>): further support for the parasite/pathogen hypothesis for the evolution of polyandry

Explanations for the evolution of multiple mating by social insect (particularly honey bee) queens have been frequently sought. An important hypothesis is that multiple mating is adaptive because it increases intracolonial genetic diversity and thereby reduces the likelihood that parasites or pathogens will catastrophically infect a colony. We tested one assumption of this model: that honey bee worker patrilines should differ in disease resistance. We used American foulbrood (caused by the bacterium Paenibacillus larvae) as a model pathogen. We found that patrilines within colonies do indeed vary in their resistance to this disease.     

Potential increase in mating frequency of queens in feral colonies of Bombus terrestris introduced into Japan

With the exception of several species, bumblebees are monandrous. We examined mating frequency in feral colonies of the introduced bumblebee Bombus terrestris in Japan. Using microsatellite markers, genotyping of sperm DNA stored in the spermatheca of nine queens detected multiple insemination paternities in one queen; the others were singly mated. The average effective paternity frequency estimated from the genotypes of queens and workers was 1.23; that estimated from the workers’ genotype alone was 2.12. These values were greater than those of laboratory-reared colonies in the native ranges of B. terrestris. The genotypes of one or two workers did not match those of their queens or showed paternities different from those of their nestmates; this may have arisen from either queen takeover or drifting of workers. These alien workers were responsible for the heterogeneous genotype distribution within each B. terrestris colony, resulting in higher estimates of paternity frequency than of insemination frequency. The high mating frequency of introduced B. terrestris may have occurred by artificial selection through mass breeding for commercialization. Moreover, polyandrous queens may be selectively advantageous, because reproduction by such queens is less likely to be disturbed by interspecific mating than that by monandrous queens.     

Postmating changes in cuticular chemistry and visual appearance in <Emphasis Type="Italic">Ectatomma tuberculatum</Emphasis> queens (Formicidae: Ectatomminae)

In the ectatommine ant Ectatomma tuberculatum, the visual appearance of queens changes after mating and ovarian development in that their cuticle turns from shiny to matte. In this study, we have shown that this change seems to be caused by 15-fold accumulation of hydrocarbons, in particular heptacosane that covers the multiple grooves present on the cuticular surface creating a wax coat in mated fully fertile queens. Analyses of the scrapped wax revealed that it is composed largely of heptacosane. Peak-by-peak comparison of the cuticular hydrocarbon (CHC) composition of mated, virgin with developed ovaries and virgin with nondeveloped ovaries revealed significant differences between the queen groups. Although the total amount of the CHC of virgin queens with developed ovaries was not higher than virgin queens that did not have developed ovaries, the composition showed a shift toward the mated queen. While it is possible that the large accumulation of hydrocarbons may give extra physical and chemical protection to queens, we propose that the switch in the relative abundance of heptacosane and nonacosane and perhaps of other components is indicative of being a mating and fertility cue. This is the first report in social insects where external chemical changes are accompanied by changes in visual appearance.     

A critical number of workers in a honeybee colony triggers investment in reproduction

Social insect colonies, like individual organisms, must decide as they develop how to allocate optimally their resources among survival, growth, and reproduction. Only when colonies reach a certain state do they switch from investing purely in survival and growth to investing also in reproduction. But how do worker bees within a colony detect that their colony has reached the state where it is adaptive to begin investing in reproduction? Previous work has shown that larger honeybee colonies invest more in reproduction (i.e., the production of drones and queens), however, the term ‘larger’ encompasses multiple colony parameters including number of adult workers, size of the nest, amount of brood, and size of the honey stores. These colony parameters were independently increased in this study to test which one(s) would increase a colony’s investment in reproduction via males. This was assayed by measuring the construction of drone comb, the special type of comb in which drones are reared. Only an increase in the number of workers stimulated construction of drone comb. Colonies with over 4,000 workers began building drone comb, independent of the other colony parameters. These results show that attaining a critical number of workers is the key parameter for honeybee colonies to start to shift resources towards reproduction. These findings are relevant to other social systems in which a group’s members must adjust their behavior as a function of the group’s size.     

No evidence of volatile chemicals regulating reproduction in a multiple queen ant

Efficient cooperation in eusocial insect colonies requires effective communication, and there is abundant evidence of non-volatile chemicals playing a role in regulating reproduction within colonies. In contrast, there have been fewer studies investigating the role of volatile chemicals. This study investigated the potential role of volatile chemicals in regulating queen reproduction either by directly inhibiting queen reproduction or by honestly signalling queen fecundity to workers. We tested this using multiple queen colonies of the ant (Leptothorax acervorum) from a functionally monogynous population where one queen monopolizes all reproduction. Nine colonies, each with an established laying queen, were split to produce two colony fragments—one containing the reproducing queen (group 1) and one containing only previously non-reproducing queens (group 2). Each group was separated by a fine wire mesh preventing physical contact, but allowing volatile chemical contact. In each group 2 fragment, we found that a single formerly non-reproductive queen commenced reproduction and that the rate of egg laying and maximum number of eggs recorded did not significantly differ between groups 1 and 2, results that do not support volatile chemicals as playing a role in regulating queen reproduction. Instead, our findings suggest that physical contact is necessary to maintain functional monogyny.     

Can Dufour��s gland compounds honestly signal fertility in the primitively eusocial wasp Ropalidia marginata?

Unlike queens of typical primitively eusocial species, Ropalidia marginata queens are docile and non-interactive, and hence cannot be using dominance to maintain their status. It appears that the queen maintains reproductive monopoly through a pheromone, of which the Dufour's gland is at least one source. Here, we reconfirm earlier results showing that queens and workers can be correctly classified on a discriminant function using the compositions of their respective Dufour's glands, and also demonstrate consistent queen-worker differences based on categories of compounds and on single compounds also in some cases. Since the queen pheromone is expected to be an honest signal of the fecundity of a queen, we investigate the correlation of Dufour's gland compounds with ovarian activation of queens. Our study shows that Dufour's gland compounds in R. marginata correlate with the state of ovarian activation of queens, suggesting that such compounds may portray the fecundity of a queen, and may indeed function as honest signals of fertility.     

A reassessment of the mating system characteristics of the army ant Eciton burchellii

In a recent study, Denny et al. (2004a) showed that queens of the army ant, Eciton burchellii, mate with multiple males and presented estimates suggesting that they mate with more males than queens of any other ant species so far investigated. They also inferred that data were consistent with queens being inseminated repeatedly throughout their life, which would be exceptional among the social Hymenoptera and contradictory to predictions from kin selection theory. In the present study, we reanalyze these data using new software and supplement them with similar microsatellite data from other colonies of the same species. Mating frequencies in E. burchellii are indeed very high (mean observed and effective queen-mating frequencies of 12.9 each) but considerably lower than the previous estimates. We show that the number of patrilines represented in the first worker offspring of a young queen is lower than in older queens but suggest that this may be due to initial sperm clumping in the queen’s sperm storage organ, rather than to repeated inseminations. Moreover, we found no evidence for repeated mating by genotyping sequential worker generations produced by a single old queen, showing that she did not obtain new inseminations despite ample opportunities for mating.