Project description:Experimental infection of (2 days old) adult honey bee workers (30 bees per replicates, 3 replicates per treatments, from 3 different colonies (one colony per cage for each treatment)) with 10^9 genome equivalent of Black Queen Cell Virus (BQCV) in 10µl of sugar solution and/or 10^5 fresh Nosema ceranae spores (control bees were given a similar bee extract in PBS, without pathogen). Bees were kept in cages of 30 bees in incubator (30°C/50%RH). At day 13 p.i., bees were flash frozen, and stored at -80°C. Brain mRNA profiles of 15 old bees were generated by deep sequencing, in triplicates except for bees infected by both Nosema ceranae and Black Queen Cell Virus (duplicates)
Project description:Experimental infection of (2 days old) adult honey bee workers (30 bees per replicates, 3 replicates per treatments, from 3 different colonies (one colony per cage for each treatment)) with 10^9 genome equivalent of Black Queen Cell Virus (BQCV) in 10µl of sugar solution and/or 10^5 fresh Nosema ceranae spores (control bees were given a similar bee extract in PBS, without pathogen). Bees were kept in cages of 30 bees in incubator (30°C/50%RH). At day 13 p.i., bees were flash frozen, and stored at -80°C.
2016-09-06 | GSE81664 | GEO
Project description:Complete coding genome sequence of Black queen cell virus isolated from honey bees in Italy
Project description:Experiment was designed (i) to analyse the strain composition of Deformed wing virus (DWV) populations in covertly and overtly infected honeybees (Apis mellifera) from Varroa-free and Varroa-infested colonies, and (ii) to determine abundance of the DWV strains following direct injection of the DWV preparations from covertly and overtly infected bees to the bee pupae haemolymph in the absence of Varroa destructor mites. Experiment included isolation of DWV preparations from the following bees: covertly-infected bees from Varroa-free colony, covertly infected bees exposed orally to the Varroa-selected DWV strains, and the overtly infected Varroa-exposed bees. Honeybee pupae were experimentally injected with those DWV preparations and sampled 4 days post injection following development of overt DWV infection. A series of the DWV cDNA fragment covering complete DWV genomic RNA sequences were amplified by RT-PCR using RNA extracted from virus preparations and the injected pupae. The cDNA preparations were sequenced using next generation(Illumina HighSeq 2000) paired-end sequencing to obtain data on the DWV strain composition.
Project description:Plant pollination by the western honey bee Apis mellifera is an irreplaceable agroecological and economic cornerstone currently under threat. Recent colony loss has been consistently linked to the increased prevalence of deformed wing virus (DWV), an Iflavirus transmitted from the ecoparasitic mite Varros destructor. While DWV has been detected in the honey bee brain and causally linked to behavioral impairment, the molecular impact of infection on brain gene expression is largely unknown. Recently, we discovered that two published and two new brain transcriptomic studies conducted in our lab contained DWV contamination in over 99% of sequenced honey bee samples. This unanticipated finding sharply contrasted with the experimental paradigms of these four studies, as no physical or behavioral signs of DWV were detected in any of the 335 individual honey bees sampled. We took this opportunity to perform a meta-analysis and test the hypothesis that DWV influences brain gene expression, a relationship which could be linked to the massive depopulation events observed around the world. Results from our study support commonalities in the molecular consequences of DWV in the honey bee brain and implicate specific genes and biological processes associated with infection. Next, we used single-cell RNA-Sequencing to implicate glia as active responders to viral infection. Finally, we performed viral gene expression analysis on a subset of samples and found DWV type A as well as a previously unreported A-B recombinant in the brain. We present this meta-analysis as a first step toward addressing a potential missing link between viral infection and behavior in honey bees.
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:Effects of behavioral maturation, diet quality and Queen Mandibular Pheromone on gene expression in the abdominal fat bodies of worker honey bees.
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:Expression profiling of honey bee brains exposed to queen mandibular pheromone. Exposure was performed in cages for 3 and 4 days; 3 and 4 days-old bees were analyzed