Queenless Colonies Research Articles

Rebel honeybee workers have a tendency to become intraspecific reproductive parasites.

Worker honeybees may reproduce in either their own or foreign colonies; the latter situation is termed intraspecific reproductive parasitism (IRP). In this study, we compared the tendency for IRP between normal honeybee workers, which are characterized by a relatively low reproductive potential, and “rebel workers”, a recently discovered subcaste of honeybee workers characterized by a high reproductive potential that develops when the colony is without a queen. We expected that the high reproductive potential of the rebel workers would influence their reproductive strategy and that these individuals would drift to other colonies to lay eggs more often than normal workers. The results confirm our expectations and show that rebel workers are more likely than normal workers to drift to foreign colonies. The rebel workers also preferred to drift to queenless colonies than to queenright colonies, while the normal workers did not show this preference. This study indicates that rebel workers have a tendency for IRP, which may be responsible for the maintenance of the rebel worker strategy in bee populations.

Control of mandibular gland pheromone synthesis by alternative splicing of the CP-2 transcription factor gemini in honeybees (Apis mellifera carnica)

The honeybee queen’s mandibular gland pheromones (QMP) are essential for the suppression of worker reproduction. Worker ovary activation is regulated by alternative splicing of a CP2-transcription factor named gemini. Since workers with activated ovaries also produce QMP in their mandibular glands, we tested whether alternative splicing of gemini also controls mandibular gland pheromone biosynthesis in workers using RNA interference. Altering the splice pattern of gemini resulted in enhanced levels of the queen-specific components of the mandibular gland pheromone in queenless honeybee workers, suggesting that gemini functions as a pleiotropic regulatory switch influencing both ovary activation and resulting in QMP synthesis in workers. Because the QMP produced by these workers suppresses ovary activation in other workers, gemini seems to be a key regulatory gene affecting reproductive hierarchies among workers in queenless colonies.

Queen mandibular pheromone modulates hemolymph ecdysteroid titers in adult Apis mellifera workers

To test the hypothesis that exposure to queen mandibular pheromone (QMP) modulates ecdysteroid production in adult worker honey bees, ecdysteroids were measured in hemolymph and other tissues of individual adult worker honey bees reared with or without QMP in cages and field colonies. Ecdysteroid titers were higher in caged workers exposed to QMP continuously from the first day of adult life than in workers reared without QMP. Statistical cluster analysis suggested the possibility that a subgroup of workers (“responders”) is more sensitive to QMP in this regard than other workers. In 12-day-old workers, ecdysteroid titers in workers reared in queenright (QR) colonies were similar to those observed in cages with QMP, but lower than those in queenless (QL) colonies. Differences in number of ovarioles or degree of ovarian activation did not correlate with hemolymph ecdysteroids. Previous studies have demonstrated ecdysteroids in hemolymph in very young adult workers and in workers in QL colonies; the present study indicates that production of ecdysteroids occurs in older adult worker honey bees in the absence of morphological signs of ovarian activation, with cage studies revealing a modulatory role for QMP masked in the complex environment of the hive.

Reproduction of rebel workers in honeybee (Apis mellifera) colonies

The honeybee is one of several eusocial species in which the queen is typically the only reproductive member of the colony; worker reproduction is mostly restricted to queenless colonies. Because workers cannot mate, they lay unfertilized eggs, which develop into males. A recent study showed that in queenless colonies, which arise after swarming, worker larvae develop into rebel workers that have greater reproductive potential than do workers reared in queenright colonies, as measured by the number of ovarioles and degree of ovary activation. However, there was no evidence that rebels had an opportunity to produce male offspring. Here, we show for the first time that rebel workers not only activate their ovaries but also produce significantly more male offspring in queenright colonies than do normal workers. Moreover, our results show that the level of rebel reproduction in queenright colonies is similar to the reproduction of normal workers in queenless colonies. This finding suggests that the ultimate factor favouring the evolution of the rebel strategy is the decrease in relatedness between the old-generation workers and the new queen’s offspring that occurs after queen exchange at swarming.

Auxiliary brood cell construction in nests of the stingless bee Plebeia lucii (Apidae: Meliponini)

Queen production in stingless bees with fusion of neighboring brood cells occurs by the perforation of the adjacent brood cell or construction of an auxiliary one. This study describes the auxiliary brood cell building behavior in queenless colonies of Plebeia lucii. Queenright and queenless (orphan) colonies were monitored, and auxiliary cell construction was video-recorded in orphan colonies. Brood cells with auxiliary cells added were analyzed with X-rays to identify the amount of food and the larval behavior into the brood cells. Plebeia lucii had specific behavioral sequence in auxiliary cell building. The addition of auxiliary cells is the main strategy to produce queens in P. lucii, mainly for the production of emergency queens in orphan colonies because queen absence triggered a high production of auxiliary cells. X-ray analyses showed that auxiliary cell addition occurred when the food in the larval brood cells had been completely eaten and showed changes in dorsoventral position of the larvae. Larvae of males did not perforate auxiliary cells, indicating that sex-related factors affect this behavior. The wax handling by workers and the fused thin and concave-shaped wall between the auxiliary and larval brood cells seems to facilitate wall perforation by the larvae.

Levels of selection shaping caste interactions during queen replacement in the honey bee, Apis mellifera

We investigated the different levels of selection that may influence the queen replacement process in honey bees by monitoring queen–queen and worker–queen interactions in queenless observation colonies containing populations of paint-marked workers that were either related or not related to introduced virgin queens experimentally reared to be of low- and high-quality, as estimated by differences in size and potential reproductive capacities. The high-quality queens were more likely to survive and become the new laying queen of their colonies. Queen survival was positively associated with both queen fighting ability and worker–queen interactions. Surviving queens were more aggressive and had greater fighting success. However, surviving queens were not bigger than killed queens nor were they the first queens to emerge, suggesting that queen size and early emergence alone are not the determining factors of queen fighting ability. The worker–queen interaction that was most strongly associated with the outcome of queen replacement was the vibration signal, which is a communication signal that workers perform on virgin queens. Surviving queens were vibrated at rates 3–4 times those experienced by killed queens and a queen’s vibration rate was positively correlated with her fighting ability and the number of rivals killed. Workers showed no consistent preferences for related vs. unrelated queens and the proportion of interactions received from related workers was not associated with any aspect of queen fighting ability and success monitored. Our results suggests that caste interactions during queen replacement have been shaped by both selection acting at the level of the individual queen (which favors higher quality queens with greater fighting ability) and selection acting at the level of the colony (which favors workers directing vibration signals towards queens with greater fighting potential), ultimately resulting in a higher quality queen becoming the new laying queen of the colony. Selection acting at the level of the individual worker through kin selection, which would favor preferential treatment of related queens, did not consistently influence caste interactions or the outcome of the replacement process. Thus, the outcome of queen replacement in honey bees may be determined primarily by a combination of a queen’s inherent fighting ability coupled with the rate at which she receives some interactions (particularly vibration signals) from workers.

Correction to ‘The role of the glucose-sensing transcription factor carbohydrate-responsive element-binding protein pathway in termite queen fertility’

Termites are among the few animals that themselves can digest the most abundant organic polymer, cellulose, into glucose. In mice and Drosophila, glucose can activate genes via the transcription factor carbohydrate-responsive element-binding protein (ChREBP) to induce glucose utilization and de novo lipogenesis. Here, we identify a termite orthologue of ChREBP and its downstream lipogenic targets, including acetyl-CoA carboxylase and fatty acid synthase. We show that all of these genes, including ChREBP, are upregulated in mature queens compared with kings, sterile workers and soldiers in eight different termite species. ChREBP is expressed in several tissues, including ovaries and fat bodies, and increases in expression in totipotent workers during their differentiation into neotenic mature queens. We further show that ChREBP is regulated by a carbohydrate diet in termite queens. Suppression of the lipogenic pathway by a pharmacological agent in queens elicits the same behavioural alterations in sterile workers as observed in queenless colonies, supporting that the ChREBP pathway partakes in the biosynthesis of semiochemicals that convey the signal of the presence of a fertile queen. Our results highlight ChREBP as a likely key factor for the regulation and signalling of queen fertility.

Reproductive Performance of Laying Worker of Apis cerana in Queenless Colonies

The Asian cavity-nesting honey bee, Apis cerana, the workers ovaries is very active in queenless colonies. We investigated the duration of egg laying by laying workers, egg-laying behavior, determine egg load in ovaries, estimate oviposition rate nad number of eggs per cell in three queenless colonies of Korean A. cerana. The number of eggs laid by laying workers at different times on the manipulated comb was checked and counted. We observed that the laying workers spent, on average, 109.2 seconds per cell. During oviposition, the laying workers showed two types of behaviors, viz; a still phase, and a recovery phase. On average, 4.0 of worker eggs per cell per week was recorded. The workers laid three different shapes of eggs in a cell. The laying workers ovaries retained 1 to 4 mature eggs. This suggests that a laying worker may lay from one to four eggs per oviposition. We did not observe false-queen in Korean A. cerana in three queenless colonies. In this study, we reported for the first time that one worker cell of A. cerana retains up to 33 eggs in queenless condition. The queenless colonies build the comb and lay eggs in new comb, and also reared the new queen without successful development of the queen larva from laying worker eggs.

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.

Honeybee worker larvae perceive queen pheromones in their food

Honeybee workers develop from fertilised eggs, but those reared in a queenless colony develop into ‘rebel’ workers, which are more queen-like than typical workers. Rebels develop after an old queen leaves with a swarm and before a new queen hatches. We hypothesised that larval food lacking queen mandibular pheromones trigger the rebel phenotype. Larvae reared under queenright or queenless conditions were additionally fed with water or a drop of macerated queen mandibular glands. After following development of the bees and subjecting them to dissection, we found that those reared with a queen or fed the macerated glands under queenless conditions developed into typical workers. Only those workers reared without a queen and without macerated glands added to their food developed into rebels; these rebels had more ovarioles, smaller hypopharyngeal glands, and larger mandibular and Dufour’s glands than did typical workers. This is the first evidence that larval perception of the presence or absence of queen pheromones causes an alternative development strategy.

Effect of Virgin Queens Storage on Their Survival Rate, Attractiveness and Acceptance by the Honey Bee Colonies

Virgin honeybee queens, were held in the centre of brood nest of both queenless and queenright strong colonies using two types of cages and fed by workers through a wire screen holes, emerging cage (EC) with two wire screen sides and Benton cage (BC) with one side of wire screen. Mean survival rate of stored queens in both cage types were not differ significantly. Storage virgin queens within queen-right colony was influenced reversely by the presence of colony laying queen as the survival rate was significantly lower (68.0%) than queens stored in queenless colonies (77.4%). The attractiveness of introduced virgin queens 30 days old to workers increased when the duration of queenlessness increase from 1 to 7 days and also, the acceptance percentage of them. The number of workers attracted by virgin queens was increased with the storage periods (3, 15 and 30days) as the lowest significant number was for the 3 days period, while there were no significant differences between the rest of periods. The younger and older queens were most significantly accepted than the intermediate ones. Workers in the 5 days queenless nuclei were more attracted and easily accepted the introduced virgin queens than those in similar queenless strong colonies.

The role of the glucose-sensing transcription factor carbohydrate-responsive element-binding protein pathway in termite queen fertility.

Termites are among the few animals that themselves can digest the most abundant organic polymer, cellulose, into glucose. In mice and Drosophila, glucose can activate genes via the transcription factor carbohydrate-responsive element-binding protein (ChREBP) to induce glucose utilization and de novo lipogenesis. Here, we identify a termite orthologue of ChREBP and its downstream lipogenic targets, including acetyl-CoA carboxylase and fatty acid synthase. We show that all of these genes, including ChREBP, are upregulated in mature queens compared with kings, sterile workers and soldiers in eight different termite species. ChREBP is expressed in several tissues, including ovaries and fat bodies, and increases in expression in totipotent workers during their differentiation into neotenic mature queens. We further show that ChREBP is regulated by a carbohydrate diet in termite queens. Suppression of the lipogenic pathway by a pharmacological agent in queens elicits the same behavioural alterations in sterile workers as observed in queenless colonies, supporting that the ChREBP pathway partakes in the biosynthesis of semiochemicals that convey the signal of the presence of a fertile queen. Our results highlight ChREBP as a likely key factor for the regulation and signalling of queen fertility.

Intraspecific worker parasitism in the common wasp, Vespula vulgaris

Insect societies display a remarkable level of cooperation, but their colonies also represent a valuable resource that can be taken advantage of by genetically unrelated individuals. Indeed, several recent studies have documented cases of intraspecific reproductive parasitism, whereby workers penetrate and lay eggs in unrelated colonies in order to have their brood raised by the host workers. Previously, it has been predicted that queenless colonies should be a prime target of such intraspecific worker parasitism, as in such colonies the parasite workers would be able to reproduce without interference from either the queen or other workers. So far, this prediction has been supported with data from the honeybee, but evidence from other social insect groups is currently lacking. Here we present the first such test in the common wasp, Vespula vulgaris. In particular, workers from queenright colonies left the natal nest at a higher rate than those from queenless colonies. However, contrary to our predictions, drifter workers targeted queenless and queenright colonies equally. Chemical data suggest that this lack of discrimination may be linked to recognition constraints and the lack of volatile signals that reliably indicate the presence or absence of the queen. In addition, in queenright colonies, drifters activated their ovaries at a rate that was ca. five times higher than the natal workers. Overall, our results suggest that also in wasps, workers can gain inclusive fitness by drifting to unrelated nests, even if the chances of successfully reproducing there may be very slim.

Physiology of reproductive worker honey bees (Apis mellifera): insights for the development of the worker caste.

Reproductive and behavioural specialisations characterise advanced social insect societies. Typically, the honey bee (Apis mellifera) shows a pronounced reproductive division of labour between worker and queen castes, and a clear division of colony roles among workers. In a queenless condition, however, both of these aspects of social organisation break down. Queenless workers reproduce, forage and maintain their colony operating in a manner similar to communal bees, rather than as an advanced eusocial group. This plasticity in social organisation provides a natural experiment for exploring physiological mechanisms of division of labour. We measured brain biogenic amine (BA) levels and abdominal fat body vitellogenin gene expression levels of workers in queenright and queenless colonies. Age, ovary activation and social environment influenced brain BA levels in honey bees. BA levels were most influenced by ovary activation state in queenless bees. Vitellogenin expression levels were higher in queenless workers than queenright workers, but in both colony environments vitellogenin expression was lower in foragers than non-foragers. We propose this plasticity in the interacting signalling systems that influence both reproductive and behavioural development allows queenless workers to deviate significantly from the typical worker bee reaction norm and develop as reproductively active behavioural generalists.

Effects of age and Reproductive Status on Tergal Gland Secretions in Queenless Honey bee Workers, Apis mellifera scutellata and A. m. capensis

Secretions from tergal glands are part of a queen's pheromonal control of worker reproduction in honey bees. However, in queenless honey bee colonies, workers compete to gain pheromonal, and hence reproductive dominance, over nestmates with ontogenetic changes in their glandular secretions that affect the behavioral or physiological responses of other individuals. Using gas chromatography/mass spectrometry, we investigated for the first time the age-dependent changes in tergal gland secretions of queenless workers of the clonal lineage of Apis mellifera capensis and workers of A. m. scutellata. The reproductive status of honey bee workers was determined by recording the presence of spermathecae and the level of ovarian activation. The tergal gland chemicals identified in both A. m. scutellata workers and A. m. capensis clone workers were oleic acid, n-tricosene, n-pentacosene, and n-heptacosene, with three additional compounds, palmitic acid, n-heneicosene, and n-nonacosene, in A. m. capensis clones. We report ethyl esters as new compounds from honey bee worker tergal gland profiles; these compounds increased in amount with age. All A. m. capensis clone workers dissected had spermathecae and showed ovarian activation from day 4, while ovarian activation only started on day 7 for A. m. scutellata workers that had no spermathecae. Tergal gland secretions were present in higher quantities in bees with activated, rather than inactive ovaries. This suggests that tergal gland secretions from reproductive workers could act as releaser and primer pheromones in synergy with other glandular compounds to achieve pheromonal and reproductive dominance.

Social Control of Reproduction and Breeding Monopolization in the Eusocial Snapping Shrimp Synalpheus elizabethae.

Understanding why individuals within altruistic societies forgo reproduction to raise others' offspring has fascinated scientists since Darwin. Although worker polymorphism is thought to have evolved only in sterile workers, worker subcastes appear to be common among social invertebrates and vertebrates. We asked whether sterility accompanies eusociality and morphological differentiation in snapping shrimps (Synalpheus)-the only known marine eusocial group. We show that workers in Synalpheus elizabethae are reproductively totipotent and that female-but not male-gonadal development and mating are mediated by the presence of a queen, apparently without physical aggression. In queenless experimental colonies, a single immature female worker typically became ovigerous, and no female workers matured in colonies with a resident queen. Thus, eusocial shrimp workers retain reproductive totipotency despite signs of morphological specialization. The failure of most female workers to mature is instead facultative and mediated by the presence of the queen, ensuring her reproductive monopoly.

Associations between reproduction and work in workers of the Asian hive bee Apis cerana

If a honey bee (Apis spp.) colony becomes queenless, about 1/3 of young workers activate their ovaries and produce haploid male-producing eggs. In doing so queenless workers maximize their inclusive fitness because the normal option of vicarious production of relatives via their queen’s eggs is no longer available. But if many workers are engaged in reproduction, how does a queenless colony continue to feed its brood and forage? Here we show that in the Asian hive bee Apis cerana hypopharyngeal gland (HPG) size is larger in queenless workers than in queenright workers and that bees undertaking brood-rearing tasks have larger HPG than same-aged bees that are foraging. In queenless colonies, workers with a smaller number of ovarioles are more likely to have activated ovaries. This reinforces the puzzling observation that a large number of ovarioles reduces reproductive success in queenless A. cerana. It further suggests that reproductive workers either avoid foraging or transition to foraging later in life than non-reproductive workers. Finally, our study also showed that ovary activation and larger-than-average numbers of ovarioles had no statistically detectable influence on foraging specialization for pollen or nectar.

Age at which larvae are orphaned determines their development into typical or rebel workers in the honeybee (Apis mellifera L.).

In the honeybee, diploid larvae fed with royal jelly develop into reproductive queens, whereas larvae fed with royal jelly for three days only and subsequently with honey and pollen develop into facultatively sterile workers. A recent study showed that worker larvae fed in a queenless colony develop into another female polyphenic form: rebel workers. These rebel workers are more queenlike and have greater reproductive potential than normal workers. However, it was unclear whether larvae orphaned at any time during their feeding period can develop into rebels. To answer this question, the anatomical features of newly emerged workers reared in queenless conditions at different ages during the larval period were evaluated. Our results showed that larvae orphaned during the final four or more days of their feeding life develop into rebel workers with more ovarioles in their ovaries, smaller hypopharyngeal glands, and larger mandibular and Dufour’s glands compared with typical workers with low reproductive potential that were reared with a queen or orphaned at the third to last or a later day of feeding life.

The effect of queen pheromone status on Varroa mite removal from honey bee colonies with different grooming ability.

The objective of this study was to assess the effects of honey bees (Apis mellifera L.) with different grooming ability and queen pheromone status on mortality rates of Varroa mites (Varroa destructor Anderson and Trueman), mite damage, and mortality rates of honey bees. Twenty-four small queenless colonies containing either stock selected for high rates of mite removal (n = 12) or unselected stock (n = 12) were maintained under constant darkness at 5 °C. Colonies were randomly assigned to be treated with one of three queen pheromone status treatments: (1) caged, mated queen, (2) a synthetic queen mandibular pheromone lure (QMP), or (3) queenless with no queen substitute. The results showed overall mite mortality rate was greater in stock selected for grooming than in unselected stock. There was a short term transitory increase in bee mortality rates in selected stock when compared to unselected stock. The presence of queen pheromone from either caged, mated queens or QMP enhanced mite removal from clusters of bees relative to queenless colonies over short periods of time and increased the variation in mite mortality over time relative to colonies without queen pheromone, but did not affect the proportion of damaged mites. The effects of source of bees on mite damage varied with time but damage to mites was not reliably related to mite mortality. In conclusion, this study showed differential mite removal of different stocks was possible under low temperature. Queen status should be considered when designing experiments using bioassays for grooming response.

Consumption of tyrosine in royal jelly increases brain levels of dopamine and tyramine and promotes transition from normal to reproductive workers in queenless honey bee colonies

Dopamine (DA) and tyramine (TA) have neurohormonal roles in the production of reproductive workers in queenless colonies of honey bees, but the regulation of these biogenic amines in the brain are still largely unclear. Nutrition is an important factor in promoting reproduction and might be involved in the regulation of these biogenic amines in the brain. To test this hypothesis, we examined the effect of oral treatments of tyrosine (Tyr; a common precursor of DA, TA and octopamine, and a component of royal jelly) in queenless workers and quantified the resulting production of biogenic amines. Tyrosine treatments enhanced the levels of DA, TA and their metabolites in the brain. Workers fed royal jelly had significantly larger brain levels of Tyr, DA, TA and the metabolites in the brains compared with those bees fed honey or sucrose (control). Treatment with Tyr also inhibited the behavior of workers outside of the hive and promoted ovarian development. These results suggest that there is a link between nutrition and the regulation of DA and TA in the brain to promote the production of reproductive workers in queenless honey bee colonies.