The virulent, emerging genotype B of Deformed wing virus is closely linked to overwinter honeybee worker loss

Bees are considered to be threatened globally, with severe overwinter losses of the most important commercial pollinator, the Western honeybee, a major concern in the Northern Hemisphere. Emerging infectious diseases have risen to prominence due to their temporal correlation with colony losses. Among these is Deformed wing virus (DWV), which has been frequently linked to colony mortality. We now provide evidence of a strong statistical association between overwintering colony decline in the field and the presence of DWV genotype-B (DWV-B), a genetic variant of DWV that has recently been shown to be more virulent than the original DWV genotype-A. We link the prevalence of DWV-B directly to a quantitative measure of overwinter decline (workforce mortality) of honeybee colonies in the field. We demonstrate that increased prevalence of virus infection in individual bees is associated with higher overwinter mortality. We also observed a substantial reduction of infected colonies in the spring, suggesting that virus-infected individuals had died during the winter. Our findings demonstrate that DWV-B, plus possible A/B recombinants exhibiting DWV-B at PCR primer binding sites, may be a major cause of elevated overwinter honeybee loss. Its potential emergence in naïve populations of bees may have far-reaching ecological and economic impacts

Elevated virulence of an emerging viral genotype as a driver of honeybee loss.

PublishedJournal ArticleEmerging infectious diseases (EIDs) have contributed significantly to the current biodiversity crisis, leading to widespread epidemics and population loss. Owing to genetic variation in pathogen virulence, a complete understanding of species decline requires the accurate identification and characterization of EIDs. We explore this issue in the Western honeybee, where increasing mortality of populations in the Northern Hemisphere has caused major concern. Specifically, we investigate the importance of genetic identity of the main suspect in mortality, deformed wing virus (DWV), in driving honeybee loss. Using laboratory experiments and a systematic field survey, we demonstrate that an emerging DWV genotype (DWV-B) is more virulent than the established DWV genotype (DWV-A) and is widespread in the landscape. Furthermore, we show in a simple model that colonies infected with DWV-B collapse sooner than colonies infected with DWV-A. We also identify potential for rapid DWV evolution by revealing extensive genome-wide recombination in vivo The emergence of DWV-B in naive honeybee populations, including via recombination with DWV-A, could be of significant ecological and economic importance. Our findings emphasize that knowledge of pathogen genetic identity and diversity is critical to understanding drivers of species decline.This work was supported by the Federal Ministry of Food, Agriculture and Consumer Protection (Germany): Fit Bee project (grant 511-06.01-28-1-71.007-10), the EU: BeeDoc (grant 244956), iDiv (2013 NGS-Fast Track grant W47004118) and the Insect Pollinators Initiative (IPI grant BB/I000100/1 and BB/I000151/1). The IPI is funded jointly by the Biotechnology and Biological Sciences Research Council, the Department for Environment, Food and Rural Affairs, the Natural Environment Research Council, the Scottish Government and the Wellcome Trust, under the Living with Environmental Change Partnership

Standard methods for Nosema research

Methods are described for working with Nosema apis and Nosema ceranae in the field and in the laboratory. For fieldwork, different sampling methods are described to determine colony level infections at a given point in time, but also for following the temporal infection dynamics. Suggestions are made for how to standardise field trials for evaluating treatments and disease impact. The laboratory methods described include different means for determining colony level and individual bee infection levels and methods for species determination, including light microscopy, electron microscopy, and molecular methods (PCR). Suggestions are made for how to standardise cage trials, and different inoculation methods for infecting bees are described, including control methods for spore viability. A cell culture system for in vitro rearing of Nosema spp. is described. Finally, how to conduct different types of experiments are described, including infectious dose, dose effects, course of infection and longevity test

Unity in defence: honeybee workers exhibit conserved molecular responses to diverse pathogens

This is the final version of the article. Available from the publisher via the DOI in this record.Background: Organisms typically face infection by diverse pathogens, and hosts are thought to have developed specific responses to each type of pathogen they encounter. The advent of transcriptomics now makes it possible to test this hypothesis and compare host gene expression responses to multiple pathogens at a genome-wide scale. Here, we performed a meta-analysis of multiple published and new transcriptomes using a newly developed bioinformatics approach that filters genes based on their expression profile across datasets. Thereby, we identified common and unique molecular responses of a model host species, the honey bee (Apis mellifera), to its major pathogens and parasites: the Microsporidia Nosema apis and Nosema ceranae, RNA viruses, and the ectoparasitic mite Varroa destructor, which transmits viruses. Results: We identified a common suite of genes and conserved molecular pathways that respond to all investigated pathogens, a result that suggests a commonality in response mechanisms to diverse pathogens. We found that genes differentially expressed after infection exhibit a higher evolutionary rate than non-differentially expressed genes. Using our new bioinformatics approach, we unveiled additional pathogen-specific responses of honey bees; we found that apoptosis appeared to be an important response following microsporidian infection, while genes from the immune signalling pathways, Toll and Imd, were differentially expressed after Varroa/virus infection. Finally, we applied our bioinformatics approach and generated a gene co-expression network to identify highly connected (hub) genes that may represent important mediators and regulators of anti-pathogen responses. Conclusions: Our meta-analysis generated a comprehensive overview of the host metabolic and other biological processes that mediate interactions between insects and their pathogens. We identified key host genes and pathways that respond to phylogenetically diverse pathogens, representing an important source for future functional studies as well as offering new routes to identify or generate pathogen resilient honey bee stocks. The statistical and bioinformatics approaches that were developed for this study are broadly applicable to synthesize information across transcriptomic datasets. These approaches will likely have utility in addressing a variety of biological questions.This article is a joint effort of the working group TRANSBEE and an outcome of two workshops kindly supported by sDiv, the Synthesis Centre for Biodiversity Sciences within the German Centre for Integrative Biodiversity Research (iDiv) Halle-Jena-Leipzig, funded by the German Science Foundation (FZT 118). New datasets were performed thanks to the Insect Pollinators Initiative (IPI grant BB/I000100/1 and BB/I000151/1), with participation of the UK-USA exchange funded by the BBSRC BB/I025220/1 (datasets #4, 11 and 14). The IPI is funded jointly by the Biotechnology and Biological Sciences Research Council, the Department for Environment, Food and Rural Affairs, the Natural Environment Research Council, the Scottish Government and the Wellcome Trust, under the Living with Environmental Change Partnershi

Water temperature, not fish morph, determines parasite infections of sympatric Icelandic threespine sticklebacks (Gasterosteus aculeatus)

Peer reviewe

Impact of asynchronous emergence of two lethal pathogens on amphibian assemblages

Emerging diseases have been increasingly associated with population declines, with co-infections exhibiting many types of interactions. The chytrid fungus (Batrachochytrium dendrobatidis) and ranaviruses have extraordinarily broad host ranges, however co-infection dynamics have been largely overlooked. We investigated the pattern of co-occurrence of these two pathogens in an amphibian assemblage in Serra da Estrela (Portugal). The detection of chytridiomycosis in Portugal was linked to population declines of midwife-toads (Alytes obstetricans). The asynchronous and subsequent emergence of a second pathogen - ranavirus - caused episodes of lethal ranavirosis. Chytrid effects were limited to high altitudes and a single host, while ranavirus was highly pathogenic across multiple hosts, life-stages and altitudinal range. This new strain (Portuguese newt and toad ranavirus – member of the CMTV clade) caused annual mass die-offs, similar in host range and rapidity of declines to other locations in Iberia affected by CMTV-like ranaviruses. However, ranavirus was not always associated with disease, mortality and declines, contrasting with previous reports on Iberian CMTV-like ranavirosis. We found little evidence that pre-existing chytrid emergence was associated with ranavirus and the emergence of ranavirosis. Despite the lack of cumulative or amplified effects, ranavirus drove declines of host assemblages and changed host community composition and structure, posing a grave threat to all amphibian populations

Divergent Macroparasite Infections in Parapatric Swiss Lake-Stream Pairs of Threespine Stickleback (Gasterosteus aculeatus).

Spatial heterogeneity in diversity and intensity of parasitism is a typical feature of most host-parasite interactions, but understanding of the evolutionary implications of such variation is limited. One possible outcome of infection heterogeneities is parasite-mediated divergent selection between host populations, ecotypes or species which may facilitate the process of ecological speciation. However, very few studies have described infections in population-pairs along the speciation continuum from low to moderate or high degree of genetic differentiation that would address the possibility of parasite-mediated divergent selection in the early stages of the speciation process. Here we provide an example of divergent parasitism in freshwater fish ecotypes by examining macroparasite infections in threespine stickleback (Gasterosteus aculeatus) of four Swiss lake systems each harbouring parapatric lake-stream ecotype pairs. We demonstrate significant differences in infections within and between the pairs that are driven particularly by the parasite taxa transmitted to fish from benthic invertebrates. The magnitude of the differences tended to correlate positively with the extent of neutral genetic differentiation between the parapatric lake and stream populations of stickleback, whereas no such correlation was found among allopatric populations from similar or contrasting habitats. This suggests that genetic differentiation is unrelated to the magnitude of parasite infection contrasts when gene flow is constrained by geographical barriers while in the absence of physical barriers, genetic differentiation and the magnitude of differences in infections tend to be positively correlated

Parasites and parallel divergence of individual MHC allelic richness in Icelandic threespine stickleback (Gasterosteus aculeatus L.): Contrasting habitats and population divergence

When individuals of a single species encounter different environments, local adaptation and thus phenotypic diversification among populations is expected. This intraspecific diversity, which is common among northern freshwater fish, can be observed even between populations that live in close proximity (sympatry). Common ecological selection pressures often lead to parallel evolution as similar habitats favour the same phenotypic optima resulting in independent evolution of the same traits towards the same direction. Local adaptation is reflected in the external morphology of the fish as many morphological traits are associated with performance and fitness i.e. locomotion activity, foraging efficiency and protection against predators. However, in environments characterized by great niche availability utilization of vacant niches can result in increased intrapopulation phenotypic variation. Differential exploitation and specialization to new alternative resources can also occur between sexes of the same population. This intersexual competition for resources can lead to sexual dimorphism and its extent has been found to increase with niche availability as sexes have the potential to explore a wider range of alternative niches. An important ecological factor that can contribute to divergence among populations is the interaction between hosts and parasites. According to theory, differential parasitism between host populations has the potential to promote adaptation to local environments and thus rapid divergence due to host-parasite co-evolutionary interactions. Parasite mediated selection has been found to operate on the highly polymorphic genes of the major histocompatibility complex (MHC) which is part of the adaptive immune system of the jawed vertebrates. In the first part of the thesis (manuscript I) threespine sticklebacks (Gasterosteus aculeatus) sampled from four lakes located in South-Western Iceland, two of which are inhabited by two distinct stickleback morphs, apparently adapted to different benthic habitats, -lava and mud-, and two single morph lakes, were screened for parasites and genotyped for MHC class IIB diversity. The level of parasitic infection was found to differ consistently between the sympatric morphs but also between populations found in similar foraging habitats with fish from the lava/rocky habitats being more heavily infected. A parallel pattern was also found in individual MHC allelic variation with lava stickleback morphs exhibiting lower levels of variation compared to the mud morphs. The parallel divergence in MHC allele number between the sympatric morphs may be caused by different selection pressures imposed by varying abundance of parasite species causing different optima to be favoured in contrasting habitats. Consequently, parasite mediated selection could play a part in the divergence of the benthic lava-mud morphs in Icelandic lakes. The second part of the thesis (manuscript II) investigates sexual dimorphism and morphological variation in relation to substrate type and lake size among populations and the sexes in threespine stickleback (Gasterosteus aculeatus). Threespine stickleback were sampled from 34 sites in 22 lakes in Iceland inhabiting three main substrate types: mud, vegetation and rock. Lake size, generally representing increased niche availability, was positively correlated with phenotypic variance within populations and sexual dimorphism, most likely representing increased ecological opportunity. Finally, our result do not support that increased sexual dimorphism restricts phenotypic variation within each sex at the population level