Summary. In the honeybee, Dufour’s gland secretion is caste specific and constitutes a component of the multi-sourced queen signal. As predicted, it is attractive to workers, which form a retinue around the scented source. Bioassays reveal the ester fraction and not the hydrocarbons to be the active con- stituents. This function of the esters was corroborated by as- says with the synthetic queen-esters mixture, which success- fully mimicked the queen’s secretion. As predicted from the queen-like secretion exhibited by egg-laying workers, their glandular secretion was also attractive to nestmates, albeit to a lesser degree than that of the queen; while that of non-egg- laying workers was totally inactive. The evolution of the multiple queen signals in honeybees can be regarded as a component in an arms race between queen and workers. We hypothesize that in response to a re- duced sensitivity to a certain queen signal, queen honeybees were selected to develop an alternative signaling-source. Du- four’s gland seems to be one of these sources. Key words: Dufour’s gland, esters, honeybees, Apis mellifera, attraction. Introduction Queen specific signal in the honey bee includes the Queen Mandibular Pheromone (QMP) (Barbier and Lederer, 1960; Barbier, 1986; Breed et al., 1992; Winston and Slessor 1998), a tergite pheromone (Wossler and Crewe, 1999), a fecal pheromone (Page et al., 1988), Dufour’s gland pheromone (Katzav-Gozansky et al., 1997), and possibly other yet unidentified pheromones. While it is conventional to attribute a certain queen-worker interaction to specific pheromones, honeybee communication is not characterized by such sim- plicity. One pheromone can possess a variety of functions, while many activities can be affected by a combination of sev- eral pheromones. Moreover, a single pheromone can act con- comitantly as both releaser and primer. In addition to the queen primer pheromone effect on workers, a typical queen releaser effect in workers is the formation of a retinue or royal court around the reproductive queen (Velthuis, 1985). Using this retinue behavior the full composition of the QMP was identified (Slessor et al., 1988) and the attractivity toward queens tergal glands and Dufour’s glands was demonstrated (Wossler and Crewe, 1999; Katzav-Gozansky et al., 2001, res- pectively). Although the attractivity of the honeybee queen was studied intensively, there are many aspects that are still elusive. It became evident as early as 1954 that the mandibu- lar glands were not the only source of pheromone production (Butler, 1954; Butler et al., 1973). Mated queens from which the mandibular glands were removed were still fully accepted by their respective colonies and successfully headed their colony for a significant period (Velthuis, 1970). This suggests that other queen pheromones produce important cues for worker honeybees, and that they can replace the QMP. One of the caste-specific glandular sources found in the honeybee is the Dufour’s gland (Katzav-Gozansky et al., 1997). While the exudates of workers are composed of a se- ries of odd n-alkanes, the glandular exudates of queens are ad- ditionally fortified with wax-type esters. It was also found that queen Dufour’s gland secretion is attractive to workers, raising the possibility that this fraction of the glandular se- cretion acts as a queen signal (Katzav-Gozansky et al., 2001). Worker-attraction towards the glandular secretion of virgin queens was also demonstrated by Abdalla and Cruz-Landim (2001). Both in vivo and in vitro studies have further demon- strated that ester biosynthesis in Dufour’s gland is not a caste- fixed phenomenon. Queenless (QL) workers that start to de- velop ovaries also biosynthesize the queen-type esters. Moreover, glands from queenright (QR) nurses incubated in vitro also produce these esters, after a certain delay (Katzav- Gozansky et al., 1997). Occasionally, under QL conditions some workers attract a small but recognizable retinue, pre- sumably by exuding some components of the queen pheromone. Such workers are designated as false queens (Crewe and Velthuis, 1980). Since Dufour’s esters seem to dif- fer significantly between QR workers and egg-laying workers, Insectes Soc. 50 (2003) 20 – 23 0020-1812/03/010020-04 © Birkhäuser Verlag, Basel, 2003 Insectes Sociaux Research article Honeybee egg-laying workers mimic a queen signal T. Katzav-Gozansky 1 , V. Soroker 2 , W. Francke 3 and A. Hefetz 1 1 Department of Zoology, George S. Wise Faculty of Life Sciences, Tel Aviv University, Ramat Aviv, Tel Aviv, Israel, e-mail: katzavt@post.tau.ac.il 2 Department of Entomology, Institute of Plant Protection, Agricultural Research Organization, The Volcani Center, Beit Dagan, Israel 3 Institut für Organische Chemie, Universität Hamburg, Hamburg, Germany Received 2 May 2002; revised 31 July 2002; accepted 12 August 2002.