Recombinants between Deformed wing virus and Varroa destructor virus-1 may prevail in Varroa destructor-infested honeybee colonies

We have used high-throughput Illumina sequencing to identify novel recombinants between deformed wing virus (DWV) and Varroa destructor virus-1 (VDV-1), which accumulate to higher levels than DWV in both honeybees and Varroa destructor mites. The recombinants, VDV-1VVD and VDV-1DVD, exhibit crossovers between the 5’-untranslated region (5’-UTR), and/or the regions encoding the structural (capsid) and non-structural viral proteins. This implies the genomes are modular and that each region may evolve independently, as demonstrated in human enteroviruses. Individual honeybee pupae were infected with a mixture of observed recombinants and DWV. The strong correlation between VDV-1DVD levels in honeybee pupae and the associated mites was observed, suggesting that this recombinant, with a DWV-derived 5’-UTR and non-structural protein region flanking VDV- 1-derived capsid encoding region, is better adapted to transmission between V. destructor and honeybees than the parental DWV or a recombinant bearing the VDV-1-derived 5’-UTR (VDV-1VVD)

極域の雪氷アルベド変更実験

第6回極域科学シンポジウム分野横断セッション：[IA] 急変する北極気候システム及びその全球的な影響の総合的解明―GRENE北極気候変動研究事業研究成果報告2015―11月19日（木）　国立極地研究所１階交流アトリウ

In Situ Hybridization Analysis of the Expression of Futsch, Tau, and MESK2 Homologues in the Brain of the European Honeybee (Apis mellifera L.)

BACKGROUND: The importance of visual sense in Hymenopteran social behavior is suggested by the existence of a Hymenopteran insect-specific neural circuit related to visual processing and the fact that worker honeybee brain changes morphologically according to its foraging experience. To analyze molecular and neural bases that underlie the visual abilities of the honeybees, we used a cDNA microarray to search for gene(s) expressed in a neural cell-type preferential manner in a visual center of the honeybee brain, the optic lobes (OLs). METHODOLOGY/PRINCIPAL FINDINGS: Expression analysis of candidate genes using in situ hybridization revealed two genes expressed in a neural cell-type preferential manner in the OLs. One is a homologue of Drosophila futsch, which encodes a microtubule-associated protein and is preferentially expressed in the monopolar cells in the lamina of the OLs. The gene for another microtubule-associated protein, tau, which functionally overlaps with futsch, was also preferentially expressed in the monopolar cells, strongly suggesting the functional importance of these two microtubule-associated proteins in monopolar cells. The other gene encoded a homologue of Misexpression Suppressor of Dominant-negative Kinase Suppressor of Ras 2 (MESK2), which might activate Ras/MAPK-signaling in Drosophila. MESK2 was expressed preferentially in a subclass of neurons located in the ventral region between the lamina and medulla neuropil in the OLs, suggesting that this subclass is a novel OL neuron type characterized by MESK2-expression. These three genes exhibited similar expression patterns in the worker, drone, and queen brains, suggesting that they function similarly irrespective of the honeybee sex or caste. CONCLUSIONS: Here we identified genes that are expressed in a monopolar cell (Amfutsch and Amtau) or ventral medulla-preferential manner (AmMESK2) in insect OLs. These genes may aid in visualizing neurites of monopolar cells and ventral medulla cells, as well as in analyzing the function of these neurons

Occurrence, function and evolutionary origins of ‘2A-like’ sequences in virus genomes

2A is an oligopeptide sequence mediating a ribosome ‘skipping’ effect, producing an apparent ‘cleavage’ of polyproteins. First identified and characterized in picornaviruses, ‘2A-like’ sequences are found in other mammalian viruses and a wide range of insect viruses. Databases were analysed using a motif conserved amongst 2A/2A-like sequences. The newly identified 2A-like sequences (30 aa) were inserted into a reporter polyprotein to determine their cleavage activity. Our analyses showed that these sequences fall into two categories. The majority mediated very high (complete) cleavage to separate proteins and a few sequences mediated cleavage with lower efficiency, generating appreciable levels of the uncleaved form. Phylogenetic analyses of 2A-like sequences and RNA-dependent RNA polymerases (RdRps) indicated multiple, independent, acquisitions of these sequences at different stages during virus evolution. Within a virus family, 2A sequences are (probably) homologous, but diverge due to other evolutionary pressures. Amongst different families, however, 2A/2A-like sequences appear to be homoplasic

Carbohydrate metabolism genes and pathways in insects: insights from the honey bee genome

Carbohydrate-metabolizing enzymes may have particularly interesting roles in the honey bee, Apis mellifera, because this social insect has an extremely carbohydrate-rich diet, and nutrition plays important roles in caste determination and socially mediated behavioural plasticity. We annotated a total of 174 genes encoding carbohydrate-metabolizing enzymes and 28 genes encoding lipid-metabolizing enzymes, based on orthology to their counterparts in the fly, Drosophila melanogaster, and the mosquito, Anopheles gambiae. We found that the number of genes for carbohydrate metabolism appears to be more evolutionarily labile than for lipid metabolism. In particular, we identified striking changes in gene number or genomic organization for genes encoding glycolytic enzymes, cellulase, glucose oxidase and glucose dehydrogenases, glucose-methanol-choline (GMC) oxidoreductases, fucosyltransferases, and lysozymes

Viruses Associated with Ovarian Degeneration in Apis mellifera L. Queens

Queen fecundity is a critical issue for the health of honeybee (Apis mellifera L.) colonies, as she is the only reproductive female in the colony and responsible for the constant renewal of the worker bee population. Any factor affecting the queen's fecundity will stagnate colony development, increasing its susceptibility to opportunistic pathogens. We discovered a pathology affecting the ovaries, characterized by a yellow discoloration concentrated in the apex of the ovaries resulting from degenerative lesions in the follicles. In extreme cases, marked by intense discoloration, the majority of the ovarioles were affected and these cases were universally associated with egg-laying deficiencies in the queens. Microscopic examination of the degenerated follicles showed extensive paracrystal lattices of 30 nm icosahedral viral particles. A cDNA library from degenerated ovaries contained a high frequency of deformed wing virus (DWV) and Varroa destructor virus 1 (VDV-1) sequences, two common and closely related honeybee Iflaviruses. These could also be identified by in situ hybridization in various parts of the ovary. A large-scale survey for 10 distinct honeybee viruses showed that DWV and VDV-1 were by far the most prevalent honeybee viruses in queen populations, with distinctly higher prevalence in mated queens (100% and 67%, respectively for DWV and VDV-1) than in virgin queens (37% and 0%, respectively). Since very high viral titres could be recorded in the ovaries and abdomens of both functional and deficient queens, no significant correlation could be made between viral titre and ovarian degeneration or egg-laying deficiency among the wider population of queens. Although our data suggest that DWV and VDV-1 have a role in extreme cases of ovarian degeneration, infection of the ovaries by these viruses does not necessarily result in ovarian degeneration, even at high titres, and additional factors are likely to be involved in this pathology

Superinfection exclusion and the long-term survival of honey bees in Varroa-infested colonies

Over the past 50 years, many millions of European honey bee (Apis mellifera) colonies have died as the ectoparasitic mite, Varroa destructor, has spread around the world. Subsequent studies have indicated that the mite's association with a group of RNA viral pathogens (Deformed Wing Virus, DWV) correlates with colony death. Here, we propose a phenomenon known as superinfection exclusion that provides an explanation of how certain A. mellifera populations have survived, despite Varroa infestation and high DWV loads. Next-generation sequencing has shown that a non-lethal DWV variant 'type B' has become established in these colonies and that the lethal 'type A' DWV variant fails to persist in the bee population. We propose that this novel stable host-pathogen relationship prevents the accumulation of lethal variants, suggesting that this interaction could be exploited for the development of an effective treatment that minimises colony losses in the future.The ISME Journal advance online publication, 27 October 2015; doi:10.1038/ismej.2015.186

Diversity in a honey bee pathogen: first report of a third master variant of the Deformed Wing Virus quasispecies

Treatment of emerging RNA viruses is hampered by the high mutation and replication rates that enable these viruses to operate as a quasispecies. Declining honey bee populations have been attributed to the ectoparasitic mite Varroa destructor and its affiliation with Deformed Wing Virus (DWV). In the current study we use next-generation sequencing to investigate the DWV quasispecies in an apiary known to suffer from overwintering colony losses. We show that the DWV species complex is made up of three master variants. Our results indicate that a new DWV Type C variant is distinct from the previously described types A and B, but together they form a distinct clade compared with other members of the Iflaviridae. The molecular clock estimation predicts that Type C diverged from the other variants ~319 years ago. The discovery of a new master variant of DWV has important implications for the positive identification of the true pathogen within global honey bee populations