Worker Ovary Development Research Articles

Surplus feeding reverses worker ovarian suppression and disrupts social cohesion in queen-right wasp colonies

Nutrition is an important component of oogenesis and ovarian development in insects. In social insect colonies where a large proportion of females are sterile, suppression of reproduction caused by differential acquisition and retention of nutrition has been hypothesized to cause worker sterility. This study, using the primitively eusocial wasp Ropalidia marginata (Lepeletier) (Hymenoptera: Vespidae), show that worker ovarian development in naturally foraging colonies is more similar to experimentally food-limited colonies than to surplus-fed colonies, indicating nutritional castration in workers whose ovaries are inhibited by food availability. Further, experimental provisioning of surplus nutrition led to higher ovarian development in workers as compared to naturally foraging and food-limited workers. Surplus feeding also led to higher nest desertion by workers, leading to a breakdown of the colony’s social structure, whereas food-limited colonies retained workers.

INSIGHTS OF CASTE DETERMINATIONS IN SOCIAL INSECTS

Eusocial insects display caste structures in which reproductive ability is possessed by a single or a few queens while all other colony members act as workers. In social insects like ants, bees, and termites, vital physiological processes are regulated at the colony scale. Females in social insects have at least one reproductive caste and one nonreproductive caste; many termites have at least two male castes. The castes have considerable anatomical, physiological, and behavioural differences in higher social insects. Organismal systems, such as pheromone sensing, hormone signaling, and brain signaling pathways, are deployed in novel circumstances to impact nestmate and colony behaviours due to physiological decentralization over evolutionary time. Significant morphogenesis with region-specific cellular proliferation and degradation occurs during soldier development through two moulting via a presoldier stage in termite. JH action has been developed, in which a high JH titer causes soldier differentiation and a low JH titer causes alate differentiation. A monogamous pair of primary reproductives (one king and one queen) generated from alates normally establishes termite colonies (winged adults). The nymph-alate pathway (sexual pathway) or the worker pathway differentiates larvae in Reticulitermes termites (neuter pathway). Haplo-diploid sex determination controls the first developmental transition, in which unfertilized (haploid) embryos become males and fertilized (diploid) embryos become females in the case of Cataglyphis ant genus. The queen’s mandibular gland secretion, a mix of fatty acids and aromatic chemicals, is critical for maintaining the reproductive division of labour in honeybees (Apis mellifera), suppressing ovary growth in workers. Besides this, the brood produced by the queen also inhibits ovary development in workers by emitting two pheromones: the brood pheromone (BP), mainly composed of esters, and the highly volatile E-b-ocimene.

Attack of the dark clones the genetics of reproductive and color traits of South African honey bees (Apis mellifera spp.).

The traits of two subspecies of western honey bees, Apis mellifera scutellata and A.m. capensis, endemic to the Republic of South Africa (RSA), are of biological and commercial relevance. Nevertheless, the genetic basis of important phenotypes found in these subspecies remains poorly understood. We performed a genome wide association study on three traits of biological relevance in 234 A.m. capensis, 73 A.m. scutellata and 158 hybrid individuals. Thirteen markers were significantly associated to at least one trait (P ≤ 4.28 × 10-6): one for ovariole number, four for scutellar plate and eight for tergite color. We discovered two possible causative variants associated to the respective phenotypes: a deletion in GB46429 or Ebony (NC_007070.3:g.14101325G>del) (R69Efs*85) and a nonsense on GB54634 (NC_007076.3:g.4492792A>G;p.Tyr128*) causing a premature stop, substantially shortening the predicted protein. The mutant genotypes are significantly associated to phenotypes in A.m. capensis. Loss-of-function of Ebony can cause accumulation of circulating dopamine, and increased dopamine levels correlate to ovary development in queenless workers and pheromone production. Allelic association (P = 1.824 x 10-5) of NC_007076.3:g.4492792A>G;p.Tyr128* to ovariole number warrants further investigation into function and expression of the GB54634 gene. Our results highlight genetic components of relevant production/conservation behavioral phenotypes in honey bees.

Histone acetylation regulates the expression of genes involved in worker reproduction in the ant Temnothorax rugatulus

BackgroundIn insect societies, queens monopolize reproduction while workers perform tasks such as brood care or foraging. Queen loss leads to ovary development and lifespan extension in workers of many ant species. However, the underlying molecular mechanisms of this phenotypic plasticity remain unclear. Recent studies highlight the importance of epigenetics in regulating plastic traits in social insects. Thus, we investigated the role of histone acetylation in regulating worker reproduction in the ant Temnothorax rugatulus. We removed queens from their colonies to induce worker fecundity, and either fed workers with chemical inhibitors of histone acetylation (C646), deacetylation (TSA), or the solvent (DMSO) as control. We monitored worker number for six weeks after which we assessed ovary development and sequenced fat body mRNA.ResultsWorkers survived better in queenless colonies. They also developed their ovaries after queen removal in control colonies as expected, but not in colonies treated with the chemical inhibitors. Both inhibitors affected gene expression, although the inhibition of histone acetylation using C646 altered the expression of more genes with immunity, fecundity, and longevity functionalities. Interestingly, these C646-treated workers shared many upregulated genes with infertile workers from queenright colonies. We also identified one gene with antioxidant properties commonly downregulated in infertile workers from queenright colonies and both C646 and TSA-treated workers from queenless colonies.ConclusionOur results suggest that histone acetylation is involved in the molecular regulation of worker reproduction, and thus point to an important role of histone modifications in modulating phenotypic plasticity of life history traits in social insects.

Cuticular and Dufour's Gland Chemistry Reflect Reproductive and Social State in the Facultatively Eusocial Sweat Bee Megalopta genalis (Hymenoptera: Halictidae).

Queen pheromones evolved independently in multiple eusocial insect lineages, in which they mediate reproductive conflict by inhibiting worker ovarian development. Although fundamentally important for reproductive division of labor - the hallmark of eusociality - their evolutionary origins are enigmatic. Here, we analyze cuticular and Dufour's gland chemistries across alternative social and reproductive phenotypes in Megalopta genalis bees (tribe Augochlorini, family Halictidae) that facultatively express simple eusociality. Reproductive bees have distinct overall glandular and cuticular chemical phenotypes compared with non-reproductive workers. On the cuticle, a likely site of signal transmission, reproductives are enriched for certain alkenes, most linear alkanes, and are heavily enriched for all methyl-branched alkanes. Chemicals belonging to these compound classes are known to function as fertility signals in other eusocial insect taxa. Some macrocyclic lactones, compounds that serve as queen pheromones in the other eusocial halictid tribe (Halictini), are also enriched among reproductives relative to workers. The intra-population facultative eusociality of M. genalis permits direct comparisons between individuals expressing alternative reproductive phenotypes - females that reproduce alone (solitary reproductives) and social queens - to highlight traits in the latter that may be important mediators of eusociality. Compared with solitary reproductives, the cuticular chemistries of queens are more strongly differentiated from those of workers, and furthermore are especially enriched for methyl-branched alkanes. Determining the pheromonal function(s) and information content of the candidate signaling compounds we identify will help illuminate the early evolutionary history of queen pheromones, chemical signals central to the organization of insect eusocial behavior.

Honeybees possess a structurally diverse and functionally redundant set of queen pheromones.

Queen pheromones, which signal the presence of a fertile queen and induce workers to remain sterile, play a key role in regulating reproductive division of labour in insect societies. In the honeybee, volatiles produced by the queen's mandibular glands have been argued to act as the primary sterility-inducing pheromones. This contrasts with evidence from other groups of social insects, where specific queen-characteristic hydrocarbons present on the cuticle act as conserved queen signals. This led us to hypothesize that honeybee queens might also employ cuticular pheromones to stop workers from reproducing. Here, we support this hypothesis with the results of bioassays with synthetic blends of queen-characteristic alkenes, esters and carboxylic acids. We show that all these compound classes suppress worker ovary development, and that one of the blends of esters that we used was as effective as the queen mandibular pheromone (QMP) mix. Furthermore, we demonstrate that the two main QMP compounds 9-ODA and 9-HDA tested individually were as effective as the blend of all four major QMP compounds, suggesting considerable signal redundancy. Possible adaptive reasons for the observed complexity of the honeybee queen signal mix are discussed.

Reproductive environment affects learning performance in bumble bees

Despite a presumed fitness advantage for individuals with well-developed cognitive abilities, learning performance is usually found to be highly variable within a population. Although little is currently known about the mechanisms responsible for maintaining such variation, there is correlative evidence to suggest that learning performance may be linked to reproductive physiology in the social insects. Bumble bee colonies naturally undergo an initial co-operative phase, when only the queen reproduces, and a subsequent competition phase when all colony members compete to produce male offspring. We experimentally induced these distinct phases by manipulating the presence/absence of the queen and assessed changes in sucrose responsiveness and learning performance. We found that nest-based workers upregulated their reproductive potential in queenless colonies, and correspondingly, these bees were more responsive to sucrose than their queenright counterparts, performing better in an olfactory learning task as a result. These findings suggest that differences in ovarian development are responsible for at least some of the remarkable variation in learning performance that can be observed among very closely related members of social insect colonies. Cognitive abilities are often assumed to be inherently adaptive, so the question of why individuals vary in their learning ability has received relatively little attention. Here, we focus on reproductive status as a proximate cause of variation in learning ability in the social insects. We show that a significant proportion of the surprising variation that exists between genetically similar colony members can be explained by worker ovarian development; reproductively active workers are more sensitive to food rewards and thus learn more quickly. Learning ability may be one of a suite of correlated traits that are linked to reproductive physiology in social insects and therefore play an important role in the evolution of division of labour.

Highly specific responses to queen pheromone in three Lasius ant species

Queen pheromones mediate the reproductive division of labor in social insect colonies and provide novel opportunities for investigating the evolution of animal communication. Previous work found that queens in the ant genus Lasius produce several 3-methylalkanes in greater relative amounts than workers do. At least one of these (3-MeC31) is a queen pheromone that regulates worker sterility in two Lasius species, although there are indications that other 3-methylalkanes might also function as queen pheromones. Here, we presented workers from three Lasius species with four different 3-methylalkanes, and measured the effect on worker ovary development. In all three species, only 3-MeC31 showed clear evidence of inhibiting worker fecundity. Our results suggest that worker ants can discriminate homologous hydrocarbons that differ in chain length and only treat specific homologs as queen pheromones. These results provide insight into the conflicting selective pressures on cuticular hydrocarbons arising from their multiple roles in signaling and adaptation to the abiotic environment.

Biological activity of the enantiomers of 3-methylhentriacontane, a queen pheromone of the ant Lasius niger.

Queen pheromones are essential for regulation of the reproductive division of labor in eusocial insect species. Although only the queen is able to lay fertilized eggs and produce females, in some cases workers may develop their ovaries and lay male-destined eggs, thus reducing the overall colony efficiency. As long as the queen is healthy, it is usually in the workers' collective interest to work for the colony and remain sterile. Queens signal their fertility via pheromones, which may have a primer effect, affecting the physiology of workers, or a releaser effect, influencing worker behavior. The queen pheromone of the ant Lasius niger was among the first queen pheromones of social insects to be identified. Its major component is 3-methylhentriacontane (3-MeC31), which is present in relatively large amounts on the queen's cuticle and on her eggs. 3-MeC31 regulates worker reproduction by inhibiting ovarian development. Most monomethyl-branched hydrocarbons can exist in two stereoisomeric forms. The correct stereochemistry is fundamental to the activity of most bioactive molecules, but this has rarely been investigated for methyl-branched hydrocarbons. Here, we tested the bioactivity of the (S)- and (R)-enantiomers of 3-MeC31, and found that whereas both enantiomers were effective in suppressing worker ovarian development, (S)-3-MeC31 appeared to be more effective at suppressing aggressive behavior by workers. This suggests that the natural pheromone may be a mixture of the two enantiomers. The enantiomeric ratio produced by queens remains unknown because of the small amounts of the compound available from each queen.

Assessing the role of β-ocimene in regulating foraging behavior of the honey bee, Apis mellifera

The larval pheromone (E)-β-ocimene (EBO) inhibits worker ovary development and accelerates the behavioral transition from nursing to foraging in the honey bee, Apis mellifera. Because EBO diffuses throughout the hive, it has been hypothesized that honey bee workers can use it to indirectly assess brood abundance. Here, we evaluate this hypothesis by testing whether EBO plays a role in the regulation of worker foraging activity. We exposed paired colonies to a pulse of either synthetic β-ocimene or paraffin oil control and recorded the subsequent number of pollen and non-pollen foragers returning to the colony. Exposure to synthetic β-ocimene moderately but significantly increased overall foraging activity, but not pollen foraging. For pollen foraging, the effect of pheromone treatment was the greatest directly after pheromone exposure.

Phenological, but not social, variation associated with climate differences in a eusocial sweat bee, Halictus ligatus, nesting in southern Ontario

Studies of annual and geographic variation in eusocial bee populations suggest that more stringent environmental conditions result in stronger reproductive skew favouring queens, while moderate conditions favour increasing worker reproduction. To test these predictions, we compared the phenology and colony development of H. ligatus nesting in St. Catharines, Ontario, Canada to a previously studied aggregation 90 km north of St. Catharines, in Victoria, Ontario. Despite the close proximity of these two locations, St. Catharines has markedly shorter winters and longer summers. Comparisons between St. Catharines in 2006 and Victoria in the 1980s and 1990s incorporate both geographic differences in climate and temporal differences due to climate change. We predicted that St. Catharines foundress queens should emerge from hibernation and initiate nests earlier in spring, giving them time to produce more workers. Since earlier studies indicated that queens have difficulty suppressing worker reproduction in larger colonies, we also predicted higher rates of worker ovarian development in St. Catharines. In spring and summer 2006, we excavated 65 H. ligatus nests, comparing their contents to 713 specimens collected in pan traps. As predicted, nests were initiated about a month earlier in St. Catharines than in Victoria, but contrary to prediction, fewer workers were produced in St. Catharines. St. Catharines workers were just as likely to have developed ovaries as Victoria workers. About 40% of St. Catharines workers were classified as reproductive, and larger reproductive workers tended to have higher ovarian scores. Early queen mortality in the longer nest cycle of St. Catharines bees may have enhanced opportunities for worker reproduction despite their smaller numbers. Novel features of H. ligatus sociobiology in St. Catharines included evidence that queens can initiate new nests following the loss of their first brood, overlap between worker and gyne production within some nests, and high rates of independent nest founding by worker-sized females, suggesting that many worker-brood females overwinter. Overall, the distinctly warmer climate of St. Catharines compared to Victoria led to earlier nest initiation and lengthening of the flight season, but not to the predicted differences in colony social organisation or queen-worker reproductive skew. A second objective of our study was to assess how well pan trap collections capture important information about demographic and social parameters important in assessing social variability in sweat bees. Nest excavations and pan traps produced similar results, suggesting that pan traps are a good alternative when nest excavations are impossible.

Bumblebee size polymorphism and worker response to queen pheromone.

Queen pheromones are chemical signals produced by reproductive individuals in social insect colonies. In many species they are key to the maintenance of reproductive division of labor, with workers beginning to reproduce individually once the queen pheromone disappears. Recently, a queen pheromone that negatively affects worker fecundity was discovered in the bumblebee Bombus terrestris, presenting an exciting opportunity for comparisons with analogous queen pheromones in independently-evolved eusocial lineages such as honey bees, ants, wasps and termites. I set out to replicate this discovery and verify its reproducibility. Using blind, controlled experiments, I found that n-pentacosane (C25) does indeed negatively affect worker ovary development. Moreover, the pheromone affects both large and small workers, and applies to workers from large, mature colonies as well as young colonies. Given that C25 is readily available and that bumblebees are popular study organisms, I hope that this replication will encourage other researchers to tackle the many research questions enabled by the discovery of a queen pheromone.

Almost royal: incomplete suppression of host worker ovarian development by a social parasite wasp

Insect social parasites, like other parasites, may benefit from inhibiting their host from reproducing (complete or partial parasitic castration) because they can then exploit more of the host’s resources for their own reproduction. In particular, social parasites that kill or expel the host queen need to prevent host workers from reproducing; this is a common worker response to the absence of their queen. Indeed, host workers would benefit from detecting the presence of the parasite and investing in direct and indirect fitness. Studying whether and how social parasites control host worker reproduction can provide information about the degree of integration of the parasite in the host colony and help identify factors regulating workers’ reproductive decisions in social insects. We investigated whether the paper wasp social parasite, Polistes sulcifer, suppresses Polistes dominula (host) worker reproduction as efficiently as the dominant host female does in queen-right colonies by comparing worker reproductive efforts in parasitized and non-parasitized (control) colonies. Our results show that 6 weeks after usurpation of their colony by the social parasite, parasitized workers (1) had more developed ovaries than control workers and (2) laid more eggs as soon as the opportunity arose. This reproductive readiness of parasitized workers was not apparent 2 weeks after colony usurpation. This suggests that P. dominula workers have evolved means to react to social parasitism, as occurs in some ants, and that the parasite has only limited control over host reproduction.

Worker self-restraint and policing maintain the queen’s reproductive monopoly in a pseudomyrmecine ant

Division of reproductive labor in insect societies is often based on worker self-restraint and both queen and worker policing. Workers of many hitherto studied wasps, bees and ants do not lay eggs in the presence of a queen. However, it is presently unclear how far these observations in a few select clades can be generalized. We investigated if and how queens maintain a reproductive monopoly in colonies of the elongate twig ant, Pseudomyrmex gracilis, a member of the previously unstudied ant subfamily Pseudomyrmecinae. Colonies are usually headed by a single, singly mated queen (monogyny, monandry). Workers therefore would be more closely related to males produced by other workers (r = 0.375) than to the sons of queens (r = 0.25). Nevertheless, workers appear to refrain from laying male-destined eggs in the presence of the queen. In queenless conditions, workers form dominance hierarchies by antennal boxing, and only one or a few high-ranking individuals readily begin to lay eggs. When returned into a queenright colony, egg-laying workers are immediately bitten, stung and expelled or killed by other workers. While the composition of cuticular hydrocarbons clearly differed between castes, it less clearly reflected worker ovarian development. An association with worker ovarian development that would allow workers to monitor the reproductive status of nestmates could only be tentatively postulated for certain substances. Our study broadens our knowledge about reproductive conflict in social Hymenoptera and shows that worker sterility in the presence of a queen is more common in monogynous, monandrous ants than expected from relatedness alone.

Two pathways ensuring social harmony

Reproductive division of labour is a characteristic trait of social insects. The dominant reproductive individual, often the queen, uses chemical communication and/or behaviour to maintain her social status. Queens of many social insects communicate their fertility status via cuticle-bound substances. As these substances usually possess a low volatility, their range in queen-worker communication is potentially limited. Here, we investigate the range and impact of behavioural and chemical queen signals on workers of the ant Temnothorax longispinosus. We compared the behaviour and ovary development of workers subjected to three different treatments: workers with direct chemical and physical contact to the queen, those solely under the influence of volatile queen substances and those entirely separated from the queen. In addition to short-ranged queen signals preventing ovary development in workers, we discovered a novel secondary pathway influencing worker behaviour. Workers with no physical contact to the queen, but exposed to volatile substances, started to develop their ovaries, but did not change their behaviour compared to workers in direct contact to the queen. In contrast, workers in queen-separated groups showed both increased ovary development and aggressive dominance interactions. We conclude that T. longispinosus queens influence worker ovary development and behaviour via two independent signals, both ensuring social harmony within the colony.

How the social parasitic bumblebee Bombus bohemicus sneaks into power of reproduction

Social parasitism is widespread in many groups of social living hymenopteran species and has also evolved in the genus Bombus. Cuckoo bumblebees (subgenus Psithyrus) are obligate brood parasites in nests of other bumblebee species. After nest usurpation and the killing of the host queen, the parasite female has to control worker reproduction in order to accomplish and maintain reproductive dominance and to ensure her reproductive success. The aim of our study was to examine whether the generalist parasitic bumblebee Bombus bohemicus monopolizes and prevents worker reproduction by physical or chemical means and to identify possible odor compounds involved therein. We performed bioassays with callow workers of the host Bombus terrestris and have shown that B. bohemicus females are able to suppress host worker ovarian development, when these host workers are under the direct influence of the parasite female. Furthermore, by chemical analyses, we have demonstrated that the parasite females adjust to the odor profiles of their host queens in order to maintain the level of fertility signaling inside the host colony although the host queen is absent. We also found that host workers change their odor profile after nest usurpation by the parasite female and consequently, we suggest that the host and parasite are caught up in a chemical arms race.

Reproductive plasticity in Polistes paper wasp workers and the evolutionary origins of sociality

Regulatory pathways in solitary species provide the raw materials for the evolution of sociality. Therefore, comparing the mechanisms that mediate reproductive plasticity in social species and their solitary ancestors can provide insight into the evolutionary origin of sociality. In many solitary insects, the effect of juvenile hormone (JH) on fertility is mediated through the fat body; individuals in good physical condition show a stronger fertility response to JH than individuals in poor physical condition. Here, we test whether a similar, condition-dependent JH response mediates fertility in workers of the primitively eusocial Polistes dominulus wasps. We test how body weight, JH, and adult nutrition influence worker ovarian development. Both JH-treatment and adult nutrition dramatically increased ovarian development. Body weight also influenced ovarian development, as large workers developed more eggs than smaller workers. Body weight and fat are strongly linked in P. dominulus workers, so these results suggest that the fat-dependent JH responsiveness common in solitary insects is conserved in social wasps. The simple, ancestral relationship between reproductive investment and physical condition may facilitate cooperation by allowing workers to adaptively allocate energy into reproduction based on their probability of successfully becoming a queen.

Caste-specific cuticular lipids in the stingless beeFriesella schrottkyi

While a queen control pheromone complex that inhibits worker ovary development has been described for honey bees, no comparable control pheromones have been identified for their sister group, the stingless bees. The aim of the present work was to search for possible control pheromones in the stingless bee Friesella schrottkyi. No volatile substances were found in the heads of queens that might serve as queen control pheromones. On the other hand, distinct differences were found between the cuticular substances of queens and workers. The major hydrocarbons were different between the two castes, and while queens contained methyl-branched alkanes and no unsaturated hydrocarbons, workers contained alkenes and alkadienes but no methyl branched hydrocarbons. Colonies deprived of a queen produced laying workers. Differences were observed in the cuticular patterns of laying workers and workers from a queen controlled colony.

A scientific note on E-β-ocimene, a new volatile primer pheromone that inhibits worker ovary development in honey bees

In social insects, larvae are completely dependent on adults to complete their development. In honey bees (Apis mellifera L.), larval development requires that larvae engage workers in nursing tasks rather than allocating energy in other activities (Le Conte and Hefetz, 2008). Indeed, in the absence of any constraint, workers tend to become active reproductive individual. The queen inhibits worker ovary development via pheromones (Hoover et al., 2003; Wossler and Crewe, 1999), but the inhibition of worker ovary development seems superior when bees are exposed to unsealed brood (Kropacova and Haslbachova, 1970, 1971). Components of the brood pheromone have already been identified and consist of a blend of 10 esters (Le Conte et al., 1989, 1990). Two of these low volatility compounds have been demonstrated to partially decrease worker ovary development (Mohammedi et al., 1998; Pankiw and Garza, 2007). Thus, we asked whether brood emits volatile compounds that could also have an effect on worker ovary activation. We identified a new highly volatile molecule from the larvae E-β-ocimene, that inhibits worker ovary maturation. We used Solid Phase Microextraction (SPME) (65 μm Carbowax) to sample volatiles emitted by different larval instars. We analyzed 20 larvae at stage 1, 2–3 or 10 larvae at stage 4–5. Larvae were kept in a 15 mL closed vial for 20 min in an incubator at 34 ◦C and 50% humidity (N = 8,

The role of the queen mandibular gland pheromone in honeybees (Apis mellifera): honest signal or suppressive agent?

Queen pheromones interfere with worker reproduction in social insects. However, there is still an unresolved question as to whether this pheromone acts as an “honest” signal for workers, giving a reliable indication of the queen’s reproductive value, or as a suppressive agent, inhibiting worker reproduction independent of the queen’s reproductive capacity. In honeybees (Apis mellifera), the queen’s mandibular gland secretion, a mix of fatty acids and some aromatic compounds, is crucial for regulating the reproductive division of labor in the colony inhibiting ovary development in workers. We quantified the mandibular gland secretions of virgin, drone-laying, and naturally mated queens using gas chromatography to test whether the queens’ mating, ovary activation, or the reproductive value for workers correlated with the composition of the secretion. Although the absolute amounts of the “queen substance” 9-oxo-2(E)-decenoic acid (9-ODA) were similar among the three groups, the proportions of 9-ODA decreased with increasing reproductive quality. Furthermore, the ratios of queen to worker compounds were similar in all three treatment groups, irrespective of the reproductive capacity. A multivariate analysis including all six compounds could not separate drone-laying queens from naturally mated ones, both with active ovaries but only the latter ensuring colony survival. We suggest that the mandibular gland pheromones are unlikely to function as reliable indicators of queen reproductive value and rather operate as an agent to suppress worker reproduction. This does not exclude the possibility that other “honest” pheromone signals exist in the honeybee colony, but these would have to arise from other semiochemicals, which could be produced by both the queen and the brood.