Project description:Flenniken - Honey bee gene expression microarray experimental design<br>To minimize variability between samples all arrayed bees were obtained from a single brood comb from a naturally mated queen, therefore all the bees were age-matched half-sisters. The bees selected for microarray analysis of virus (Sindbis-eGFP) co-injected with either virus-specific-dsRNA (vs-dsRNA) or non-specific dsRNA (ns-dsRNA) exhibited the reduced virus phenotype that was seen in the majority of the bees assayed. The five representative bees from each condition (v, v+vs-dsRNA, v+ns-dsRNA, dsRNA, and mock/injected with buffer) selected for microarray analysis were free of pre-existing conditions (assessed by APM analysis) (Runckel, Flenniken et al., 2011). To facilitate gene expression comparisons between multiple treatment groups we utilized a reference-design strategy in which each Cy5-labeled experimental sample was hybridized with a standardized Cy3-labeled reference sample. A complex RNA mixture representing hundreds of bees of various ages exposed to difference treatment groups, served as the reference RNA sample.
Project description:We used microarrays to monitor expression patterns of several thousand genes in the brains of same-aged (10 day old) virgin queens, sterile workers, and reproductive workers in honey bees (Apis mellifera).
Project description:BACKGROUND: Social insects, such as honey bees, use molecular, physiological and behavioral responses to combat pathogens and parasites. The honey bee genome contains all of the canonical insect immune response pathways, and several studies have demonstrated that pathogens can activate expression of immune effectors. Honey bees also use behavioral responses, termed social immunity, to collectively defend their hives from pathogens and parasites. These responses include hygienic behavior (where workers remove diseased brood) and allo-grooming (where workers remove ectoparasites from nestmates). We have previously demonstrated that immunostimulation causes changes in the cuticular hydrocarbon profiles of workers, which results in altered worker-worker social interactions. Thus, cuticular hydrocarbons may enable workers to identify sick nestmates, and adjust their behavior in response. Here, we test the specificity of behavioral, chemical and genomic responses to immunostimulation by challenging workers with a panel of different immune stimulants (saline, Sephadex beads and Gram-negative bacteria E. coli). RESULTS: While only bacteria-injected bees elicited altered behavioral responses from healthy nestmates compared to controls, all treatments resulted in significant changes in cuticular hydrocarbon profiles. Immunostimulation caused significant changes in expression of hundreds of genes, the majority of which have not been identified as members of the canonical immune response pathways. Furthermore, several new candidate genes that may play a role in cuticular hydrocarbon biosynthesis were identified. Finally, we identified common genes regulated by pathogen challenge in honey bees and other insects, suggesting that immune responses are conserved at the molecular level. CONCLUSIONS: These studies suggest that honey bee genomic responses to immunostimulation are substantially broader than expected, and may mediate the behavioral changes associated with social immunity by orchestrating changes in chemical signaling.
Project description:Background: Honey bee is a major insect used for pollination of a number of commercial crops worldwide. However, the number of managed honey bee colonies has recently declined in several countries, and a number of possible causes are proposed. Although the use of honey bees for pollination can be considered as disruption of the habitat, its effects on honey bees' physiology have never been addressed. In Japan, more than 100 thousands colonies are annually used for pollination, and intriguingly 80% of them are used in greenhouses. Recently, honey bee colonies have often collapsed when they are introduced into greenhouses. Thus, to suppress colony collapses and maintain the number of worker bees in the colonies are essential for successful long-term pollination in greenhouses and recycling honey bee colonies. Honey bee hives were installed into strawberry and eggplant greenhouses, and then the gene expression profiles of the honey bees were examined at the different time periods. Total 16 samples with two replicates were analyzed.
Project description:The aim of this work was to extend the previous gene expression analysis ofM- M- nurse and forager gene expression, but using the Apis oligo-array designed from the recently sequenced genome.M- M- This study included 28000 features which included ~11000 predicted genes.M- M- The intial study was done using a microarray design based on PCR products of 5500 features derived from an expressed sequence tag analysis of a brain cDNA library.M- M- The variables studied were age (young and old), cast (nurse and forager) and differential gene expression.
Project description:As a matter of fact, honeybees are vital for the pollination of more than 80 crops of agricultural interest. However, population decline has become an important global issue causing significant concerns among agricultural experts and the broader public. For this, parasites are known to be the major culprits responsible for the losses of millions of honeybee colonies so far. Among these parasites, Varroa destructor has been identified as a major cause for global losses in Western honeybee (Apis mellifera) colonies. Hygienic behavior (HB), on the other hand, is a collective response by adult honeybees to defend against parasites and diseases that is known to involve in resistance towards Varroosis. Even with the efforts made to elucidate the molecular mechanism underlying HB, it is still not understood. In our study, we have studied the proteomic correlates to HB using a honeybee line (selected for Varroa-specific HB for over a decade in Germany). We sampled individual worker bees from this line that showed HB after closer infrared video observations and compared the proteomes of their mushroom bodies and antennae with those of workers that came from the same set of colonies but didn't show the behaviour. Furthermore, we compared the pupal hemolymph for worker bees of the selected HB line and a control line using state-of-the art techniques of proteomics. We identified a total of 8609 proteins (covered >55% of the honeybee proteome) from these three honeybee tissues. This is the most comprehensive proteomic study of the honeybee HB to date, and the first to focus on individual bees expressing Varroa-specific HB. These results have significantly advanced our knowledge on the biology underlining HB to a new level. The uniquely found functional classes and pathways by the proteins identified in each tissue suggest that hygienic bees have shaped distinct proteome settings to underpin the HB. Moreover, during analysis of pupal hemolymph proteome, the HB-line has adapted a unique strategy to boost an individual and social immunity and drove pupal organogenesis via energy metabolism and protein biosynthesis. Moreover, in the mushroom bodies of different HB phenotypic worker bees, the hygienic bees have enhanced their neuronal sensitivity to promote the execution of HB by activation of synaptic vesicles and calcium channel activities. Moreover, in the antennae of two HB phenotypic worker bees, the hygienic bees have demonstrated strengthening of their sensitivity associated with olfactory senses and signal transmissions, which is important to input a strong signal to the mushroom bodies and initiate HB. In conclusion, our novel findings have significantly extended our understandings of the molecular mechanisms that underline the HB to combat Varroa infestation. Furthermore, we identified a wide array of novel markers that are useful for accelerating marker associated selection of HB to aid in the natural resistance to a parasite blamed for a global decline in honeybee health.
Project description:Bees were trained to visit food sources at different times of day. By collecting bees just before the training times, we were able to analyze the genes that are associated with the time of day, the time of training, the activity state, or genes associated with unique spatiotemporal memories