ABSTRACT: 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
Project description:Scouts and non-scouts (recruits) were collected by using a novelty-seekingM-^] assay. Experiment was conducted in a large outdoor screened enclosure, which enabled us to exert complete control over the location and number of food resources while at the same time studying naturalistic honey bee foraging behavior. Foragers were first trained to a color-marked training feederM-^] that contained unscented 50% sucrose solution (m/v); this initially was the only food source available to them. After 2-3 days of training, a novel feederM-^] was set up in another location in the enclosure, with different color markings and an odor cue. The training feeder was maintained, providing the bees with two possible foraging locations, a familiar and a novel. Scouts were identified as bees that switched foraging from the training feeder to the novel feeder; only bees seen foraging at the novel feeder two or more times and at least once at the training feeder were collected as scouts. Non-scouts (recruits) were collected at the end of the experiments; these were bees that continued to forage at the training feeder, and were never observed to switch to the novel feeder.
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:Specific genes or encoded proteins are involved in regulating various learning models of different species through certain signaling pathways,but whether there are also regulatory genes during bimodal learning and memory is largely unknown. Using a multi-omics approach to examine gene expression changes in bees brain performed with three different learning assays, a general up-regulation of genes and proteins were observed in bimodal learning compared to controls. Protein-protein network predictions of differential proteins together with FISH assays suggest ALDH7A1 may be involved in regulation of bimodal learning and memory. Injecting siRNA-ALDH7A1 to the bee brain results in significant inhibition the expressions of ALDH7A1 and regucalcin, and increase β-alanine content. Interestingly, we found that loss of ALDH7A1 only affect visual-olfactory bimodal learning and memory, but not single visual or olfactory conditioned learning after ALDH7A1-RNAi in bees. Therefore, our data suggests that ALDH7A1 may affect bimodal learning and memory though controlling β-alanine related plasticity mechanisms.
Project description:Bees from 3 unrelated colonies were injected with 1ul PBS extract containing 10^9 genome equivalent of Deformed wIng virus (DWV) and/or fed 10µl sucrose solution containing 10^5 fresh Nosema ceranae spores. Control bees were injected and fed with an equivalent DWV- and Nosema-free extract respectively). Bees were kept in cages of 21 bees (7 from each colony), and each cage was replicated 5 times per each of the 4 treatments). Bess were kept in an incubator at 30°C/50%RH. At day 12 p.i., bees were flash frozen in liquid nitrogen, and stored at -80°C. Bee abdomen RNA was sent to Christina Grozinger lab (Penn State, USA). RNA was pooled for 3 abdomens per replicate for 5 replicates per treatment. Arrays were hybridized in a dye-swap loop design.
Project description:The drug phenobarbital induces cytochrome P450 monooxygenase (P450) gene expression in many animals, but no changes in P450 expression, or expression of any detoxification genes, were observed in worker honey bees fed on phenobarbital-candy relative to bees fed plain candy. Keywords: Expression profiling by array Five replicate microarray hybridizations were performed comparing transcript abundance in phenobarbital-fed and control-fed adult worker bees. Each total RNA pool was derived from an independent collection of 10 worker bees.
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:Expression profiling of honey bee brains exposed to brood pheromone. Exposure was performed in colonies and young (5 days-old) and old bees (15 days-old) were analyzed .
Project description:This experiment was designed to monitor the gene expression changes of young bees raised with and without queen mandibular pheromone (QMP). This experiment was a timecourse, comparing QMP- to QMP+ bees over 4 days of exposure. Bees that had eclosed over 16 hours were collected from a brood frame and placed in cages, 35 bees/cage. 8 hours later, queen mandibular pheromone (QMP, 0.1 queen equivalents) treatment was initiated, and fresh QMP was placed in the cage every day. One hour after QMP treatment had been refreshed, QMP- and QMP+ cages were collected into liquid nitrogen. 8 cages of each were collected on each day of the timecourse (1 day of treatment = Day 1, as well as Day 2, Day 3, and Day4).
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:Increasing evidence suggests microRNAs (miRNAs) control levels of mRNA expression during development of the nervous system and during sensory elicited remodelling of the brain. We used an associative olfactory learning paradigm (proboscis extension response) in the honeybee Apis mellifera to detect gene expression changes in the brain. Transcriptome analysis of bees trained to associate an odor with a reward and control bees exposed to air without reward, helped us abstract mRNA-miRNA interactions for empirical testing. Functional studies, feeding cholesterol-conjugated antisense RNA to bees resulted in the inhibition of miR-210 and of miR-932 that is embedded within the neuroligin 2 (Nlg2) gene involved in synapse development. Loss of miR-932 prevents long-term memory formation but not learning. We validated 3M-bM-^@M-^YUTR target site interactions of miR-932 and show miR-932 dysregulates actin, a key cytoskeletal molecule involved in neuronal development and activity-dependent plasticity of the brain. The analysis used Air group (no odor learning) as control sample for comparison to two groups of odor-conditioned bees: linalool and floral mix.