Genotypic variability for rates of behavioral development in worker honeybees (Apis mellifera L.)

We demonstrate the effects of age and genotype on the likelihoodthat an individual worker honeybee (Apis mellifera L.) willbecome a forager. We established experimental colonies thatwere each initially composed of identifiable, nonforaging workersof similar ages (1–5 days old). Workers in each test colonywere the progeny of two queens, providing age and genotypicgroups of workers for comparisons. We then recorded the daythat each worker was first observed foraging. Older workerswere more likely to become foragers under our experimental conditions,even when age differences were just 1 day. At a given age, workersfrom different queens varied in their likelihood of becomingforagers. However, we found that neither age nor genotype (queensource) directly affected the likelihood that a forager wouldrevert to within-nest, larval care activities after the removalof nonforaging bees from colonies. The likelihood of revertingwas only dependent on how long a worker was a forager beforeremoving the nonforagers of the colony.     

Differences in sNPF Receptor-Expressing Neurons in Brains of Fire Ant (Solenopsis invicta Buren) Worker Subcastes: Indicators for Division of Labor and Nutritional Status?

In the red imported fire ant, Solenopsis invicta Buren, the neuronal and molecular mechanisms related to worker division of labor are poorly understood. Workers from different subcastes (major, medium and minors) perform different tasks, which are loosely associated with their size. We hypothesized that the short neuropeptide F (sNPF) signaling system (NPY-like) could be involved in mechanisms of worker division of labor and sensing or responding to colony nutritional requirements. Thus, we investigated the expression of the short neuropeptide F receptor (sNPFR) in the brain and subesophageal ganglion (SEG) of workers from colonies with and without brood. Across worker subcastes a total of 9 clusters of immunoreactive sNPFR cells were localized in the brain and the subesophageal ganglion (SEG); some of these cells were similar to those observed previously in the queen. Worker brain sNPFR cell clusters were found in the protocerebrum near mushroom bodies, in the central complex and in the lateral horn. Other sNPFR immunoreactive cells were found at the edge of the antennal lobes. Across subcastes, we observed both a constant and a differential pattern of sNPFR clusters, with a higher number of sNPFR cells found in minor than in major workers. Those sNPFR cells detected in all worker subcastes appear to be involved in olfaction or SEG functions. The differential expression of clusters in subcastes suggests that sNPFR signaling is involved in regulating behaviors associated with specific subcastes and thus, division of labor. Some sNPFR cells appear to be involved in nutrient sensing and/or brood care, feeding behavior and locomotion. In colonies without brood, workers showed a lower cluster number, and an overall reduced sNPFR signal. Our results suggest the sNPF signaling system is a candidate for the neurobiological control of worker division of labor and sensing brood presence, perhaps correlating with protein requirements and availability.     

Caste specialization in food storage in the dimorphic antColobopsis nipponicus (Wheeler)

Summary Food-storage abilities of both minor and major workers of the dimorphic antColobopsis nipponicus were examined to prove the hypothesis that major workers of this species have a trophic role in addition to a defensive one. Both worker subcastes accumulated water, that was supplied with water-soluble food, as well as fat, that probably originated from food given. However, the major workers accumulated much larger amounts of water and fat than did the minor workers. Difference in water accumulation increased as food supply increased. In spite of their difference in body weight, the residual amount of fat after starvation did not differ between the two subcastes. Although the amount of stored fat increased with colony size in both subcastes, the stored fat per dry weight of the major worker was significantly larger than that of the minor worker regardless of the colony size. When workers of a colony were starved, minor workers with a single major worker survived significantly longer than those without a major worker. In addition, about half the behavioral acts of major workers were regurgitation for minor workers. These results demonstrate that major workers ofC. nipponicus functioned as a trophic caste. On the basis of the results, quantitative relationships of storage abilities between the two worker subcastes are discussed.     

Social parasitism by Cape honeybee workers in colonies of their own subspecies (Apis mellifera capensis Esch.)

Social parasitism is widespread in the eusocial insects. Although social parasites often show a reduced worker caste, unmated workers can also parasitize colonies. Cape honeybee workers, Apis mellifera capensis, can establish themselves as social parasites in host colonies of other honeybee subspecies. However, it is unknown whether social parasitism by laying workers also occurs among Cape honeybee colonies. In order to address this question we genotyped worker offspring of six queenless A. m. capensis colonies and determined the maternity of the reproducing workers. We found that three non-nestmate workers dominated reproduction in a host colony and produced 62.5% of the progeny. Our results show that social parasitism by laying workers is a naturally occurring part of the biology of Cape honeybees. However, such social parasitism is not frequently found (6.41% of the total worker offspring) probably due to co-evolutionary processes among A. m. capensis resulting in an equilibrium between selection for reproductive dominance in workers, colony maintenance and queen adaptation. Received 28 July 2005; revised 19 September and 11 November 2005; accepted 16 November 2005.     

蜜蜂无政府主义蜂群的研究进展

西方蜜蜂Apis mellifera作为典型的社会性昆虫, 最重要的特征是生殖劳动分工。蜂王垄断蜂群的生殖权利, 工蜂生殖功能受到抑制, 从事除产卵和交配以外的所有职能。而在无政府主义蜂群中, 即使蜂王存在, 也有较多工蜂的卵巢激活并产卵, 蜂群中大多数雄蜂是工蜂的后代。这些特殊蜂群为正常蜂群工蜂不育机制研究提供了绝佳的反例材料。本文对无政府主义蜂群的行为特征、 产生条件、 遗传基础等研究进行了综述。无政府主义蜂群中有较多的工蜂产卵, 且工蜂所产卵能够逃避工蜂监督, 这种行为的产生受环境、 遗传组成、 基因表达等多种因素的影响, 并且遗传结构体系复杂, 参与调控的基因数量多。无政府主义蜂群行为机制的研究为工蜂不育机制的揭示及其他社会性昆虫工职不育基因的筛选和功能研究提供借鉴。     

蜜蜂大脑的分区与功能

蜜蜂Apis mellifera L.是神经生物学研究的重要模式生物。尽管工蜂脑的体积不足1mm3,包含的神经元数量不到百万,但却拥有丰富的个体和社会行为,甚至还有学习、记忆、认知等高级行为。如此微小的大脑也是通过不同结构分区来实现其丰富复杂的行为。本文对蜜蜂大脑的精细解剖结构以及脑区功能研究进行了综述,为昆虫科学和神经生物学研究提供参考。     

Worker honeybee brain proteome

A large-scale mapping of the worker honeybee brain proteome was achieved by MudPIT. We identified 2742 proteins from forager and nurse honeybee brain samples; 17% of the total proteins were found to be differentially expressed by spectral count sampling statistics and a G-test. Sequences were compared with the EuKaryotic Orthologous Groups (KOG) catalog set using BLASTX and then categorized into the major KOG categories of most similar sequences. According to this categorization, nurse brain showed increased expression of proteins implicated in translation, ribosomal structure, and biogenesis (14.5%) compared with forager (1.8%). Experienced foragers overexpressed proteins involved in energy production and conversion, showing an extensive difference in this set of proteins (17%) in relation to the nurse subcaste (0.6%). Examples of proteins selectively expressed in each subcaste were analyzed. A comparison between these MudPIT experiments and previous 2-DE experiments revealed nine coincident proteins differentially expressed in both methodologies.     

Cape honeybees, Apis mellifera capensis, police worker-laid eggs despite the absence of relatedness benefits

In the Cape honeybee, Apis mellifera capensis, workers lay diploid(female) eggs via thelytoky. In other A. mellifera subspecies,workers lay haploid (male) eggs via arrhenotoky. When thelytokousworker reproduction occurs, worker policing has no relatednessbenefit because workers are equally related to their sisterworkers' clonal offspring and their mother queen's female offspring.We studied worker policing in A. m. capensis and in the arrhenotokousAfrican honeybee A. m. scutellata by quantifying the removalrates of worker-laid and queen-laid eggs. Discriminator coloniesof both subspecies policed worker-laid eggs of both their ownand the other subspecies. The occurrence of worker policing,despite the lack of relatedness benefit, in A. m. capensis stronglysuggests that worker reproduction is costly to the colony andthat policing is maintained because it enhances colony efficiency.In addition, because both subspecies policed each others eggs,it is probable that the mechanism used in thelytokous A. m.capensis to discriminate between queen-laid and worker-laideggs is the same as in arrhenotokous A. m. scutellata.     

Studies on calmodulin-binding proteins (CaMBPs) in the cilia of Tetrahymena

As a first step to elucidate the involvement of calmodulin in Ca2+-dependent regulation of ciliary motility, molecular species and properties of calmodulin-binding proteins (CaMBPs) in Tetrahymena cilia were investigated by a modified [125I]calmodulin overlay method. At least 36 kinds of CaMBPs were detected. All the CaMBPs bound to calmodulin in Ca2+-dependent and calmodulin-specific manners, but they showed different Ca2+-dependencies. Several of CaMBPs bound to calmodulin in the presence of 100 microM trifluoperazine, several did in the presence of 8 M urea, and a few of them were highly sensitive to trypsin digestion. Among these CaMBPs, we noticed a 95 000-dalton (D) CaMBP present in the outerdoublet microtubule fraction, which possessed some attributes of the calmodulin counterpart suggested from the results of our previous paper [12]. We discussed a possibility that this protein might correspond to one of the protein components of the interdoublet link.     

Pheromonal dominance and the selection of a socially parasitic honeybee worker lineage (Apis mellifera capensis Esch.)

The recent invasion by self-replicating socially parasitic Cape honeybee workers, Apis mellifera capensis, of colonies of the neighbouring African subspecies Apis mellifera scutellata represents an opportunity to study evolution of intraspecific parasitism in real time. As honeybee workers compete pheromonally for reproductive dominance, and as A. m. capensis workers readily produce queen-like pheromones, we hypothesized that these semiochemicals promoted the evolution of intraspecific social parasitism. Remarkably, the offspring of a single worker became established as a parasite in A. m. scutellata's range. This could have resulted from extreme selection among different clonal parasitic worker lineages. Using pheromonal contest experiments, we show that the selected parasitic lineage dominates in the production of mandibular gland pheromones over all other competitors to which it is exposed. Our results suggest that mandibular gland pheromones played a key role in the evolution of intraspecific social parasitism in the honeybee and in the selection of a single genotype of parasitic workers.     

Calmodulin-binding domains in Alzheimer's disease proteins: extending the calcium hypothesis

The calcium hypothesis of Alzheimer's disease (AD) invokes the disruption of calcium signaling as the underlying cause of neuronal dysfunction and ultimately apoptosis. As a primary calcium signal transducer, calmodulin (CaM) responds to cytosolic calcium fluxes by binding to and regulating the activity of target CaM-binding proteins (CaMBPs). Ca(2+)-dependent CaMBPs primarily contain domains (CaMBDs) that can be classified into motifs based upon variations on the basic amphiphilic alpha-helix domain involving conserved hydrophobic residues at positions 1-10, 1-14 or 1-16. In contrast, an IQ or IQ-like domain often mediates Ca(2+)-independent CaM-binding. Based on these attributes, a search for CaMBDs reveals that many of the proteins intimately linked to AD may be calmodulin-binding proteins, opening new avenues for research on this devastating disease.     

Target molecules of calmodulin on microtubules of Tetrahymena cilia

In the course of an attempt to isolate the calmodulin-binding proteins (CaMBPs) from cilia of Tetrahymena, it was found that some CaMBPs tend to interact with axonemal microtubules. The present study demonstrates this interaction by cosedimentation experiments using in vitro polymerized Tetrahymena axonemal microtubules and Tetrahymena CaMBPs purified from axonemes by calmodulin affinity column chromatography. Analysis by the [125I]calmodulin overlay method showed that at least three CaMBPs (Mr69, 45, and 37 kDa) cosediment with microtubules. Furthermore, without any addition of exogenous CaMBPs, microtubules purified after three cycles of temperature-dependent polymerization and depolymerization included the above CaMBPs and additional CaMBPs (Mr30, 26, and 22 kDa) which could not cosediment with microtubules. From the results, we have classified these microtubule-associated CaMBPs into two groups: (i) CaMBPs which interact with microtubules only during polymerization (30, 26, and 22 kDa), and (ii) CaMBPs which interact not only with microtubules during polymerization, but also with polymerized microtubules (69, 45, and 37 kDa). These results suggest that the microtubule-associated CaMBPs, especially those of the latter group, are located on the surface of ciliary microtubules, and may become the target molecules of calmodulin at Ca2+-triggered ciliary reversal.     

Functional flexibility of the honey bee hypopharyngeal gland in a dequeened colony

The role of the worker honey bee Apis mellifera L. changes depending on age after eclosion (age polyethism): young workers (nurse bees) take care of their brood by synthesizing and secreting brood food (royal jelly), while older workers (foragers) forage for nectar and process it into honey. Previously, we showed that the major proteins synthesized in the hypopharyngeal gland of the worker change from brood food proteins to alpha-glucosidase at the single secretory cell level in parallel with this age polyethism [Kubo et al., J. Biochem. 119, 291-295 (1996); Ohashi et al., Eur. J. Biochem. 249, 797-802 (1997)]. Here, we examined whether the function of the hypopharyngeal gland has flexibility depending on the colony conditions, by creating a dequeened colony in which the older workers were compelled to feed the drone larvae. It was found that most of the older workers in the dequeened colony synthesized brood food proteins as did nurse bees. Furthermore, the percentage of workers that synthesized brood food proteins was maintained at 80-90% of the total workers for at least two months, as in a normal colony. These results indicate that the function of the hypopharyngeal gland cells of the worker has flexibility and can, if necessary, be maintained as that of the nurse bee, depending on the condition of the colony.     

Identification of honeybee antennal proteins/genes expressed in a sex- and/or caste selective manner

We identified three candidate proteins/genes involved in caste and/or sex-specific olfactory processing in the honeybee Apis mellifera L., that are differentially expressed between the antennae of the worker, queen, and drone honeybees using SDS-polyacrylamide gel electrophoresis or the differential display method. A protein was identified, termed D-AP1, that was expressed preferentially in drone antennae when compared to those of workers. cDNA cloning revealed that D-AP1 is homologous to carboxylesterases. Enzymatic carboxylesterase activity in the drone antennae was higher than in the workers, suggesting its dominant function in the drone antennae. In contrast, two proteins encoded by genes termed W-AP1 and Amwat were expressed preferentially in worker antennae when compared to those of queens. W-AP1 is homologous to insect chemosensory protein, and Amwat encodes a novel secretory protein. W-AP1 is expressed selectively in worker antennae, while Amwat is expressed both in the antennae and legs of the workers. These findings suggest that these proteins are involved in the antennal function characteristic to drone or worker honeybees.     

Gaseous neuromodulator-related genes expressed in the brain of honeybee Apis mellifera

Nitric oxide (NO), hydrogen sulfide (H2S), and carbon monoxide (CO) are thought to act as gaseous neuromodulators in the brain across species. For example, in the brain of honeybee Apis mellifera, NO plays important roles in olfactory learning and discrimination, but the existence of H2S- and CO-mediated signaling pathways remains unknown. In the present study, we identified the genes of nitric oxide synthase (NOS), soluble guanylyl cyclase (sGC), cystathionine beta-synthase (CBS), and heme oxygenase (HO) from the honeybee brain. The honeybee brain contains at least one gene for each of NOS, CBS, and HO. The deduced proteins for NOS, CBS, and HO are thought to contain domains to generate NO, H2S, and CO, respectively, and to contain putative Ca2+/calmodulin-binding domains. On the other hand, the honeybee brain contains three subunits of sGC: sGCalpha1, sGCbeta1, and sGCbeta3. Phylogenetic analysis of sGC revealed that Apis sGCalpha1 and sGCbeta1 are closely related to NO- and CO-sensitive sGC subunits, whereas Apis sGCbeta3 is closely related to insect O2-sensitive sGC subunits. In addition, we performed in situ hybridization for Apis NOS mRNA and NADPH-diaphorase histochemistry in the honeybee brain. The NOS gene was strongly expressed in the optic lobes and in the Kenyon cells of the mushroom bodies. NOS activity was detected in the optic lobes, the mushroom bodies, the central body complex, the lateral protocerebral lobes, and the antennal lobes. These findings suggest that NO is involved in various brain functions and that H2S and CO can be endogenously produced in the honeybee brain.