Reproductive workers are seldom seen in colonies with a laying queen.
Honey bee workers are sexually undeveloped females and under normal hive conditions do not lay eggs. However, workers have small ovaries which under the right conditions can produce eggs which will develop into drone progeny. Since workers are incapable of mating they can only produce unfertilized eggs. Reproductive workers are seldom seen in colonies with a laying queen (Visscher 1989; Page and Erickson 1988) because a suite of pheromones derived from the queen and the brood inhibits ovarian development in workers. If a colony loses its queen and fails to replace her, the queen pheromone will cease to be present and the brood pheromone will gradually disappear as the brood emerges. By the time all of the brood has emerged the workers’ ovaries will have started to develop, although considerable difference in the rate of development occurs in different individuals, as does the number of eggs a particular worker can produce when its ovaries are fully developed (Morse and Hooper 1985). Another mechanism that limits successful worker reproduction is a behavior known as worker policing: the removal of worker-laid eggs by other workers (Dampney et al. 2002). Visscher (1996) estimates that in queenright colonies of mixed European origin, only approximately 7% of all male eggs originate from laying workers.
While workers with fully developed ovaries are rare in queenright honey bee colonies, Smith et al. (2013) showed that partial ovary development is common. Across nine studies, an average of 6% to 43% of workers had partially developed ovaries in queenright colonies with naturally mated queens. Workers with partially developed ovaries have ovaries that are neither resting (i.e., no swelling of the ovarioles) nor at an advanced stage of development (i.e. completely elongated eggs visible within ovarioles) (Velthuis 1970). There was substantial variation across these studies in the proportion of workers that had partially developed ovaries, which is probably attributable to differences in year, location, season, genetics and dissection methodology (Backx et al. 2012; Hoover et al. 2006). Nevertheless, it is clear that partial ovary development is consistently observed among workers in queenright colonies. This observation raises the question: if workers with only partially developed ovaries are effectively sterile, what is the significance of this incomplete investment in reproductive physiology?
Hoover et al. (2006) examined the effect of larval and adult nutrition on worker ovary development. Workers were fed high or low-pollen diets as larvae, and high or low-protein diets as adults. Workers fed low-protein diets at both life stages had the lowest levels of ovary development, followed by those fed high-protein diets as larvae and low-quality diets as adults, and then those fed diets poor in protein as larvae but high as adults. Workers fed high-protein diets at both life stages had the highest levels of ovary development. The increases in ovary development due to improved dietary protein in the larval and adult life stages were additive. Adult diet also had an effect on body mass. The results demonstrate that both carry-over of larval reserves and nutrients acquired in the adult life stage are important to ovary development in worker honey bees. Carry-over from larval development, however, appears to be less important to adult fecundity than is adult nutrition. Seasonal trends in worker ovary development and mass were examined throughout the brood rearing season. Worker ovary development was lowest in Spring, highest in mid-Summer, and intermediate in Fall.
While workers with fully developed ovaries are rare in queenright honey bee colonies, Smith et al. showed that partial ovary development is common.
Ratnieks (1993) investigated whether worker policing via the selective removal of worker-laid male eggs occurs in normal colonies with a queen. Queenright colonies were set up with the queen below a queen excluder. Frames of worker brood and drone comb were placed above the queen excluder. Daily inspections of the drone frames revealed the presence of a few eggs, presumably laid by workers, at a rate of one egg per 16,000 drone cells. Eighty-five percent of these eggs were removed within one day and only two percent hatched. Dissections of workers revealed that about one worker in 10,000 had a fully developed egg in her body. These data show that worker egg-laying and worker policing are both normal, though rare, in queenright colonies.
Lin et al. (1999) examined the factors that might influence ovary development in worker honey bees. Queenless workers at different ages (< 12 hours, and four, eight, and 21 days) were tested in cages for ovarian development. Newly emerged, four- and eight-day-old, and 21-day-old workers had medium-, large- and small-sized ovaries, respectively, suggesting that of the worker ages tested only four- and eight-day-old workers are likely to become egg layers in a queenless colony. Also, they compared ovarian development of newly emerged workers that were caged for 14 days and allowed to consume either pollen or royal jelly to that of another group of workers similarly caged but screened so that they could only obtain food via trophallaxis from young bees. Ovaries of newly emerged workers that received food from young bees were as well developed as those of newly emerged workers allowed to take pollen or royal jelly directly. Screened workers also had lower but still elevated vitellogenin levels compared with bees having direct access to food. These results indicate that nurse-age bees functioning as pollen-digesting units affect the ovarian development of other workers and to a lesser extent vitellogenesis via food exchange. They compared the influence of group sizes of 25, 125, and 600 bees per cage on ovarian development for 14 days. The two groups of 25 and 125 bees had similar mean ovary scores, and higher scores than a group of 600 bees. Their findings suggest that nurse-age bees could play an important role in mediating worker fertility via trophallaxis, possibly by differentiating worker dominance status, and generally only young workers become fertile when a queen is lost in a colony.
Hoover et al. (2003) investigated the identity of the queen-produced compounds that inhibit worker honey bee ovary development. They examined the effects of synthetic honey bee queen mandibular pheromone (QMP), four newly identified queen retinue pheromone components (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), and whole-queen extracts on ovary development of caged worker bees. The newly identified compounds did not inhibit worker ovary development alone, nor did they improve the efficacy of QMP when applied in combination. QMP was as effective as queen extracts at ovary regulation. Caged workers in the QMP and queen extract treatments had better developed ovaries than did workers remaining in queenright colonies. They concluded that QMP is responsible for the ovary-regulating pheromonal capability of queens from European-derived Apis mellifera subspecies.
When workers activate their ovaries and begin to lay eggs, this physiological change is accompanied by a shift in their pheromonal bouquet, which becomes more queen-like.
When workers activate their ovaries and begin to lay eggs, this physiological change is accompanied by a shift in their pheromonal bouquet, which becomes more queen-like. Tan et al. (2012) found that the pheromonal components HOB, 9-ODA, HVA, 9-HAD, 10-HDAA, 10-HDA have significantly higher amounts in laying workers than in non-laying workers.
Genetic markers were used to study the reproductive behavior of worker honey bees. Five experiments were conducted that demonstrate the significance of worker reproduction. Biases were found in the egg-laying success of workers belonging to different subfamilies within queenless colonies, however, members of all subfamilies laid eggs. These biases were probably not a consequence of direct reproductive competition among subfamily members but most likely represent genetic variability for the timing of the onset of oviposition. Workers preferentially oviposit in drone-sized cells, demonstrating a caste-specific adaptation for oviposition behavior. Drone brood production is highly synchronous within colonies and can result in the production of more than 6,000 drones before colonies die (Page and Erickson 1988).
Wegener et al. (2010) investigated whether differences in the reproductive biology of queens and laying workers are reflected in their eggs. They first tested the capacity of queen- and worker-laid male eggs to withstand dry conditions, by incubating samples at 30.0, 74.9 and 98.7% relative humidity. They found that worker-laid eggs were more sensitive to desiccation. Secondly, they measured the weight and quantities of vitellin, total protein, lipid, glycogen, and free carbohydrate in queen- and worker-laid eggs. Although worker-laid eggs were found to be heavier than queen-laid eggs in two of the four replicates, no systematic differences were found regarding nutrient content. Finally, they compared the duration of embryo development in the two egg types. Worker-laid eggs developed more slowly than queen-laid eggs in two out of three replicates, suggesting that they may only be partly mature at the moment they are laid.
Miller and Ratnieks (2001) measured changes in worker egg-removal behavior, ovary development, and egg-laying rate in hives following the removal of their queens. They carried out weekly assays of worker removal of experimentally transferred eggs, dissection and inspection of worker bee ovaries, and daily checks of worker oviposition. Following queen removal, the egg-removal rate by workers generally first increased, then decreased or leveled off over the four-week time course of the experiment; this behavior was closely synchronized with the increase in worker ovary development and egg-laying.
The physiological state and behavior of laying workers partly resemble those of queens. Laying workers have low juvenile hormone titers and relatively high vitellogenin levels in the hemolymph (blood) similar to queens. Nakaoka et al. (2008) investigated whether the physiological state of laying workers is more similar to that of nurse bees or foragers by examining the hypopharyngeal gland (produces brood-food) and hemolymph vitellogenin titers. In a normal colony, nurse bees have well-developed hypopharyngeal glands that synthesize royal jelly proteins and high hemolymph vitellogenein tiers, whereas foragers have shrunken hypopharyngeal glands and low hemolymph vitellogenin titers. In queenless colonies, however, laying workers tended to have more developed hypopharyngeal glands and to synthesize royal jelly proteins, whereas workers with shrunken hypopharyngeal glands tended to synthesize α-glucosidase (which is needed for processing nectar into honey) and to have undeveloped ovaries. Furthermore, the workers with developed ovaries had higher vitellogenin titers than nurse bees, whereas, those with undeveloped ovaries had lower vitellogenin titers. These findings indicate that the physiological state of laying workers is similar to that of nurse bees, but opposite that of foragers.
Three experiments were performed to determine the role of juvenile hormone (JH) in worker reproduction in queenless colonies of honey bees. In the first experiment, egg-laying workers had low hemolymph titers of JH, as did bees engaged in brood care, while foragers had significantly higher titers. Experiment 2 confirmed these findings by demonstrating that laying workers have significantly lower rates of JH biosynthesis than foragers do. In the third experiment, ovary development was inhibited slightly by application of the JH analog methoprene to one-day-old bees, but was not affected by application to older bees, at least some already displaying egg-laying behavior. These results, which are consistent with earlier findings for queen honey bees, are contrary to a common model of insect reproduction, in which elevated JH titers trigger ovary development, which then leads to oviposition (Robinson et al. 1992).
Vasfi Gencer and Kahya (2011) compared sperm traits of small drones from laying worker colonies (LWC) with those of large drones from queenright colonies (QRC). The drones from QRC were 50.4 percent heavier (221.6 mg) than the drones from laying worker colonies (147. 3 mg). The mean volume of ejaculate of drones from QRC (1.01µl) was 53 percent larger than that of drones from LWC (0.66 µl). The mean sperm number in drones from QRC (7.320 x 106) was significantly higher than that of drones from LWC (4.425 x 106). Sperm concentration of drones from queenright colonies (7.256 x 106/ µl) was significantly higher than that of drones from laying worker colonies (6.661 x 106/ µl). In addition, the drones from queenright colonies (33.155 x 103 mg) produced 3.189 x 103 more sperm cells per mg body mass than drones from laying worker colonies (29.966 x 103 mg). No significant differences were found between drones from QRC and LWC in sperm viability and sperm length.
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