Immunogold Localization of Vitellogenin in the Ovaries, Hypopharyngeal Glands and Head Fat Bodies of Honeybee Workers, Apis Mellifera

Vitellogenin is a yolk precursor protein in most oviparous females. In the advanced eusocial honeybee, Apis mellifera (Hymenoptera: Apidae), vitellogenin has recently attracted much interest as this protein, in addition to a classical function in oocyte development in the reproductive queen caste, has evolved functions in the facultatively sterile female worker caste not documented in other species. However, research on the spatial dynamics of vitellogenin in various tissues is not easily performed with available tools. Here we present an immunogold staining procedure that visualizes honeybee vitellogenin in resin embedded tissue. To establish the protocol, we used ovaries of worker bees from colonies with and without a queen. Under the first condition, vitellogenin is assumed not to be present in the workers' ovaries. Under the second condition, the ovaries of worker bees become vitellogenic, with abundant opportunities for detection of complex patterns of vitellogenin uptake and storage. By use of this experimental setup, the staining method is shown to be both sensitive and specific. To demonstrate the functional significance of the protocol, it was subsequently used to identify vitellogenin protein in the hypopharyngeal glands (brood food producing head glands) of nursing worker bees and in adjacent head fat body cells for the first time. Localization of vitellogenin in these tissues supports previously hypothesized roles of vitellogenin in social behavior. This protocol thus provides deeper insights into the functions of vitellogenin in the honeybee

The Gene vitellogenin Has Multiple Coordinating Effects on Social Organization

Temporal division of labor and foraging specialization are key characteristics of honeybee social organization. Worker honeybees (Apis mellifera) initiate foraging for food around their third week of life and often specialize in collecting pollen or nectar before they die. Variation in these fundamental social traits correlates with variation in worker reproductive physiology. However, the genetic and hormonal mechanisms that mediate the control of social organization are not understood and remain a central question in social insect biology. Here we demonstrate that a yolk precursor gene, vitellogenin, affects a complex suite of social traits. Vitellogenin is a major reproductive protein in insects in general and a proposed endocrine factor in honeybees. We show by use of RNA interference (RNAi) that vitellogenin gene activity paces onset of foraging behavior, primes bees for specialized foraging tasks, and influences worker longevity. These findings support the view that the worker specializations that characterize hymenopteran sociality evolved through co-option of reproductive regulatory pathways. Further, they demonstrate for the first time how coordinated control of multiple social life-history traits can originate via the pleiotropic effects of a single gene that affects multiple physiological processes

The Making of a Queen: TOR Pathway Is a Key Player in Diphenic Caste Development

Honey bees (Apis mellifera) provide a principal example of diphenic development. Excess feeding of female larvae results in queens (large reproductives). Moderate diet yields workers (small helpers). The signaling pathway that links provisioning to female developmental fate is not understood, yet we reasoned that it could include TOR (target of rapamycin), a nutrient- and energy-sensing kinase that controls organismal growth.Here, the role of Apis mellifera TOR (amTOR) in caste determination is examined by rapamycin/FK506 pharmacology and RNA interference (RNAi) gene knockdown. We show that in queen-destined larvae, the TOR inhibitor rapamycin induces the development of worker characters that are blocked by the antagonist FK506. Further, queen fate is associated with elevated activity of the Apis mellifera TOR encoding gene, amTOR, and amTOR gene knockdown blocks queen fate and results in individuals with worker morphology.A much-studied insect dimorphism, thereby, can be governed by the TOR pathway. Our results present the first evidence for a role of TOR in diphenic development, and suggest that adoption of this ancestral nutrient-sensing cascade is one evolutionary pathway for morphological caste differentiation in social insects

PDK1 and HR46 Gene Homologs Tie Social Behavior to Ovary Signals

The genetic basis of division of labor in social insects is a central question in evolutionary and behavioral biology. The honey bee is a model for studying evolutionary behavioral genetics because of its well characterized age-correlated division of labor. After an initial period of within-nest tasks, 2–3 week-old worker bees begin foraging outside the nest. Individuals often specialize by biasing their foraging efforts toward collecting pollen or nectar. Efforts to explain the origins of foraging specialization suggest that division of labor between nectar and pollen foraging specialists is influenced by genes with effects on reproductive physiology. Quantitative trait loci (QTL) mapping of foraging behavior also reveals candidate genes for reproductive traits. Here, we address the linkage of reproductive anatomy to behavior, using backcross QTL analysis, behavioral and anatomical phenotyping, candidate gene expression studies, and backcross confirmation of gene-to-anatomical trait associations. Our data show for the first time that the activity of two positional candidate genes for behavior, PDK1 and HR46, have direct genetic relationships to ovary size, a central reproductive trait that correlates with the nectar and pollen foraging bias of workers. These findings implicate two genes that were not known previously to influence complex social behavior. Also, they outline how selection may have acted on gene networks that affect reproductive resource allocation and behavior to facilitate the evolution of social foraging in honey bees

Behavioral genomics of honeybee foraging and nest defense

The honeybee has been the most important insect species for study of social behavior. The recently released draft genomic sequence for the bee will accelerate honeybee behavioral genetics. Although we lack sufficient tools to manipulate this genome easily, quantitative trait loci (QTLs) that influence natural variation in behavior have been identified and tested for their effects on correlated behavioral traits. We review what is known about the genetics and physiology of two behavioral traits in honeybees, foraging specialization (pollen versus nectar), and defensive behavior, and present evidence that map-based cloning of genes is more feasible in the bee than in other metazoans. We also present bioinformatic analyses of candidate genes within QTL confidence intervals (CIs). The high recombination rate of the bee made it possible to narrow the search to regions containing only 17–61 predicted peptides for each QTL, although CIs covered large genetic distances. Knowledge of correlated behavioral traits, comparative bioinformatics, and expression assays facilitated evaluation of candidate genes. An overrepresentation of genes involved in ovarian development and insulin-like signaling components within pollen foraging QTL regions suggests that an ancestral reproductive gene network was co-opted during the evolution of foraging specialization. The major QTL influencing defensive/aggressive behavior contains orthologs of genes involved in central nervous system activity and neurogenesis. Candidates at the other two defensive-behavior QTLs include modulators of sensory signaling (Am5HT(7) serotonin receptor, AmArr4 arrestin, and GABA-B-R1 receptor). These studies are the first step in linking natural variation in honeybee social behavior to the identification of underlying genes

Characterisation of time-dependent mechanical behaviour of trabecular bone and its constituents

Trabecular bone is a porous composite material which consists of a mineral phase (mainly hydroxyapatite), organic phase (mostly type I collagen) and water assembled into a complex, hierarchical structure. In biomechanical modelling, its mechanical response to loads is generally assumed to be instantaneous, i.e. it is treated as a time-independent material. It is, however, recognised that the response of trabecular bone to loads is time-dependent. Study of this time-dependent behaviour is important in several contexts such as: to understand energy dissipation ability of bone; to understand the age-related non-traumatic fractures; to predict implant loosening due to cyclic loading; to understand progressive vertebral deformity; and for pre-clinical evaluation of total joint replacement. To investigate time-dependent behaviour, bovine trabecular bone samples were subjected to compressive loading, creep, unloading and recovery at multiple load levels (corresponding to apparent strain of 2,000-25,000 με). The results show that: the time-dependent behaviour of trabecular bone comprises of both recoverable and irrecoverable strains; the strain response is nonlinearly related to applied load levels; and the response is associated with bone volume fraction. It was found that bone with low porosity demonstrates elastic stiffening followed by elastic softening, while elastic softening is demonstrated by porous bone at relatively low loads. Linear, nonlinear viscoelastic and nonlinear viscoelastic-viscoplastic constitutive models were developed to predict trabecular bone’s time-dependent behaviour. Nonlinear viscoelastic constitutive model was found to predict the recovery behaviour well, while nonlinear viscoelastic-viscoplastic model predicts the full creep-recovery behaviour reasonably well. Depending on the requirements all these models can be used to incorporate time-dependent behaviour in finite element models. To evaluate the contribution of the key constituents of trabecular bone and its microstructure, tests were conducted on demineralised and deproteinised samples. Reversed cyclic loading experiments (tension to compression) were conducted on demineralised trabecular bone samples. It was found that demineralised bone exhibits asymmetric mechanical response - elastic stiffening in tension and softening in compression. This tension to compression transition was found to be smooth. Tensile multiple-load-creep-unload-recovery experiments on demineralised trabecular samples show irrecoverable strain (or residual strain) even at the low stress levels. Demineralised trabecular bone samples demonstrate elastic stiffening with increasing load levels in tension, and their time-dependent behaviour is nonlinear with respect to applied loads . Nonlinear viscoelastic constitutive model was developed which can predict its recovery behaviour well. Experiments on deproteinised samples showed that their modulus and strength are reasonably well related to bone volume fraction. The study considers an application of time-dependent behaviour of trabecular bone. Time-dependent properties are assigned to trabecular bone in a bone-screw system, in which the screw is subjected to cyclic loading. It is found that separation between bone and the screw at the interface can increase with increasing number of cycles which can accentuate loosening. The relative larger deformation occurs when this system to be loaded at the higher loading frequency. The deformation at the bone-screw interface is related to trabecular bone’s bone volume fraction; screws in a more porous bone are at a higher risk of loosening

Volatilné látky v zahraničních šumivých vínach

This thesis deals with the sensory and analytical analysis of volatile substances in foreign sparkling wines. In total there were selected and evaluated 32 samples from 13 countries of the world. Most of the samples came from the France. Theoretical overview describes the history of the production of sparkling wines and its individual steps as were created. Describe today in the world used and available methods of production and its technology. It focuses on a different styles and types of produced wines and their terroir and describes the volatile substances they contain. The experimental section contains a description of the methods of sensory and analytical analysis. The results section evaluates all wines using sensory and analytical methods. The results of the work were in addition to the verbal description transferred to the charts and graphs and compared between their parameters. These were then treated in STATISTICA software

Selection for high and low, colony weight gain in the honey bee, Apis mellifera, using selected queens and random males

Seasonal colony weight gain (honey production) in the honey bee, Apis mellifera, can be modified by selection. Two-way selection for strains of honey bees exhibiting high or low seasonal colony weight gain was made for 3 generations. In each generation, daughter queens were reared from selected colonies and permitted to mate naturally with unselected males. Two strains of honey bees were produced that differed significantly with respect to seasonal colony weight gain. These results suggest that commercial and hobbyist beekeepers can increase the efficiency of honey production by using a simple selection and mating scheme. Several variables were also evaluated as indicators of seasonal colony weight gain. Short-term colony weight gain was significantly correlated with seasonal colony weight gain and is a useful aid to selection. Early-winter colony weight, late-winter colony weight, early-spring colony weight, and winter weight loss were not correlated with seasonal colony weight gain and do not appear to be useful aids to selection

Biodemographic analysis of male honey bee mortality, Aging Cell 4

Abstract: Biodemographic studies of insects have significantly enhanced our understanding of the biology of aging. Eusocial insects have evolved to form different groups of colony members that are specialized for particular tasks and highly dependent on each other. These different groups (castes and sexes) also differ strongly in their life expectancy but relatively little is known about their mortality dynamics. In this study we present data on the age-specific flight activity and mortality of male honey bees from two different genetic lines that are exclusively dedicated to reproduction. We show that males initiating flight at a young age experience more flight events during their lifetime. No (negative) relation between the age at flight initiation and lifespan exists, as might be predicted on the basis of the antagonistic pleiotropy theory of aging. Furthermore, we fit our data to different aging models and conclude that overall a slight deceleration of the age-dependent mortality increase at advanced ages occurs. However, mortality risk increases according to the Gompertz-Makeham model when only days with flight activity (active days) are taken into account. Our interpretation of the latter is that two mortality components act on honey bee males during flight: increasing, age-dependent deaths (possibly from wear-and-tear), and age-independent deaths (possibly due to predation). The overall mortality curve is caused by the interaction of the distribution of age at foraging initiation and the mortality function during the active (flight) lifespan

Strain patterns during tensile, compressive, and shear fatigue of human cortical bone and implications for bone biomechanics

It is a common theme in basic bone biomechanics and in biomechanical applications that much of the behavior can be determined and is dictated by the level of strain, whether this pertains to bone physiology, bone remodeling, osseoinduction, osseointegration, or the development of damage. The development of damage, demonstrated by stiffness loss measurements, has already been reported in detail in the literature. However, the systematic study of the development of ‘‘plastic’’ (residual) strains, which are associated with the inelastic mechanical behavior of bone tissue, has generally been overlooked. The present study compares the rates at which the elastic (ea) and plastic components (ep) of strain developed during tensile, compressive, and shear fatigue in human cortical bone of six individuals aged between 53 and 79 years. The overall hypothesis of this investigation is that there is a common underlying factor in the damage-related behaviorof bone, which may allow us to link together the various aspects of the damage related behavior of bone. Therate of development of plastic strain (Dep/DN) and the rate of growth in elastic strain amplitude (Dea/DN) are described as a function of the stress (r), and/or stress-normalized by the modulus of elasticity (r/E). The implications of our findings are discussed with respect to simple models/mechanisms, which may underlie the observed behavior