Varroa mites and viruses are the currently the high-profile suspects in collapsing bee colonies. Therefore, seasonal variation in varroa load and viruses (Acute-Kashmir-Israeli complex (AKI) and Deformed Wing Virus (DWV)) were monitored in a year-long study. We investigated the viral titres in honey bees and varroa mites from 23 colonies (15 apiaries) under three treatment conditions: Organic acids (11 colonies), pyrethroid (9 colonies) and untreated (3 colonies). Approximately 200 bees were sampled every month from April 2011 to October 2011, and April 2012. The 200 bees were split to 10 subsamples of 20 bees and analysed separately, which allows us to determine the prevalence of virus-infected bees. The treatment efficacy was often low for both treatments. In colonies where varroa treatment reduced the mite load, colonies overwintered successfully, allowing the mites and viruses to be carried over with the bees into the next season. In general, AKI and DWV titres did not show any notable response to the treatment and steadily increased over the season from April to October. In the untreated control group, titres increased most dramatically. Viral copies were correlated to number of varroa mites. Most colonies that collapsed over the winter had significantly higher AKI and DWV titres in October compared to survivors. Only treated colonies survived the winter. We discuss our results in relation to the varroa-virus model developed by Stephen Martin.

Aquaglyceroporins form the subset of the aquaporin water channel family that is permeable to glycerol and certain small, uncharged solutes. AQP9 has unusually broad solute permeability and is expressed in hepatocyte plasma membranes. Proteoliposomes reconstituted with expressed, purified rat AQP9 protein were compared with simple liposomes for solute permeability. At pH 7.5, AQP9 proteoliposomes exhibited Hg 2؉ -inhibitible glycerol and urea permeabilities that were increased 63-fold and 90-fold over background. ␤-Hydroxybutyrate permeability was not increased above background, and osmotic water permeability was only minimally elevated. During starvation, the liver takes up glycerol for gluconeogenesis. Expression of AQP9 in liver was induced up to 20-fold in rats fasted for 24 -96 h, and the AQP9 level gradually declined after refeeding. No changes in liver AQP9 levels were observed in rats fed ketogenic diets or high-protein diets, but AQP9 levels were elevated in livers of rats made diabetic by streptozotocin injection. When blood glucose levels of the diabetic rats were restored to normal by insulin treatments, the AQP9 levels returned to baseline. Confocal immunofluorescence revealed AQP9 immunostaining on the sinusoidal surfaces of hepatocyte plates throughout the livers of control rats. Denser immunostaining was observed in the same distribution in livers of fasted and streptozotocin-treated rats. We conclude that AQP9 serves as membrane channel in hepatocytes for glycerol and urea at physiological pH, but not for ␤-hydroxybutyrate. In addition, levels of AQP9 expression fluctuate depending on the nutritional status of the subject and the circulating insulin levels. T he AQP9 cDNA was first isolated during efforts to clone urea transporters by expression in oocytes (1). Expressed in testes, leukocytes, and brain, AQP9 is abundant in liver (1) where it resides in hepatocyte plasma membranes facing the sinusoids (2-4). The coding sequence of AQP9 is more closely related to AQP3 (5) and AQP7 (6), which are both permeated by glycerol and water. This subset of proteins, referred to as aquaglyceroporins, is functionally distinct from the water-selective homologs AQP1, AQP2, AQP4, and AQP5 (7). The original studies of Xenopus laevis oocytes expressing rat AQP9 reported permeability to a wide range of 14 C-or 3 H-labeled solutes including polyols, carbamides, purines, pyrimidines, nucleosides, and monocarboxylates (1). Glycerol and urea permeability have been confirmed with AQP9 oocytes (8), but studies of proteoliposomes reconstituted with purified AQP9 protein have not been reported.The physiological functions of AQP9 are uncertain. During prolonged fasting, glycerol released from adipocytes via AQP7 may be taken up by the liver via AQP9 for gluconeogenesis. Urea, a byproduct of amino acid deamination, and ␤-hydroxybutyrate, an alternative fuel, may be released from liver via AQP9. An elegant series of recent studies of adipocyte AQP7 and liver AQP9 mRNAs and promoters suggested that the genes are coordinately ...

-Samples of adult honey bees from apiaries with unusually high winter mortality and brood from hives with symptoms of disease were tested for presence of acute bee paralysis virus (ABPV), black queen cell virus (BQCV), chronic bee paralysis virus (CBPV), deformed wing virus (DWV), Kashmir bee virus (KBV) and sacbrood virus (SBV) by RT-PCR. All six viruses were detected, but the frequencies varied significantly: SBV was detected in 78 apiaries, DWV in 55, ABPV in 11, CBPV in 4, BQCV in 1 and KBV in 1. This is the first record of KBV in Denmark. A large majority of the bee samples were infected with one or more viruses. Single, dual and triple infections were observed. Nucleotide sequences of the PCR products from each virus were determined and found to be 98-99% identical to GenBank accessions except CBPV, which was only 88-90% identical to known CBPV sequences.Kashmir bee virus / acute bee paralysis virus / black queen cell virus / chronic bee paralysis virus / deformed wing virus / sacbrood virus / multiple virus infection

BackgroundNatural products are an important source of drugs and other commercially interesting compounds, however their isolation and production is often difficult. Metabolic engineering, mainly in bacteria and yeast, has sought to circumvent some of the associated problems but also this approach is impeded by technical limitations. Here we describe a novel strategy for production of diverse natural products, comprising the expression of an unprecedented large number of biosynthetic genes in a heterologous host.ResultsAs an example, genes from different sources, representing enzymes of a seven step flavonoid pathway, were individually cloned into yeast expression cassettes, which were then randomly combined on Yeast Artificial Chromosomes and used, in a single transformation of yeast, to create a variety of flavonoid producing pathways. Randomly picked clones were analysed, and approximately half of them showed production of the flavanone naringenin, and a third of them produced the flavonol kaempferol in various amounts. This reflected the assembly of 5–7 step multi-species pathways converting the yeast metabolites phenylalanine and/or tyrosine into flavonoids, normally only produced by plants. Other flavonoids were also produced that were either direct intermediates or derivatives thereof. Feeding natural and unnatural, halogenated precursors to these recombinant clones demonstrated the potential to further diversify the type of molecules that can be produced with this technology.ConclusionThe technology has many potential uses but is particularly suited for generating high numbers of structurally diverse compounds, some of which may not be amenable to chemical synthesis, thus greatly facilitating access to a huge chemical space in the search for new commercially interesting compounds

The well-being of a colony and replenishment of the workers depends on a healthy queen. Diseases in queens are seldom reported, and our knowledge on viral infection in queens is limited. In this study, 86 honey bee queens were collected from beekeepers in Denmark. All queens were tested separately by two real-time PCRs: one for the presence of deformed wing virus (DWV), and one that would detect sequences of acute bee-paralysis virus, Kashmir bee virus and Israeli acute paralysis virus (AKI complex). Worker bees accompanying the queen were also analysed. The queens could be divided into three groups based on the level of infection in their head, thorax, ovary, intestines and spermatheca. Four queens exhibited egg-laying deficiency, but visually all queens appeared healthy. Viral infection was generally at a low level in terms of AKI copy numbers, with 134/430 tissues (31 %) showing the presence of viral infection ranging from 101 to 105 copies. For DWV, 361/340 tissues (84 %) showed presence of viral infection (DWV copies ranging from 102 to 1012), with 50 tissues showing viral titres >107 copies. For both AKI and DWV, the thorax was the most frequently infected tissue and the ovaries were the least frequently infected. Relative to total mass, the spermatheca showed significantly higher DWV titres than the other tissues. The ovaries had the lowest titre of DWV. No significant differences were found among tissues for AKI. A subsample of 14 queens yielded positive results for the presence of negative-sense RNA strands, thus demonstrating active virus replication in all tissues.