Larval juvenile hormone treatment affects pre-adult development,but not adult age at onset of foraging in worker honey bees (Apis mellifera)

Previous research has shown that juvenile hormone (JH) titers increase as adult worker honey bees age and treatments with JH, JH analogs and JH mimics induce precocious foraging. Larvae from genotypes exhibiting faster adult behavioral development had significantly higher levels of juvenile hormone during the 2nd and 3rd larval instar. It is known that highly increased JH during this period causes the totipotent female larvae to differentiate into a queen. We treated third instar larvae with JH to test the hypothesis that this time period may be a developmental critical period for organizational effects of JH on brain and behavior also in the worker caste, such that JH treatment at a lower level than required to produce queens will speed adult behavioral development in workers. Larval JH treatment did not influence adult worker behavioral development. However, it made pre-adult development more queen-like in two ways: treated larvae were capped sooner by adult bees, and emerged from pupation earlier. These results suggest that some aspects of honey bee behavioral development may be relatively insensitive to pre-adult perturbation. These results also suggest JH titer may be connected to cues perceived by the adult bees indicating larval readiness for pupation resulting in adult bee cell capping behavior.     

Juvenile hormone esterase activity in the pupating and diapausing larvae of Sesamia nonagrioides

The development of the Mediterranean corn borer, Sesamia nonagrioides, under long-day (LD) photoperiod is associated with juvenile hormone (JH) decline and pupation in the 5th or 6th larval instar. The larvae grown under short-day (SD) conditions maintain a moderate JH titer and enter diapause during which they undergo several extra larval molts. Both types of larvae exhibit similar levels of juvenile hormone esterase (JHE) activity that increases in each instar during the period of low ecdysteroid titer and drops when the titer rises to a molt-inducing peak. A suppression of JHE activity within 24h after application of an ecdysteroid agonist suggests that the drop of activity is a rapid and possibly direct response to ecdysteroids or their agonist. Esterase inhibitor 3-octylthio-1,1,1-trifluoro-2-propanone (OTFP) suppressed more than 98% of the JHE activity without affecting pupation timing and adult development. The data indicate that JHE is not crucial for the switch between larval development, diapause, and metamorphosis in S. nonagrioides.     

Methoprene does not affect juvenile hormone titers in honey bee (Apis mellifera) workers

Methoprene, a juvenile hormone (JH) analog, is a widely used insecticide that also accelerates behavioral development in honey bees (Apis mellifera). JH regulates the transition from nursing to foraging in adult worker bees, and treatment with JH or methoprene have both been shown to induce precocious foraging. To determine how methoprene changes honey bee behavior, we compared JH titers of methoprene‐treated and untreated bees. Behavioral observations confirmed that methoprene treatment significantly increased the number of precocious foragers in 3 out of 4 colonies. In only 1 out of 4 colonies, however, was there a significant difference in JH titers between the methoprene‐treated and control bees. Further, in all 4 colonies, there was no significant differences in JH titers between precocious and normal‐aged foragers. These results suggest that methoprene did not directly affect the endogenous JH secreted by corpora allata. Because methoprene caused early foraging without changing workers’ JH titers, we conclude that methoprene most likely acts directly on the JH receptors as a substitute for JH.     

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 JH's 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.Abbreviations JH Juvenile hormone - RIA radioimmunoassay - CA corpora allata - HPLC high performance liquid chromatography - TLC thin layer chromatography     

Methoprene does not affect food preferences and foraging performance in honey bee workers

The fundamental determinants of division of labor among honey bee workers are age, genotype, and environment. These determinants work through intermediate physiological channels to realize particular patterns of division of labor. The change of juvenile hormone (JH) titer in worker bees is one such channel. Previous studies concentrated on the impact of JH on timing of in-hive and foraging activity. Here we examined the effects of JH on task specialization and the collection of pollen or nectar by same-age bees and we tested the possible impact on JH titer on foraging performance. Methoprene treatments were conducted after workers began to forage inside a flight room. We found that methoprene, a JH analogue, had no effect on preferences for pollen or nectar and, also, did not influence nectar foraging rate, nectar load size, and foraging span.     

Juvenile hormone paces behavioral development in the adult worker honey bee

Behavioral development in the adult worker honey bee (Apis mellifera), from performing tasks inside the hive to foraging, is associated with an increase in the blood titer of juvenile hormone III (JH), and hormone treatment results in precocious foraging. To study behavioral development in the absence of JH we removed its glandular source, the corpora allata, in 1-day-old adult bees. The age at onset of foraging for allatectomized bees in typical colonies was significantly older compared with that of sham-operated bees in 3 out of 4 colonies; this delay was eliminated by hormone replacement in 3 out of 3 colonies. To determine the effects of corpora allata removal on sensitivity to changes in conditions that influence the rate of behavioral development, we used "single-cohort" colonies (composed of only young bees) in which some colony members initiate foraging precociously. The age at onset of foraging for allatectomized bees was significantly older compared with that of sham-operated bees in 2 out of 3 colonies, and this delay was eliminated by hormone replacement. Allatectomized bees initiated foraging at significantly younger ages in single-cohort colonies than in typical colonies. These results demonstrate that JH influences the pace of behavioral development in honey bees, but is not essential for either foraging or altering behavioral development in response to changes in conditions.     

Colony Integration in Honey Bees: Mechanisms of Behavioral Reversion

After confirming that worker honey bees (Apis mellifera) can revert from foraging to brood care, we determined whether juvenile hormone (JH) mediates this form of plasticity in behavioral development and whether worker age and genotype influence the probability of its expression. Measurements of JH titers support the hypothesis that plasticity in honey bee behavioral development is a consequence of modulation of JH by extrinsic factors. Observations of individually marked bees in a colony composed of two phenotypically distinguishable subfamilies revealed that the likelihood of undergoing behavioral reversion was influenced by worker age but not by worker genotype. The effect of worker age on reversion is consistent with a previously formulated model for the regulation of age polyethism in honey bees that predicts that workers of different ages have different response thresholds for task-associated stimuli. The lack of a genotypic effect on reversion is in contrast to results for other forms of behavioral plasticity.     

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.     

Rates of juvenile hormone synthesis control caste differentiation in the stingless bee Scaptotrigona postica depilis

Summary In social insects the expression of caste-specific characters is controlled by juvenile hormone (JH) during definite sensitive periods in preimaginal development. For a number of stingless bee species the existence of such a JH-sensitive period has already been demonstrated. Queen development can be induced by topical JH applications during the cocoon spinning phase of the last larval instar. Neither JH titers nor rates of JH synthesis were known so far for this subfamily of eusocial bees distinguished by a pronounced caste dimorphism. As the pantropically distributed stingless bees with approximately 400 recent species are the largest group of social bees, JH synthesis was studied in one of the species that can be kept under laboratory conditions. An in vitro radiochemical assay was used to measure stage- and caste-specific activities of the corpora allata (CA). For the first time in a eusocial hymenopteran species it was demonstrated how the endocrine system is reacting to trophogenic stimuli capable to induce caste differentiation during larval development. Generally JH synthesis in queen CA was found to be 30–80% higher than in workers during the penultimate and last larval instar, but a strong and distinct caste-specific modulation of JH synthesis was only observed right before the onset of a JH-sensitive period in the cocoon spinning phase of the fifth instar.     

Juvenile hormone levels in honey bee (Apis mellifera L.) foragers: foraging experience and diurnal variation

A rising blood titer of juvenile hormone (JH) in adult worker honey bees is associated with the shift from working in the hive to foraging. We determined whether the JH increase occurs in anticipation of foraging or whether it is a result of actual foraging experience and/or diurnal changes in exposure to sunlight. We recorded all foraging flights of tagged bees observed at a feeder in a large outdoor flight cage. We measured JH from bees that had taken 1, 3-5, or >100 foraging flights and foragers of indeterminate experience leaving or entering the hive. To study diurnal variation in JH, we sampled foragers every 6h over one day. Titers of JH in foragers were high relative to nurses as in previous studies, suggesting that conditions in the flight cage had no effect on the relationship between foraging behavior and JH. Titers of JH in foragers showed no significant effects of foraging experience, but did show significant diurnal variation. Our results indicate that the high titer of JH in foragers anticipates the onset of foraging and is not affected by foraging experience, but is modulated diurnally.     

Limits on volume changes in the mushroom bodies of the honey bee brain

The behavioral maturation of adult worker honey bees is influenced by a rising titer of juvenile hormone (JH), and is temporally correlated with an increase in the volume of the neuropil of the mushroom bodies, a brain region involved in learning and memory. We explored the stability of this neuropil expansion and its possible dependence on JH. We studied the volume of the mushroom bodies in adult bees deprived of JH by surgical removal of the source glands, the corpora allata. We also asked if the neuropil expansion detected in foragers persists when bees no longer engage in foraging, either because of the onset of winter or because colony social structure was experimentally manipulated to cause some bees to revert from foraging to tending brood (nursing). Results show that adult exposure to JH is not necessary for growth of the mushroom body neuropil, and that the volume of the mushroom body neuropil in adult bees is not reduced if foraging stops. These results are interpreted in the context of a qualitative model that posits that mushroom body neuropil volume enlargement in the honey bee has both experience-independent and experience-dependent components.     

A whole genome screening and RNA interference identify a juvenile hormone esterase-like gene of the diamondback moth,Plutella xylostella

Juvenile hormone (JH) plays a crucial role in preventing precocious metamorphosis and stimulating reproduction. Thus, its hemolymph titer should be under a tight control. As a negative controller, juvenile hormone esterase (JHE) performs a rapid breakdown of residual JH in the hemolymph during last instar to induce a larval-to-pupal metamorphosis. A whole genome of the diamondback moth (DBM), Plutella xylostella, has been annotated and proposed 11 JHE candidates. Sequence analysis using conserved motifs commonly found in other JHEs proposed a putative JHE (Px004817). Px004817 (64.61 kDa, pI = 5.28) exhibited a characteristic JHE expression pattern by showing high peak at the early last instar, at which JHE enzyme activity was also at a maximal level. RNA interference of Px004817 reduced JHE activity and interrupted pupal development with a significant increase of larval period. This study identifies Px004817 as a JHE-like gene of P. xylostella.     

The critical period for caste determination in Bombus terrestris and its juvenile hormone correlates

The critical period for caste determination and its juvenile hormone (JH III) correlates were studied in Bombus terrestris. Larvae of known age and instar were taken from young colonies, in which they would have been reared as workers, and placed into groups of queenless workers. Under these conditions the critical age for caste determination was 5 days, during the second instar. Endocrine correlates of caste determination were obtained by determining profiles of juvenile hormone titer and juvenile hormone biosynthesis, measured by chiral-specific radioimmunoassay and the in vitro radiochemical assay, respectively. By the middle of the second instar prospective queen larvae had significantly higher rates of juvenile hormone biosynthesis and juvenile hormone titer than prospective worker larvae. Based on the coincidence of timing of both the critical period and the appearance of caste-specific juvenile hormone titer, we suggest that juvenile hormone plays a role in the mechanisms that control caste determination in B. terrestris.     

Developmental expression and hormonal regulation of different isoforms of the transcription factor E75 in the tobacco hornworm Manduca sexta

E75A and E75B, isoforms of the E75 orphan nuclear receptor, are sequentially up-regulated in the abdominal epidermis of the tobacco hornworm Manduca sexta by 20-hydroxyecdysone (20E) during larval and pupal molts, with E75A also increasing at pupal commitment (Zhou et al., Dev. Biol. 193, 127-138, 1998). We have now cloned E75C and show that little is expressed in the epidermis during larval life with trace amounts seen just before ecdysis. Instead, E75C is found in high amounts during the development of the adult wings as the ecdysteroid titer is rising, and this increase was prevented by juvenile hormone (JH) that prevented adult development. By contrast, E75D is expressed transiently during the larval and pupal molts as the ecdysteroid titer begins to decline and again just before ecdysis, but in the developing adult wings is expressed on the rise of 20E. Removal of the source of JH had little effect on either E75C or E75D mRNA expression during the larval and pupal molts. At the time of pupal commitment, in vitro experiments show that 20E up-regulates E75D and JH prevents this increase. Neither E75A nor E75D mRNA was up-regulated by JH alone. Thus, E75C is primarily involved in adult differentiation whereas E75D has roles both during the molt and pupal commitment.