A Closer Look
Queen’s Court
By: Clarence Collison
When the queen is stationary on the comb she is surrounded by a circle of workers known as the “court” or “retinue” that face toward her and feed, palpate and lick her. Usually, the retinue is composed of eight or 10 workers. The functions of the workers making up the queen’s court (retinue) are to feed her, groom her, remove her waste and to lick pheromones from her body. The queen rarely, if ever, feeds herself. Secretions from mandibular glands and hypopharyngeal glands form the proteinaceous jelly that is fed to queens. The amount the queen is fed by adult workers in her court is related to the queen’s egg laying rate. “The queen produces QRP (Queen Retinue Pheromone) which contains QMP (Queen Mandibular Pheromone) and at least four additional pheromones. QRP elicits retinue behavior from younger workers, who touch, lick and groom the queen. In the process they acquire QMP and the other queen pheromones, which are then distributed throughout the colony” (Schneider, 2015). Queen mandibular pheromone is a blend of five different pheromones produced by the mandibular glands (9–oxo-trans-2–decenoic acid [9-ODA], ± 9-hydroxydec-2-enoic acid [+/- 9-HAD], methyl-p-hydroxybenzoate [HOB] and 4-hydroxy-3-methyoxyphenylethanol [HVA]).
The queen mandibular pheromone (QMP), consisting of the five synergistic components listed previously, is the only pheromone chemically identified in the honey bee queen, but this pheromone does not fully duplicate the pheromonal activity of a full queen extract. To identify the remaining unknown compounds for retinue attraction, honey bee colonies were selectively bred to have low response to synthetic QMP and high response to a queen extract in a laboratory retinue bioassay. Workers from these colonies were then used in the bioassay to guide the isolation and identification of the remaining active components. Four new compounds were identified from several glandular sources that account for the majority of the difference in retinue attraction between synthetic QMP and queen extract: methyl (Z)-octadec-9-enoate (methyl oleate), (E)-3-(4-hydroxy-3-methoxyphenyl)-prop-2-en-1-ol (coniferyl alcohol), hexadecan-1-ol and (Z9,Z12,Z15)-octadeca-9,12,15-trienoic acid (linolenic acid). These compounds were inactive alone or in combination, and they only elicited attraction in the presence of QMP. There was still unidentified activity remaining in the queen extract. The queen therefore produces a synergistic, multiglandular pheromone blend of at least nine compounds for retinue attraction, the most complex pheromone blend known for inducing a single behavior in any organism (Keeling et al., 2003).
Queen attendance behavior of workers from selected honey bee colonies with high and low worker retinue response to synthetic queen mandibular gland pheromone (QMP) was investigated. Antennating, licking, grooming and feeding of the queen by workers from high and low responding colonies were examined. High and low QMP responding workers did not attend the queen differently. However, workers originating from different colonies antennated and licked the queen more frequently than others, suggesting there may be a genetic basis for queen attendance behavior not necessarily associated with response to QMP. The median age of queen attendance was independent of strain (Pankiw et al., 1995).
The behavior of queen and worker honey bees was observed using small colonies in observation hives. Workers paid more attention to queens which had been mated for two months or more than to those which were newly mated; virgin queens received the least attention. Queens received most attention when they were stationary and least when they were walking over the comb; virgin queens were most active. The queen pheromone component 9–oxo-trans-2–decenoic acid stimulated ‘court’ behavior when presented on small polyethylene blocks, but workers responded aggressively to complete extracts of queens’ heads. Both the heads and abdomens of mated queens received much attention from court workers, but the abdomens were palpated by more workers for longer and were licked much more. The queens’ thoraces were least attended. Abdominal tergites posterior to tergite glands were licked for longer than those anterior to the glands. Only worker bees very near to the queen reacted to her and joined her ‘court’. No evidence was found of a diel periodicity in the behavior of a queen or her ‘court’. During the Winter the queen’s court was smaller than in Summer and she walked less and laid fewer eggs. When colonies were fed with sucrose syrup in Winter, their queens laid more eggs and workers reared more brood but there was no change in the attention received by the queens (Free et al., 1992).
Allen (1960) investigated the behavior of the queen’s attendants. One hundred ninety-six individually marked bees were seen to examine the queen and the durations of their visits were recorded. All gradations of behavior varying from an apparent desire to avoid the queen to a strong desire to remain with her were found. The behavior of individual bees varied on different visits, but no regular sequence could be found. More than half the total number of visits had a duration of 30 seconds or less while the remainder were longer than 30 seconds, with a maximum of 41 minutes. About 8% of the visits were very short and terminated abruptly and with signs of alarm, but no reason for this type of behavior was apparent. The ages of the attendants varied from a few hours to 52 days but no obvious difference in age-distribution between the attendants remaining for very short periods and those remaining for longer was found. In relation to the total numbers of marked bees of different ages present, the proportion of workers acting as attendants showed no systematic variations, although in a colony where the average age was low (as in Summer), the actual numbers of young attendants would apparently greatly exceed those of the older ones. The age-range of attendants licking the queen was similar to the overall age-range of the attendants. The ages of the attendants feeding the queen in two colonies ranged from one day to 23 days. No bee was seen to feed the queen during the day of its emergence, although the greatest numbers recorded occurred in the next few days of adult life. No systematic changes were apparent in the duration of feeds given by bees of different ages, the mean value for two colonies being 44 seconds. Observations made during the season of active brood-rearing up to swarming time (April-July) showed that the number and duration of feeds supplied to the queen tended to rise until two to three weeks before the swarm left, but in this latter period both number and duration decreased, approximately from the time that queen cell formation started. Thus, the queen received gradually diminishing quantities of food in the period of swarm preparation. Egg-production over the same period showed a similar trend.
In another study, workers gathered on cages containing a mated laying queen placed above a normal queenright colony. Fewer workers gathered on a cage when they could not touch the caged queen, or when she was dead, or when her mandibular glands had been removed. The number of workers on the cage was correlated with the amount of 9-oxodecenoic acid (primary component of queen mandibular pheromone) in the queen’s head. Workers gathered on cages containing only 9-oxodecenoic acid on filter-paper that the bees could not touch. ‘Court’ formation round a free queen was not diminished by removing her mandibular glands. The head of queens whose mandibular glands had been removed contained traces of 9-oxodecenoic acid. Up till now, no unknown substance affecting worker bees’ behavior was detected in the queen’s head. No sounds likely to be specifically communicative were heard from the queen or from workers that formed a ‘court’ round her. Workers did not move from afar to form a ‘court’ round a queen. It is concluded that in an undisturbed colony with a mated laying queen there is no attraction of workers towards the queen except over very short distances (Butler et al., 1973).
Honey bee colonies maintain viable queens in part through communication with queen mandibular pheromone (QMP), a mixture that signals the queen’s presence and reproductive quality to workers. In turn, workers are thought to provide retinue queen care or replace queens partially based on QMP profiles. The effects of seasonal dearth were examined (overwintering in a warm subtropical location) on queen-worker interactions. Retinue worker responses to continuously ovipositing queens were considered in view of QMP signaling and queen reproductive quality. QMP signaling was estimated from QMP residues recovered from nest worker bodies, which is the primary mode of QMP transfer from the queen to the colony at large. QMP residues varied seasonally but not at all with queen reproductive quality (spermatheca sperm storage, ovary protein and lipid contents). 9-HDA and 9-ODA were lower in January than other months. HOB decreased from July to January, while HVA, a component associated with mated queens, increased sharply in January. Despite these seasonal signaling differences, retinue workers attended queens at similar levels through the months. In terms of reproductive quality, queens did not differ over the months in matedness (spermatheca sperm storage) or physiological age (protein carbonyl content) but varied in nutrient allocation to reproductive and non-reproductive tissues. Queen ovaries contained more protein in September than in November, and more lipid in July and September than in November and January. Queen fat bodies had more protein in July than September or November, but less lipid in July and September than November or January. Retinue worker responses did not vary with seasonal QMP changes, but reflected overall continuous brood rearing efforts and queen matedness throughout the year. The absence of seasonal differences in worker responses to QMP should be considered in the broader context of continuous reproductive efforts in warm subtropical colonies (Carroll et al., 2023).
Honey bee queen attendants disperse queen pheromones to supplement pheromone dispersal by direct queen-worker contacts. With time they lose their dispersal function exponentially due mainly to volatilization of queen pheromones carried on their bodies. The elimination of those airborne pheromones together with the air while ventilating the hive is balanced by pheromone release by the queen. This equilibrium results in a certain level of queen pheromones in the broodnest. The change of the pheromone level (for example, due to loss of the colony of its queen) can serve as a signal to alter the behavior of the workers and the state of the colony (Juška et al., 1981).
The substance produced by the mandibular gland complex (HQMC) of queen honey bees is considered to be responsible for retinue formation in honey bees. The retinue response includes the licking and antennating behavior which signals the presence of a dominant reproductive queen and thereby establishes and stabilizes the social fabric of the colony. Despite 25 years of research, none of the known constituents of the mandibular gland complex extract has been effective in eliciting full retinue behavior. We report that the mandibular-gland-based retinue response is mediated by five semio-chemicals. Each component is weakly active alone, but the complete blend imparts activity equivalent to HQMC extract at a level as low as 10–7 of that present in a queen (Slessor et al., 1988).
Ferguson and Free (1980) observations provided strong circumstantial evidence that workers in a queen’s court obtain queen pheromone on their antennae, and that queen pheromone is distributed through the colony during antennal contact between workers. Workers that had just left the court of a mated or virgin queen had an increased tendency to make reciprocated antennal contacts with other workers. This tendency was reinforced when the workers concerned licked the queen in addition to palpating her with their antennae, probably because they spent longer in the court. The first workers contacted by those leaving the court also had an increased tendency to make reciprocated antennal contacts. Antennal contacts were more likely to be initiated by bees other than those from the queen’s court. Workers from the court and those they first contacted participated in food transfer more than did workers selected at random. Workers that licked a virgin queen subsequently participated in food transfer more than those that palpated her with their antennae only. Within about five minutes of leaving the court, a worker’s participation in reciprocated antennal contacts and in food transfer diminished to the level of non-court workers. Workers chosen at random made more transient and non-reciprocated antennal contacts when in a colony with a mated queen than when in a colony with a virgin queen.
The ontogeny of the five queen mandibular gland semiochemicals that initiate and maintain the retinue behavior of worker honey bees was investigated by quantitative splitless capillary gas chromatography. No detectable pheromone is present at the time of eclosion, but decenoic acid levels build up rapidly during the first week of the queen’s life. Two aromatic components attain detectable levels later, with the more plentiful methylp-hydroxybenzoate preceding the 4-hydroxy-3-methoxyphenylethanol. Pheromone levels are maximal in mature, mated, laying queens. The ratio of (R,E)-(−)-9-hydroxy-2-decenoic acid to the (S,E)-(+) enantiomer increases with the age of the queen. Pheromone levels in queen mandibular glands are largely unaffected by queen banking, restraint with workers in mailing cages, and limited storage on dry ice. All major body parts of typical queens, especially the head and legs, have sufficient mandibular exudate to be highly attractive to worker bees (Slessor et al., 1990).
References
Allen, M.D. 1960. The honey bee queen and her attendants. Anim. Behav. 8: 201-208.
Butler, C.G., R.K. Callow, C.G. Koster and J. Simpson 1973. Perception of the queen by workers in the honey bee colony. J. Apic. Res. 12: 159-166.
Carroll, M.J., N.J. Brown, Z. Ruetz, V.A. Ricigliano, and K.E. Anderson 2023. Honey bee retinue workers respond similarly to queens despite seasonal differences in Queen Mandibular Pheromone (QMP) signaling. PLoS ONE 18(9): e0291710
Ferguson, A.W. and J. B. Free 1980. Queen pheromone transfer within honey bee colonies. Physiol. Entomol. 5: 358-366.
Free, J.B., A.W. Ferguson and J.R. Simpkins 1992. The behaviour of queen honey bees and their attendants. Physiol. Entomol. 17: 43-55.
Juška, A., T.D. Seeley and H.H.W. Velthuis 1981. How honey bee queen attendants become ordinary workers. J. Insect. Physiol. 27: 515-519.
Keeling, C.I., K.N. Slessor, H.A. Higo and M.L.Winston 2003. New components of the honey bee (Apis mellifera L.) queen retinue pheromone. Proc. Natl. Acad. Sci. USA 100 (8): 4486-4491.
Pankiw, T., M.L. Winston and K.N. Slessor 1995. Queen attendance behavior of worker honey bees (Apis mellifera L.) that are high and low responding to queen mandibular pheromone. Insect. Soc. 42: 371-378.
Schneider, S.S. 2015. The honey bee colony: life history. In: The Hive And The Honey Bee (Ed. J.M. Graham), Dadant and Sons, Hamilton, IL, pp. 73-109.
Slessor, K.N., L.-A. Kaminski, G.G.S. King, J.H. Borden and M.L. Winston 1988. Semiochemical basis of the retinue response to queen honey bees. Nature 332: 354-356.
Slessor, K.N., L.A. Kaminski, G.G.S. King and M.L. Winston 1990. Semiochemicals of the honey bee queen mandibular glands. J. Chem. Ecol. 16: 851-860.
Clarence Collison is an Emeritus Professor of Entomology and Department Head Emeritus of Entomology and Plant Pathology at Mississippi State University, Mississippi State, MS.
