David W. Borst

Physiological correlates of division of labor among similarly aged honey bees

Hormone analyses and exocrine gland measurements were made to probe for physiological correlates of division of labor among similarly aged adult worker honey bees (Apis mellifera L.). Middle-age bees (ca. 2 weeks old) performing different tasks showed significant differences in both juvenile hormone (JH) biosynthesis rates and hemolymph titers; guards and undertakers had high JH, and wax producers and food storers, low JH. Guards and undertakers had similar hormone levels to foragers, even though they were 10 days younger than foragers. No differences in JH were detected among young bees (1-week-old queen attendants and nurses) or older bees (3–4 week-old pollen foragers, non-pollen foragers, and soldiers). Hypopharyngeal gland size was inversely correlated with worker age and rate of JH biosynthesis, but soldiers had significantly larger hypopharyngeal glands than did foragers, despite their similar age and JH level. Results from soldiers indicate that exocrine gland development is not always linked with age-related behavior and endocrine development; they also support the recent claim that soldiers constitute a group of older bees that are distinct from foragers. Hormonal analyses indicate that the current model of JHs role in honey bee division of labor needs to be expanded because high levels of JH are associated with several other tasks besides foraging. JH may be involved in the regulation of division of labor among similarly aged workers in addition to its role in age-related division of labor.

Methyl farnesoate and its role in crustacean reproduction and development

Studies with Libinia emarginata suggest that methyl farnesoate (MF), a product of the mandibular organs (MOs), may be a crustacean juvenile hormone. In order to better understand the significance of this compound in crustacean physiology, we first investigated the presence of MF in other decapods. MF was synthesized and secreted by MOs from all species tested. However, large differences in the level of MF secretion were observed between species and also between individuals of a species. For example, the level of secretion by MOs from L. emarginata was 100-fold greater than that observed in MOs from Homarus americanus. Analysis of hemolymph from these two species by GC-MS indicated comparable differences in the amount of MF present. Differences in the level of MF secretion by MOs from individuals of a species appear to reflect the physiological roles of this compound. For example, a close relationship was seen between MF secretion and gametogenesis in females of L. emarginata. Finally, treatment of lobster larvae with seawater containing MF caused a small but significant delay in their metamorphosis when compared with untreated larvae. These data suggest that MF affects reproduction in a manner similar to the effects of JH on insects, and may also have effects on the development of crustacean larvae. Taken together, these data support the classification of MF as a crustacean JH.

Juvenile hormone titers, juvenile hormone biosynthesis, ovarian development and social environment in Bombus terrestris.

The effects of the social environment and age on juvenile hormone (JH) and reproduction were investigated by measuring ovarian development, hemolymph levels of JH III, and rates of JH biosynthesis from the same individual bumble bees (Bombus terrestris). Differences in social environment were associated with differences in rates of JH biosynthesis, JH titer and ovarian development. Young queenless workers had a higher rate of JH biosynthesis, JH titer and ovarian development than queenright (QR) workers of similar age. Dominant workers in QR colonies had a higher rate of JH biosynthesis, JH titer and ovarian development than low ranked workers of similar size. There was a positive correlation between JH titer and ovarian development, but no correlation between rate of JH biosynthesis and ovarian development or between JH biosynthesis and JH titer. Both JH titer and rate of JH biosynthesis increased with age from emergence to 3 days of age, but 6-day-old workers, egg-laying workers, and actively reproducing queens had high JH titers and highly developed ovaries but low rates of JH biosynthesis. These results show that reproduction in B. terrestris is strongly affected by the social environment and the influence of the environment on reproduction is mediated by JH. Our data also indicate that the rate of JH biosynthesis measured in vitro is not a reliable indicator of JH titer or ovarian development in B. terrestris; possible reasons are discussed.

Rapid elevation of juvenile hormone titer during behavioral assessment of the breeding resource by the burying beetle, Nicrophorus orbicollis

Female burying beetles (Nicrophorus orbicollis) rapidly complete ovarian maturation upon discovering a suitable carrion resource for breeding. In this study, we examined changes in hemolymph titers of juvenile hormone (JH) over the first 30 days of adult female life, and in response to the discovery of a mouse carcass. Levels of JH were found to increase gradually over the first 20 days, and then increased abruptly within 24 h of discovery of a carcass. Changes in JH titer were correlated with increases in ovarian mass and length of terminal oocytes. To more precisely determine the timing of the endocrine response to a carcass, hemolymph titers of JH were measured 2, 10, 20 and 60 min after carcass discovery. Titers of JH were significantly elevated (112% over controls) in just 10 min. To confirm this resource discovery-related increase in JH, hemolymph samples were taken from the same individuals both 2 days before, and 10 min after, discovery of a carcass. Again, JH titers rose significantly (170%) in beetles 10 min after carcass discovery. Prominent behaviors observed during the 10-min period following discovery included palpating, lifting, walking around the carcass and making forays into the surrounding soil. Feeding did not occur. These results suggest that the rapid JH surge in female burying beetles is triggered by information obtained during behavioral assessment of a breeding resource, and not by mating or feeding cues. The association between ovarian development and JH titer further suggests a role for JH in co-ordinating reproduction in a temporally and spatially unpredictable environment.

Methyl farnesoate: Its site of synthesis and regulation of secretion in a juvenile crustacean

Abstract Analogy to Insecta suggests that JH(s) may play important roles in the development and reproduction of Crustacea. It has recently been shown by Laufer et al. (1987), using LC and GC/MS that the mandibular organ of adult crabs synthesized methyl farnesoate, the unepoxidated from of JH III, and that synthesis varied according to the sex, as a function of eyestalk ablation, and stage of vitellogenesis. In experiments reported here, we found that eyestalk removal from juvenile spider crabs, Libinia emarginata , resulted in a two-fold increase in the rate of MF synthesis by MOs in vitro . Furthermore, 2 h incubations of MOs from eyestalk ablated animals with eyestalk extracts inhibited the rate of synthesis of MF by about 60%. These results suggest that MF may be a gonad stimulating hormone (GSH) reported by other workers. Secretion by the MO is inhibited by an eyestalk factor (MO-IH), and MO-IH may be similar or identical to the gonad inhibiting hormone (GIH) reported by others.

Effects of social conditions on Juvenile Hormone mediated reproductive development in Bombus terrestris workers

Abstract. During the annual life cycle of the bumble bee Bombus terrestris (L.) colony, there is a stage characterized by worker reproduction in the presence of the queen. It has been proposed that this is a result of a decrease in queen inhibition. This hypothesis was examined by studying the effects of queens taken from colonies at different stages of development on several aspects of worker physiology and behaviour: rates of Juvenile Hormone (JH) release in vitro, ovary development, and behaviour associated with reproduction. After optimizing and validating the radiochemical assay for JH release for bumble bee workers, we found that queenless workers had significantly more developed ovaries and higher rates of release of JH than did queenright workers, confirming and extending previous findings that suggest that bumblebee ovarian development is under JH control. Mated queens, separated from their colony and brood, can have the same inhibitory effect on the reproductive development of callow workers. In contrast, workers confined with virgin queens or in queenless groups demonstrated a significantly higher rate of release of JH, overt aggression and threatening behaviours. However, there were no differences in rates of release of JH between workers confined in groups in the laboratory with queens taken from colonies either before or after the onset of worker reproduction. Furthermore, overt aggression and threatening behaviours were similar and low in both types of groups. These results gave no support to the hypothesis that a decrease in queen inhibition is associated with the onset of worker reproduction. We also show that young workers reared in colonies either before or after worker reproduction occurs, or in queenless colonies, all demonstrated similar, low rates of release of JH. These results suggest that older workers may inhibit the corpora allata of younger workers in queenless colonies.

Hemolymph levels of methyl farnesoate increase in response to osmotic stress in the green crab, Carcinus maenas

The salinity of estuarine environments can vary widely, exposing resident organisms to considerable osmotic stress. The green crab Carcinus maenas is well known for its ability to osmoregulate in response to such stress. Therefore, we tested the relationship between osmoregulation and hemolymph levels of methyl farnesoate (MF), a compound previously shown to rise in response to various types of environmental stresses. When crabs were transferred from 100% seawater to dilute (hypo-osmotic) seawater, hemolymph osmolality dropped rapidly, reaching an acclimation level 48 h after transfer. Hemolymph levels of MF also rose in these animals after a delay of 6 h, and reached a maximum level at 48 h. MF levels remained elevated as long as the crabs were maintained in dilute seawater, and quickly returned to basal levels when the animals were returned to full strength seawater. In most (but not all) animals, MF levels were elevated when hemolymph osmolality fell below the isosmotic point (approx. 800 mOsm/kg). These data suggest that MF may have a role in osmoregulation by this species. In addition, the elevation of MF by hypo-osmotic seawater suggests an experimental strategy for manipulating MF levels in crustaceans.

Caste Determination in Bombus terrestris: Differences in Development and Rates of JH Biosynthesis between Queen and Worker Larvae.

To study the possible role of juvenile hormone in caste determination in Bombus terrestris, we measured development and rates of juvenile hormone biosynthesis in vitro in larvae destined to develop into either workers or queens. Larvae of both castes developed through four instars and had the same growth rates. However, the duration of the instars was longer for queen larvae, and their head width at the third and fourth instars was significantly larger. After validating the well-known radiochemical assay of JH for bumble bee larvae, we show that worker larvae corpora allata exhibited a constant and low rate of JH biosynthesis, never more than 5 pmol JH/h/pair. Queen larvae, in contrast, had two peaks of JH biosynthesis: a small one during the first instar, which has previously been correlated with caste determination; and a large peak, previously undetected, above 40 pmol JH/h/pair, during the second and third instars. We suggest that caste determination in this species is mediated by JH and that the duration of larval instars is a key factor. The possibility that the queen influences caste determination via an effect on instar duration is also discussed. Copyright 1997 Elsevier Science Ltd. All rights reserved

Regulation of methyl farnesoate in the hemolymph and mandibular organ of the lobster, Homarus americanus

Methyl farnesoate (MF) was measured in the hemolymph and mandibular organs (MO) of the lobster. Although a few animals had detectable MF levels in their hemolymph, this compound was undetectable (less than 0.4 ng/ml) in the hemolymph of most animals. One day after bilateral eyestalk ablation, MF was detected in the hemolymph of all animals, reaching variable levels (2.0-31.2 ng/ml) by the fourth day. Unilateral eyestalk ablation caused a smaller increase in hemolymph levels of MF. Similarly, the MF content of the MO, the only known site of MF synthesis, was low in intact lobsters (8.1 ng/gland) and was elevated in unilaterally and bilaterally eyestalk-ablated animals (54.1 and 106.9 ng/gland, respectively). When extracts of the sinus gland (SG), a source of neuropeptides in the eyestalk, were injected into bilaterally ablated lobsters, hemolymph levels of MF dropped to undetectable levels in 2 to 3 hr. The response to SG extract was dose dependent, and MF levels recovered by 12 to 24 hr after treatment. SG extract also lowered the MF content in the MO from 267.6 to 6.6 ng/gland after 4 hr. These results indicate that MF in the hemolymph and MO is negatively regulated by a factor(s) from the SG.

The effect of queen-worker conflict on caste determination in the bumblebee Bombus terrestris

Abstract Endocrine analyses were used to investigate the well-known association between queen production and the onset of worker reproduction (termed the competition phase, CPh) in Bombus terrestris. Larvae that reached the age of 5 days before the CPh had a worker-like profile: low juvenile hormone (JH) biosynthesis rates and low JH hemolymph titers. In contrast, larvae that reached the age of 5 days during the CPh had a queen-like profile: high JH biosynthesis rates and high hemolymph JH levels. Larval fate could be manipulated by transplanting egg cells into host colonies with different social structures. There was a steep rise in JH production in larvae transplanted into colonies near or during the CPh. This indicates that during colony development, larvae switch from the ”worker developmental pathway” to the ”queen developmental pathway,” and that the switch is socially regulated. In small rearing groups, larvae reared with queens before the CPh developed into workers, whereas those reared with queens after the CPh developed into queens. Variation in worker type (naive or experienced) did not affect caste determination. Therefore, we hypothesize that queens produce a pheromone that directly inhibits queen differentiation by larvae. We also present two alternative scenarios that explain the timing of gyne production in B. terrestris, one based on ecological constraints and the other based on queen-worker competition.